399 results on '"Inhwan, Hwang"'
Search Results
102. Interactions between Transmembrane Helices within Monomers of the Aquaporin AtPIP2;1 Play a Crucial Role in Tetramer Formation
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Jouhyun Jeon, Yongjik Lee, Juhun Lee, Dong-Wook Lee, Inhwan Hwang, Jin-Kwan Han, Hyun-Kyung Lee, Myoung Hui Lee, Wonhee Han, Yun-Joo Yoo, Jin Seok Kim, Dae Heon Kim, Yunje Cho, and Junho Lee
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0106 biological sciences ,0301 basic medicine ,Mutant ,Arabidopsis ,Aquaporin ,Plant Science ,Biology ,Aquaporins ,Endoplasmic Reticulum ,01 natural sciences ,Protein Structure, Secondary ,Cell membrane ,03 medical and health sciences ,Tetramer ,medicine ,Molecular Biology ,Alanine ,Arabidopsis Proteins ,Endoplasmic reticulum ,Cell Membrane ,Transmembrane domain ,030104 developmental biology ,medicine.anatomical_structure ,Biochemistry ,Biophysics ,Protein Multimerization ,Biogenesis ,010606 plant biology & botany - Abstract
Aquaporin (AQP) is a water channel protein found in various subcellular membranes of both prokaryotic and eukaryotic cells. The physiological functions of AQPs have been elucidated in many organisms. However, understanding their biogenesis remains elusive, particularly regarding how they assemble into tetramers. Here, we investigated the amino acid residues involved in the tetramer formation of the Arabidopsis plasma membrane AQP AtPIP2;1 using extensive amino acid substitution mutagenesis. The mutant proteins V41A/E44A, F51A/L52A, F87A/I91A, F92A/I93A, V95A/Y96A, and H216A/L217A, harboring alanine substitutions in the transmembrane (TM) helices of AtPIP2;1 polymerized into multiple oligomeric complexes with a variable number of subunits greater than four. Moreover, these mutant proteins failed to traffic to the plasma membrane, instead of accumulating in the endoplasmic reticulum (ER). Structure-based modeling revealed that these residues are largely involved in interactions between TM helices within monomers. These results suggest that inter-TM interactions occurring both within and between monomers play crucial roles in tetramer formation in the AtPIP2;1 complex. Moreover, the assembly of AtPIP2;1 tetramers is critical for their trafficking from the ER to the plasma membrane, as well as water permeability.
- Published
- 2016
103. Efficacy and safety of mFOLFIRINOX in patients with borderline resectable and locally advanced unresectable pancreatic cancer: Intention-to-treat population analysis
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Jae Ho Byun, Kyu-Pyo Kim, Inhwan Hwang, Heung-Moon Chang, Changhoon Yoo, Baek-Yeol Ryoo, and Jae Ho Jeong
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Oncology ,Unresectable Pancreatic Cancer ,Cancer Research ,medicine.medical_specialty ,education.field_of_study ,Disease entity ,Intention-to-treat analysis ,business.industry ,Population ,Locally advanced ,medicine.disease ,Borderline resectable ,Internal medicine ,Pancreatic cancer ,medicine ,In patient ,education ,business - Abstract
720 Background: Borderline resectable pancreatic cancer (BRPC) and locally advanced unresectable pancreatic cancer (LAUPC) are heterogeneous disease entity with various prognosis. Based on the phase III PRODIGE trial, (m)FOLFIRINOX has been widely used for the management of patients with BRPC and LAUPC. Considering the lack of large phase 3 trial of (m)FOLFIRINOX for BRPC and LAUPC, real-life evidences of (m)FOLFIRINOX are needed. Methods: In this retrospective analysis, 199 patients who received at least one dose of (m)FOLFIRINOX between February 2013 and January 2017 were included. Endpoints of this study were objective response rates (ORR), surgical resection rate, progression-free survival (PFS) and overall survival (OS). Results: Median age was 60 years (range, 33-79) and 62.3% of patients were male. Pancreas head (n=112, 56.3%) was the most common primary tumor site, followed by body (n=42, 21.1%) and multifocal (n=34, 17.1%). By an independent radiology review, patients were classified to BRPC (n=75, 37.7%) and LAUPC (n=124, 62.3%). With median 40.3 months (95% CI, 36.7-43.8) of follow-up duration in surviving patients, ORR was 26.6% (n=53), median PFS and OS were 10.6 months (95% CI, 9.5-11.7) and 17.1 months (95% CI, 13.2-20.9), respectively. There was no difference in PFS and OS between BRPC and LAUPC (median PFS, 11.1 months [95% CI, 8.8-13.5] vs. 10.1 months [95% CI, 8.4-11.8], p=0.47); (median OS, 18.4 months [95% CI, 16.1-20.8] vs. 17.1 months [95% CI, 13.2-20.9], p=0.50). Curative-intent surgery (R0 and R1) was done in 63 patients (33.2%, 49 for R0 and 14 for R1) after treatment with (m)FOLFIRINOX. Resection rates were 58.2% in BRPC patients and 19.4% in LAUPC patients (p
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- 2020
104. Expression of seven carbonic anhydrases in red alga Gracilariopsis chorda and their subcellular localization in a heterologous system, Arabidopsis thaliana
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Inhwan Hwang, Jeong Hee Kim, Dong-Wook Lee, Hwan Su Yoon, JunMo Lee, and Md. Abdur Razzak
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0106 biological sciences ,0301 basic medicine ,Gene isoform ,Signal peptide ,Glycosylation ,Sequence analysis ,Chorda ,Green Fluorescent Proteins ,Arabidopsis ,Golgi Apparatus ,Plant Science ,Endoplasmic Reticulum ,01 natural sciences ,Gene Expression Regulation, Enzymologic ,03 medical and health sciences ,Carbonic anhydrase ,Arabidopsis thaliana ,Computer Simulation ,Gene ,Phylogeny ,Carbonic Anhydrases ,biology ,Protoplasts ,General Medicine ,biology.organism_classification ,Subcellular localization ,Plants, Genetically Modified ,Protein Transport ,030104 developmental biology ,Biochemistry ,Rhodophyta ,Vacuoles ,biology.protein ,Agronomy and Crop Science ,010606 plant biology & botany ,Subcellular Fractions - Abstract
Red alga, Gracilariopsis chorda, contains seven carbonic anhydrases that can be grouped into α-, β- and γ-classes. Carbonic anhydrases (CAHs) are metalloenzymes that catalyze the reversible hydration of CO2. These enzymes are present in all living organisms and play roles in various cellular processes, including photosynthesis. In this study, we identified seven CAH genes (GcCAHs) from the genome sequence of the red alga Gracilariopsis chorda and characterized them at the molecular, cellular and biochemical levels. Based on sequence analysis, these seven isoforms were categorized into four α-class, one β-class, and two γ-class isoforms. RNA sequencing revealed that of the seven CAHs isoforms, six genes were expressed in G. chorda in light at room temperature. In silico analysis revealed that these seven isoforms localized to multiple subcellular locations such as the ER, mitochondria and cytosol. When expressed as green fluorescent protein fusions in protoplasts of Arabidopsis thaliana leaf cells, these seven isoforms showed multiple localization patterns. The four α-class GcCAHs with an N-terminal hydrophobic leader sequence localized to the ER and two of them were further targeted to the vacuole. GcCAHβ1 with no noticeable signal sequence localized to the cytosol. The two γ-class GcCAHs also localized to the cytosol, despite the presence of a predicted presequence. Based on these results, we propose that the red alga G. chorda also employs multiple CAH isoforms for various cellular processes such as photosynthesis.
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- 2018
105. Nab-paclitaxel plus gemcitabine versus FOLFIRINOX as the first-line chemotherapy for patients with metastatic pancreatic cancer: retrospective analysis
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Sang Soo Lee, Jae Ho Jeong, Kyu-Pyo Kim, Sang Hyun Shin, Dong Wan Seo, Myung-Hwan Kim, Sung Koo Lee, Inhwan Hwang, Jihoon Kang, Seung-Mo Hong, Jae Hoon Lee, Heung-Moon Chang, Baek-Yeol Ryoo, Do Hyun Park, Changhoon Yoo, Ki Byung Song, Tae Jun Song, Dae Wook Hwang, and Song Cheol Kim
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0301 basic medicine ,Oncology ,Male ,medicine.medical_specialty ,Paclitaxel ,FOLFIRINOX ,medicine.medical_treatment ,Leucovorin ,Irinotecan ,Deoxycytidine ,Disease-Free Survival ,Metastasis ,03 medical and health sciences ,0302 clinical medicine ,Internal medicine ,Pancreatic cancer ,Albumins ,Metastatic pancreatic cancer ,Antineoplastic Combined Chemotherapy Protocols ,medicine ,Retrospective analysis ,Organometallic Compounds ,Humans ,Pharmacology (medical) ,Nab-paclitaxel ,Aged ,Retrospective Studies ,Pharmacology ,Chemotherapy ,business.industry ,medicine.disease ,Gemcitabine ,Oxaliplatin ,Pancreatic Neoplasms ,Drug Combinations ,030104 developmental biology ,030220 oncology & carcinogenesis ,Female ,Fluorouracil ,business ,medicine.drug - Abstract
Purpose nab-paclitaxel plus gemcitabine (AG) and FOLFIRINOX have been established as standard first-line treatment in metastatic pancreatic cancer (mPC). We performed retrospective analysis comparing the efficacies of AG and FOLFIRINOX in daily practice setting. Materials and Methods We analyzed 308 patients who presented initially as mPC and received AG (n = 149) or FOLFIRINOX (n = 159) as first-line treatment between 2013 and 2016. Primary endpoints were progression-free survival (PFS) and overall survival (OS). Result There were no significant differences between the two groups in terms of baseline characteristics, except older age and higher Charlson Comorbidity Index (CCI) score in AG group. The response rates (34% vs 34%) and median PFS (6.8 vs 5.1 months) were comparable between two groups (p = 0.88 and p = 0.19, respectively), while median OS was significantly better with AG than FOLFIRINOX (11.4 vs 9.6 months; p = 0.002). Elevated baseline CA19–9 level and liver metastasis were independent adverse prognostic factors for PFS and OS. In subgroup analyses, PFS with AG was better in patients with age ≥ 65 years, peritoneal metastasis, and higher CCI than that with FOLFIRINOX. Conclusion Both AG and FOLFIRINOX showed comparable efficacy outcomes in daily practice setting. AG might be preferentially considered in patients with peritoneal metastasis, comorbid medical conditions or old age.
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- 2018
106. AtCAP2 is crucial for lytic vacuole biogenesis during germination by positively regulating vacuolar protein trafficking
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Yun Kwon, Inhwan Hwang, Jinbo Shen, Myoung Hui Lee, Liwen Jiang, and Kyoung Rok Geem
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0106 biological sciences ,0301 basic medicine ,Protein storage vacuole ,Arabidopsis ,Germination ,Protein degradation ,01 natural sciences ,03 medical and health sciences ,Syntaxin ,Lytic vacuole ,Protein kinase A ,Organelle Biogenesis ,Multidisciplinary ,Arabidopsis Proteins ,Chemistry ,Peripheral membrane protein ,Signal transducing adaptor protein ,Cell biology ,Protein Transport ,030104 developmental biology ,PNAS Plus ,Seeds ,Vacuoles ,Microtubule-Associated Proteins ,Biogenesis ,010606 plant biology & botany - Abstract
Protein trafficking is a fundamental mechanism of subcellular organization and contributes to organellar biogenesis. AtCAP2 is an Arabidopsis homolog of the Mesembryanthemum crystallinum calcium-dependent protein kinase 1 adaptor protein 2 (McCAP2), a member of the syntaxin superfamily. Here, we show that AtCAP2 plays an important role in the conversion to the lytic vacuole (LV) during early plant development. The AtCAP2 loss-of-function mutant atcap2-1 displayed delays in protein storage vacuole (PSV) protein degradation, PSV fusion, LV acidification, and biosynthesis of several vacuolar proteins during germination. At the mature stage, atcap2-1 plants accumulated vacuolar proteins in the prevacuolar compartment (PVC) instead of the LV. In wild-type plants, AtCAP2 localizes to the PVC as a peripheral membrane protein and in the PVC compartment recruits glyceraldehyde-3-phosphate dehydrogenase C2 (GAPC2) to the PVC. We propose that AtCAP2 contributes to LV biogenesis during early plant development by supporting the trafficking of specific proteins involved in the PSV-to-LV transition and LV acidification during early stages of plant development.
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- 2018
107. Evolution and Design Principles of the Diverse Chloroplast Transit Peptides
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Dong Wook, Lee and Inhwan, Hwang
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Chloroplast Proteins ,endosymbiosis ,protein import into chloroplasts ,chloroplast evolution ,food and beverages ,transit peptide ,Minireview ,Protein Sorting Signals ,Symbiosis - Abstract
Chloroplasts are present in organisms belonging to the kingdom Plantae. These organelles are thought to have originated from photosynthetic cyanobacteria through endosymbiosis. During endosymbiosis, most cyanobacterial genes were transferred to the host nucleus. Therefore, most chloroplast proteins became encoded in the nuclear genome and must return to the chloroplast after translation. The N-terminal cleavable transit peptide (TP) is necessary and sufficient for the import of nucleus-encoded interior chloroplast proteins. Over the past decade, extensive research on the TP has revealed many important characteristic features of TPs. These studies have also shed light on the question of how the many diverse TPs could have evolved to target specific proteins to the chloroplast. In this review, we summarize the characteristic features of TPs. We also highlight recent advances in our understanding of TP evolution and provide future perspectives about this important research area.
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- 2018
108. A cost-effective and high standard purification approach for the production of bioactive recombinant human interleukin-6 in Nicotiana benthamiana
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Md Reyazul Islam, Jeon-Soo Lee, and Inhwan Hwang
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- 2018
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109. Feasibility of heart transplantation for patients with amyloid cardiomyopathy accompanying extracardiac organ involvement
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Tae Jin Yun, H. Chae, Jea-Joong kim, Dok Hyun Yoon, Min-Seok Kim, Inhwan Hwang, Cheolwon Suh, Jae Won Lee, Sung Ho Jung, and Chan-Jeoung Park
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Heart transplantation ,Cancer Research ,medicine.medical_specialty ,Oncology ,business.industry ,Internal medicine ,medicine.medical_treatment ,Cardiology ,Medicine ,Organ involvement ,Hematology ,business ,Amyloid cardiomyopathy - Published
- 2019
110. Multi-modal human action recognition using deep neural networks fusing image and inertial sensor data
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Songhwai Oh, Geonho Cha, and Inhwan Hwang
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Artificial neural network ,Computer science ,business.industry ,010401 analytical chemistry ,Feature extraction ,ComputingMethodologies_IMAGEPROCESSINGANDCOMPUTERVISION ,02 engineering and technology ,Sensor fusion ,01 natural sciences ,Convolutional neural network ,0104 chemical sciences ,Recurrent neural network ,Inertial measurement unit ,0202 electrical engineering, electronic engineering, information engineering ,020201 artificial intelligence & image processing ,Computer vision ,Artificial intelligence ,business ,Wireless sensor network ,Pose - Abstract
Human action recognition has been studied in many fields including computer vision and sensor networks using inertial sensors. However, there are limitations such as spatial constraints, occlusions in images, sensor unreliability, and the inconvenience of users. In order to solve these problems we suggest a sensor fusion method for human action recognition exploiting RGB images from a single fixed camera and a single wrist mounted inertial sensor. These two different domain information can complement each other to fill the deficiencies that exist in both image based and inertial sensor based human action recognition methods. We propose two convolutional neural network (CNN) based feature extraction networks for image and inertial sensor data and a recurrent neural network (RNN) based classification network with long short term memory (LSTM) units. Training of deep neural networks and testing are done with synchronized images and sensor data collected from five individuals. The proposed method results in better performance compared to single sensor-based methods with an accuracy of 86.9% in cross-validation. We also verify that the proposed algorithm robustly classifies the target action when there are failures in detecting body joints from images.
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- 2017
111. Wearable gesture control of agile micro quadrotors
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Songhwai Oh, Yunho Choi, and Inhwan Hwang
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0209 industrial biotechnology ,Quadcopter ,business.industry ,Computer science ,Wearable computer ,02 engineering and technology ,Smartwatch ,020901 industrial engineering & automation ,Recurrent neural network ,Inertial measurement unit ,Gesture recognition ,Joystick ,0202 electrical engineering, electronic engineering, information engineering ,020201 artificial intelligence & image processing ,Computer vision ,Artificial intelligence ,business ,Gesture - Abstract
Quadrotor unmanned aerial vehicles (UAVs) have seen a surge of use in various applications due to its structural simplicity and high maneuverability. However, conventional control methods using joysticks prohibit novices from getting used to maneuvering quadrotors in short time. In this paper, we suggest the use of a wearable device, such as a smart watch, as a new remote-controller for a quadrotor. The user's command is recognized as gestures using the 9-DoF inertial measurement unit (IMU) of a wearable device through a recurrent neural network (RNN) with long short-term memory (LSTM) cells. Our implementation also makes it possible to align the heading of a quadrotor with the heading of the user. Our implementation allows nine different gestures and the trained RNN is used for real-time gesture recognition for controlling a micro quadrotor. The proposed system exploits available sensors in a wearable device and a quadrotor as much as possible to make the gesture-based control intuitive. We have experimentally validated the performance of the proposed system by using a Samsung Gear S smart watch and a Crazyflie Nano Quadcopter.
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- 2017
112. Physiological Functions of the COPI Complex in Higher Plants
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Yong Won Kang, Hyun Sook Pai, Hee-Kyung Ahn, Hye Min Lim, and Inhwan Hwang
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cell plate formation ,autophagy ,Golgi Apparatus ,virus-induced gene silencing ,Apoptosis ,Biology ,Coatomer Protein ,Article ,Coat Protein Complex I ,symbols.namesake ,Tobacco ,Molecular Biology ,COPII ,Secretory pathway ,Cytokinesis ,Plant Proteins ,Vesicular-tubular cluster ,fungi ,Golgi localization ,Cell Biology ,General Medicine ,Cell plate ,COPI ,COP-Coated Vesicles ,Golgi apparatus ,Cell biology ,Protein Transport ,cell death ,Biochemistry ,Coatomer ,symbols ,RNA Interference ,COPI vesicle - Abstract
COPI vesicles are essential to the retrograde transport of proteins in the early secretory pathway. The COPI coatomer complex consists of seven subunits, termed α-, β-, β'-, γ-, δ-, e-, and ζ-COP, in yeast and mammals. Plant genomes have homologs of these subunits, but the essentiality of their cellular functions has hampered the functional characterization of the subunit genes in plants. Here we have employed virus-induced gene silencing (VIGS) and dexamethasone (DEX)-inducible RNAi of the COPI subunit genes to study the in vivo functions of the COPI coatomer complex in plants. The β'-, γ-, and δ-COP subunits localized to the Golgi as GFP-fusion proteins and interacted with each other in the Golgi. Silencing of β'-, γ-, and δ-COP by VIGS resulted in growth arrest and acute plant death in Nicotiana benthamiana, with the affected leaf cells exhibiting morphological markers of programmed cell death. Depletion of the COPI subunits resulted in disruption of the Golgi structure and accumulation of autolysosome-like structures in earlier stages of gene silencing. In tobacco BY-2 cells, DEX-inducible RNAi of β'-COP caused aberrant cell plate formation during cytokinesis. Collectively, these results suggest that COPI vesicles are essential to plant growth and survival by maintaining the Golgi apparatus and modulating cell plate formation.
- Published
- 2015
113. Sequence Motifs in Transit Peptides Act as Independent Functional Units and Can Be Transferred to New Sequence Contexts
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Seungjin Woo, Inhwan Hwang, Dong-Wook Lee, and Kyoung Rok Geem
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Chloroplast ,Rubisco Small Subunit ,Physiology ,Botany ,Genetics ,Sequence (biology) ,Plant Science ,Computational biology ,Biology ,Sequence motif ,Translocon - Abstract
A large number of nuclear-encoded proteins are imported into chloroplasts after they are translated in the cytosol. Import is mediated by transit peptides (TPs) at the N termini of these proteins. TPs contain many small motifs, each of which is critical for a specific step in the process of chloroplast protein import; however, it remains unknown how these motifs are organized to give rise to TPs with diverse sequences. In this study, we generated various hybrid TPs by swapping domains between Rubisco small subunit (RbcS) and chlorophyll a/b-binding protein, which have highly divergent sequences, and examined the abilities of the resultant TPs to deliver proteins into chloroplasts. Subsequently, we compared the functionality of sequence motifs in the hybrid TPs with those of wild-type TPs. The sequence motifs in the hybrid TPs exhibited three different modes of functionality, depending on their domain composition, as follows: active in both wild-type and hybrid TPs, active in wild-type TPs but inactive in hybrid TPs, and inactive in wild-type TPs but active in hybrid TPs. Moreover, synthetic TPs, in which only three critical motifs from RbcS or chlorophyll a/b-binding protein TPs were incorporated into an unrelated sequence, were able to deliver clients to chloroplasts with a comparable efficiency to RbcS TP. Based on these results, we propose that diverse sequence motifs in TPs are independent functional units that interact with specific translocon components at various steps during protein import and can be transferred to new sequence contexts.
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- 2015
114. Targeting and biogenesis of transporters and channels in chloroplast envelope membranes: Unsolved questions
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Inhwan Hwang and Young Jun Oh
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Signal peptide ,Chloroplasts ,Ion Transport ,Nuclear gene ,Arabidopsis Proteins ,Physiology ,Arabidopsis ,Membrane Transport Proteins ,food and beverages ,Translation (biology) ,Cell Biology ,Biology ,Chloroplast membrane ,Ribosome ,Cell biology ,Chloroplast ,Chloroplast DNA ,Biochemistry ,Calcium ,Molecular Biology ,Biogenesis - Abstract
Chloroplasts produce carbohydrates, hormones, vitamins, amino acids, pigments, nucleotides, ATP, and secondary metabolites. Channels and transporters are required for the movement of molecules across the two chloroplast envelope membranes. These transporters and channel proteins are grouped into two different types, including β-barrel proteins and transmembrane-domain (TMD) containing proteins. Most β-barrel proteins are localized at the outer chloroplast membrane, and TMD-containing proteins are localized at the inner chloroplast membrane. Many of these transporters and channels are encoded by nuclear genes; therefore, they have to be imported into chloroplasts after translation on cytosolic ribosomes. These proteins should have specific targeting signals for their final destination in the chloroplast membrane and for assembly into specific complexes. In this review, we summarize recent progress in the identification, functional characterization, and biogenesis of transporters and channels at the chloroplast envelope membranes, and discuss outstanding questions regarding transporter and channel protein biogenesis.
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- 2015
115. VibePhone: efficient surface recognition for smartphones using vibration
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Songhwai Oh, Jungchan Cho, and Inhwan Hwang
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0209 industrial biotechnology ,Computer science ,business.industry ,020206 networking & telecommunications ,Context (language use) ,02 engineering and technology ,Accelerometer ,Object (computer science) ,Vibrator (mechanical) ,020901 industrial engineering & automation ,Artificial Intelligence ,Pattern recognition (psychology) ,0202 electrical engineering, electronic engineering, information engineering ,Object type ,Context awareness ,Computer vision ,Computer Vision and Pattern Recognition ,Artificial intelligence ,Haptic perception ,business - Abstract
With various sensors in a smartphone, it is now possible to obtain information about a user and her surroundings, such as the location and the activity of the smartphone user, and the obtained context information is being used to provide new services to the users. In this paper, we propose VibePhone, which uses a built-in vibrator and accelerometer, for efficiently recognizing the type of surfaces contacted by a smartphone, enabling the sense of touch to smartphones. In particular, this paper focuses on developing a succinct set of features that are useful for recognizing surface types for reducing computation and memory requirements, which will in turn reduce the power consumption of the device. For humans and animals, the sense of touch, which is obtained from the texture of an object by scrubbing its surface, is fundamental for both recognizing and learning the properties of objects. Since a smartphone cannot physically scrub the contacting surface, we emulate the touch by generating vibrations and analyzing accelerometer readings. While the recognition of the object type by vibration alone is an extremely difficult task, even for a human, we demonstrate that it is possible to distinguish object types into broad categories where a phone is usually placed. For efficiency, VibePhone uses only a small subset of the most informative features of accelerometer readings using feature selection and reduces the computation time by 92 % using only 15 % of features, while maintaining performance. We believe that our analysis of vibrations about contacted surfaces can provide an important insight for the haptic perception in future smartphones, enabling new experiences to the users.
- Published
- 2015
116. Evolution of rubisco complex small subunit transit peptides from algae to plants
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Yun-Joo Yoo, Dong-Wook Lee, Md. Abdur Razzak, and Inhwan Hwang
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0106 biological sciences ,0301 basic medicine ,Ribulose-Bisphosphate Carboxylase ,Amino Acid Motifs ,Arabidopsis ,lcsh:Medicine ,Chlamydomonas reinhardtii ,medicine.disease_cause ,01 natural sciences ,Article ,Evolution, Molecular ,Chloroplast Proteins ,03 medical and health sciences ,Chlorophyta ,Protein targeting ,Botany ,medicine ,Amino Acid Sequence ,Plastid ,lcsh:Science ,Multidisciplinary ,biology ,lcsh:R ,RuBisCO ,food and beverages ,Plants ,biology.organism_classification ,Chloroplast ,030104 developmental biology ,Biochemistry ,biology.protein ,lcsh:Q ,Peptides ,Sequence motif ,010606 plant biology & botany - Abstract
Chloroplasts evolved from a free-living cyanobacterium acquired by the ancestor of all photosynthetic eukaryotes, including algae and plants, through a single endosymbiotic event. During endosymbiotic conversion, the majority of genes in the endosymbiont were transferred to the host nucleus and many of the proteins encoded by these genes must therefore be transported into the chloroplast after translation in the cytosol. Chloroplast-targeted proteins contain a targeting signal, named the transit peptide (TP), at the N-terminus. However, the evolution of TPs is not well understood. In this study, TPs from RbcS (rubisco small subunit) were compared between lower and higher eukaryotes. Chlamydomonas reinhardtii RbcS (CrRbcS) TP was non-functional in Arabidopsis. However, inclusion of a critical sequence motif, FP-RK, from Arabidopsis thaliana RbcS (AtRbcS) TP allowed CrRbcS TP to deliver proteins into plant chloroplasts. The position of the FP-RK motif in CrRbcS TP was critical for function. The QMMVW sequence motif in CrRbcS TP was crucial for its transport activity in plants. CrRbcS TPs containing additional plant motifs remained functional in C. reinhardtii. These results suggest that TPs evolved by acquiring additional sequence motifs to support protein targeting to chloroplasts during evolution of land plants from algae.
- Published
- 2017
117. SCYL2 Genes Are Involved in Clathrin-Mediated Vesicle Trafficking and Essential for Plant Growth
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Dong-Wook Lee, Daniel P. Schachtman, Ji-Yul Jung, Inhwan Hwang, and Stephen B. Ryu
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0106 biological sciences ,0301 basic medicine ,Physiology ,Arabidopsis ,Plant Development ,Plant Science ,Root hair ,Biology ,Protein Serine-Threonine Kinases ,01 natural sciences ,Clathrin ,Plant Roots ,Clathrin Heavy Chains ,03 medical and health sciences ,Bimolecular fluorescence complementation ,Genetics ,Arabidopsis Proteins ,Vesicle ,fungi ,food and beverages ,Clathrin-Coated Vesicles ,Articles ,Qb-SNARE Proteins ,Cell biology ,Transport protein ,Vesicular transport protein ,Adaptor Proteins, Vesicular Transport ,030104 developmental biology ,biology.protein ,Soluble NSF attachment protein ,010606 plant biology & botany ,trans-Golgi Network - Abstract
Protein transport between organelles is an essential process in all eukaryotic cells and is mediated by the regulation of processes such as vesicle formation, transport, docking, and fusion. In animals, SCY1-LIKE2 (SCYL2) binds to clathrin and has been shown to play roles in trans-Golgi network-mediated clathrin-coated vesicle trafficking. Here, we demonstrate that SCYL2A and SCYL2B, which are Arabidopsis (Arabidopsis thaliana) homologs of animal SCYL2, are vital for plant cell growth and root hair development. Studies of the SCYL2 isoforms using multiple single or double loss-of-function alleles show that SCYL2B is involved in root hair development and that SCYL2A and SCYL2B are essential for plant growth and development and act redundantly in those processes. Quantitative reverse transcription-polymerase chain reaction and a β-glucuronidase-aided promoter assay show that SCYL2A and SCYL2B are differentially expressed in various tissues. We also show that SCYL2 proteins localize to the Golgi, trans-Golgi network, and prevacuolar compartment and colocalize with Clathrin Heavy Chain1 (CHC1). Furthermore, bimolecular fluorescence complementation and coimmunoprecipitation data show that SCYL2B interacts with CHC1 and two Soluble NSF Attachment Protein Receptors (SNAREs): Vesicle Transport through t-SNARE Interaction11 (VTI11) and VTI12. Finally, we present evidence that the root hair tip localization of Cellulose Synthase-Like D3 is dependent on SCYL2B. These findings suggest the role of SCYL2 genes in plant cell developmental processes via clathrin-mediated vesicle membrane trafficking.
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- 2017
118. SH3 Domain-Containing Protein 2 Plays a Crucial Role at the Step of Membrane Tubulation during Cell Plate Formation
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Liwen Jiang, Youngdae Yoon, Hong Hanh, Wonhwa Cho, Inhwan Hwang, Kaveesha J. Wijesinghe, Hyeran Kim, Zizhen Liang, Robert V. Stahelin, Gyeongik Ahn, Dae Heon Kim, Kristen A. Johnson, Byung-Ho Kang, Indira Singaram, and Xiaohong Zhuang
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0301 basic medicine ,Dynamins ,Leading edge ,Membrane tubulation ,Arabidopsis Proteins ,Vesicle ,Arabidopsis ,Cell Biology ,Plant Science ,Cell plate ,Biology ,biology.organism_classification ,Plants, Genetically Modified ,SH3 domain ,Cell biology ,03 medical and health sciences ,030104 developmental biology ,BAR domain ,Arabidopsis thaliana ,Carrier Proteins ,Cytokinesis ,Research Articles ,trans-Golgi Network - Abstract
During cytokinesis in plants, trans-Golgi network-derived vesicles accumulate at the center of dividing cells and undergo various structural changes to give rise to the planar cell plate. However, how this conversion occurs at the molecular level remains elusive. In this study, we report that SH3 Domain-Containing Protein 2 (SH3P2) in Arabidopsis thaliana plays a crucial role in converting vesicles to the planar cell plate. SH3P2 RNAi plants showed cytokinesis-defective phenotypes and produced aggregations of vesicles at the leading edge of the cell plate. SH3P2 localized to the leading edge of the cell plate, particularly the constricted or curved regions of the cell plate. The BAR domain of SH3P2 induced tubulation of vesicles. SH3P2 formed a complex with dynamin-related protein 1A (DRP1A) and affected DRP1A accumulation to the cell plate. Based on these results, we propose that SH3P2 functions together with DRP1A to convert the fused vesicles to tubular structures during cytokinesis.
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- 2017
119. Sorting of nuclear-encoded chloroplast membrane proteins
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Junho Lee, Dong-Wook Lee, and Inhwan Hwang
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0301 basic medicine ,food and beverages ,Membrane Proteins ,Plant Science ,Biology ,medicine.disease_cause ,Chloroplast membrane ,Cell biology ,Transport protein ,Twin-arginine translocation pathway ,03 medical and health sciences ,Chloroplast Proteins ,Protein Transport ,030104 developmental biology ,Membrane ,Membrane protein ,Thylakoid ,Protein targeting ,medicine - Abstract
Among the many organelles in eukaryotic cells, chloroplasts have the most complex structure, with multiple suborganellar membranes, making protein targeting to chloroplasts, particularly to various suborganellar membranes, highly challenging. Multiple mechanisms function in the biogenesis of chloroplast membrane proteins. Nuclear-encoded nascent proteins can be targeted to the outer envelope membrane directly from the cytosol after translation, but their targeting to the inner envelope and thylakoid membranes requires multiple steps, including cytosolic sorting, translocation across the envelope membranes, sorting in the stroma, and insertion into their target membranes. In this review, we discuss the current knowledge about the sorting mechanisms of proteins to the two envelope membranes and the thylakoid membrane, along with perspectives for future research.
- Published
- 2017
120. Transit peptide elements mediate selective protein targeting to two different types of chloroplasts in the single-cell C4 species Bienertia sinuspersici
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Diana Wimmer, Inhwan Hwang, Vinay Shekhar, Philipp Bohnhorst, and Sascha Offermann
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0106 biological sciences ,0301 basic medicine ,Chloroplasts ,sequence motifs ,ved/biology.organism_classification_rank.species ,Cell ,c-4 photosynthesis ,bundle-sheath ,Biology ,medicine.disease_cause ,01 natural sciences ,kranz anatomy ,Article ,import apparatus ,03 medical and health sciences ,Subcellular Protein Targeting ,Transit Peptide ,Protein targeting ,Botany ,medicine ,metabolic pathways ,RNA, Messenger ,Photosynthesis ,Dewey Decimal Classification::500 | Naturwissenschaften ,Plant Proteins ,cycloptera chenopodiaceae ,aralocaspica chenopodiaceae ,Multidisciplinary ,Amaranthaceae ,ved/biology ,Protoplasts ,Bienertia sinuspersici ,food and beverages ,distinct pathways ,Transport protein ,Cell biology ,Chloroplast ,Plant Leaves ,Protein Transport ,030104 developmental biology ,medicine.anatomical_structure ,ddc:500 ,Function (biology) ,leaf development ,010606 plant biology & botany - Abstract
Bienertia sinuspersici is a terrestrial plant that performs C4 photosynthesis within individual cells through operating a carbon concentrating mechanism between different subcellular domains including two types of chloroplasts. It is currently unknown how differentiation of two highly specialized chloroplasts within the same cell occurs as no similar cases have been reported. Here we show that this differentiation in photosynthetic cells of B. sinuspersici is enabled by a transit peptide (TP) mediated selective protein targeting mechanism. Mutations in the TPs cause loss of selectivity but not general loss of chloroplast import, indicating the mechanism operates by specifically blocking protein accumulation in one chloroplast type. Hybrid studies indicate that this selectivity is transferable to transit peptides of plants which perform C4 by cooperative function of chloroplasts between two photosynthetic cells. Codon swap experiments as well as introducing an artificial bait mRNA show that RNA affects are not crucial for the sorting process. In summary, our analysis shows how the mechanism of subcellular targeting to form two types of chloroplast within the same cell can be achieved. This information is not only crucial for understanding single-cell C4 photosynthesis; it provides new insights in control of subcellular protein targeting in cell biology. DFG/OF106/1-1 DFG/OF 106/1-1 Cooperative Research Program for Agriculture Science & Technology Development Rural Development Administration, Republic of Korea
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- 2017
121. Adaptor proteins in protein trafficking between endomembrane compartments in plants
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Myoung Hui Lee and Inhwan Hwang
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Cytosol ,biology ,Biochemistry ,Vesicle ,biology.protein ,Signal transducing adaptor protein ,Clathrin adaptor proteins ,Endomembrane system ,Plant Science ,Lytic vacuole ,Endocytosis ,Clathrin ,Cell biology - Abstract
Clathrin is a highly conserved coat protein that plays a critical role in lipid vesicle-mediated trafficking at multiple routes in various post-Golgi compartments. It consists of large and small subunits, and exists in the cytosol as triskelions composed of three pairs of small and large subunits. For vesicle formation, the triskelions are recruited to the membrane of specific compartments where they undergo self-polymerization to produce coats for lipid vesicles. However, clathrin has no ability to bind directly to lipid membranes. Therefore, accessory proteins are necessary for its recruitment to the donor compartment where vesicles are formed. A large number of accessory proteins, called adaptor proteins, have been identified and characterized extensively at the molecular and cellular levels in animal cells and yeast. Recently, the roles of many adaptor proteins have been elucidated in plant cells. As expected from the conserved nature of lipidmediated trafficking in eukaryotic cells, these plant adaptor proteins for clathrin show a high degree of functional conservation with those found in animal cells and yeast. At the same time, they are also involved in plant-specific processes such as the transition from the PSV to the lytic vacuole and cell-plate formation. Here, we summarize recent advances in the physiological roles of adaptor proteins in plant cells.
- Published
- 2014
122. An Ankyrin Repeat Domain of AKR2 Drives Chloroplast Targeting through Coincident Binding of Two Chloroplast Lipids
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Hwan Su Yoon, Gwang Hyeon Gwon, Seohyeon Song, Antonina Silkov, Eun Chan Yang, Barry Honig, Kyungyoung Song, Dae Heon Kim, Younghyun Kim, Wonhwa Cho, Inhwan Hwang, Zheng Yi Xu, Yunje Cho, and Mi Jeong Park
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Models, Molecular ,Chloroplasts ,Molecular Sequence Data ,Arabidopsis ,Plasma protein binding ,Biology ,Cyanobacteria ,General Biochemistry, Genetics and Molecular Biology ,Cytosol ,Protein structure ,Amino Acid Sequence ,Binding site ,Symbiosis ,Molecular Biology ,Peptide sequence ,Binding Sites ,Sequence Homology, Amino Acid ,Arabidopsis Proteins ,Galactolipids ,Phosphatidylglycerols ,Cell Biology ,Chloroplast outer membrane ,Lipids ,Ankyrin Repeat ,Protein Structure, Tertiary ,Chloroplast ,Biochemistry ,Ankyrin repeat ,Function (biology) ,Molecular Chaperones ,Protein Binding ,Developmental Biology - Abstract
SummaryIn organellogenesis of the chloroplast from endosymbiotic cyanobacteria, the establishment of protein-targeting mechanisms to the chloroplast should have been pivotal. However, it is still mysterious how these mechanisms were established and how they work in plant cells. Here we show that AKR2A, the cytosolic targeting factor for chloroplast outer membrane (COM) proteins, evolved from the ankyrin repeat domain (ARD) of the host cell by stepwise extensions of its N-terminal domain and that two lipids, monogalactosyldiacylglycerol (MGDG) and phosphatidylglycerol (PG), of the endosymbiont were selected to function as the AKR2A receptor. Structural analysis, molecular modeling, and mutational analysis of the ARD identified two adjacent sites for coincidental and synergistic binding of MGDG and PG. Based on these findings, we propose that the targeting mechanism of COM proteins was established using components from both the endosymbiont and host cell through a modification of the protein-protein-interacting ARD into a lipid binding domain.
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- 2014
123. OPTIMUS:online persistent tracking and identification of many users for smart spaces
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Inhwan Hwang, Dong-Hoon Lee, and Songhwai Oh
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business.industry ,Computer science ,Association (object-oriented programming) ,Smart spaces ,Accelerometer ,Tracking (particle physics) ,Computer Science Applications ,Identification (information) ,Hardware and Architecture ,Histogram ,Pattern recognition (psychology) ,Computer vision ,Computer Vision and Pattern Recognition ,Artificial intelligence ,Smart camera ,business ,Software - Abstract
A smart space, which is embedded with networked sensors and smart devices, can provide various useful services to its users. For the success of a smart space, the problem of tracking and identification of smart space users is of paramount importance. We propose a system, called Optimus, for persistent tracking and identification of users in a smart space, which is equipped with a camera network. We assume that each user carries a smartphone in a smart space. A camera network is used to solve the problem of tracking multiple users in a smart space and information from smartphones is used to identify tracks. For robust tracking, we first detect human subjects from images using a head detection algorithm based on histograms of oriented gradients. Then, human detections are combined to form tracklets and delayed track-level association is used to combine tracklets to build longer trajectories of users. Last, accelerometers in smartphones are used to disambiguate identities of trajectories. By linking identified trajectories, we show that the average length of a track can be lengthened by over six times. The performance of the proposed system is evaluated extensively in realistic scenarios.
- Published
- 2014
124. Abscisic Acid Uridine Diphosphate Glucosyltransferases Play a Crucial Role in Abscisic Acid Homeostasis in Arabidopsis
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Ting Dong, Inhwan Hwang, Zheng-Yi Xu, Dae Heon Kim, Yongjik Lee, and Young Min Park
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Osmosis ,Physiology ,Recombinant Fusion Proteins ,Green Fluorescent Proteins ,Abscisic acid homeostasis ,Mutant ,Arabidopsis ,Germination ,Plant Science ,Sodium Chloride ,Hydroxylation ,Uridine Diphosphate ,chemistry.chemical_compound ,Cytosol ,Gene Expression Regulation, Plant ,Stress, Physiological ,RNA interference ,Genetics ,Homeostasis ,Arabidopsis thaliana ,Abscisic acid ,Sequence Homology, Amino Acid ,biology ,Arabidopsis Proteins ,organic chemicals ,fungi ,Glycosyltransferases ,food and beverages ,Articles ,biology.organism_classification ,Protein Transport ,Uridine diphosphate ,Phenotype ,Solubility ,Biochemistry ,chemistry ,Glucosyltransferases ,Mutation ,RNA Interference ,Ectopic expression ,Abscisic Acid - Abstract
The phytohormone abscisic acid (ABA) is crucial for plant growth and adaptive responses to various stress conditions. Plants continuously adjust the ABA level to meet physiological needs, but how ABA homeostasis occurs is not fully understood. This study provides evidence that UGT71B6, an ABA uridine diphosphate glucosyltransferase (UGT), and its two closely related homologs, UGT71B7 and UGT71B8, play crucial roles in ABA homeostasis and in adaptation to dehydration, osmotic stress, and high-salinity stresses in Arabidopsis (Arabidopsis thaliana). UGT RNA interference plants that had low levels of these three UGT transcripts displayed hypersensitivity to exogenous ABA and high-salt conditions during germination and exhibited a defect in plant growth. However, the ectopic expression of UGT71B6 in the atbg1 (for β-glucosidase) mutant background aggravated the ABA-deficient phenotype of atbg1 mutant plants. In addition, modulation of the expression of the three UGTs affects the expression of CYP707A1 to CYP707A4, which encode ABA 8'-hydroxylases; four CYP707As were expressed at higher levels in the UGT RNA interference plants but at lower levels in the UGT71B6:GFP-overexpressing plants. Based on these data, this study proposes that UGT71B6 and its two homologs play a critical role in ABA homeostasis by converting active ABA to an inactive form (abscisic acid-glucose ester) depending on intrinsic cellular and environmental conditions in plants.
- Published
- 2014
125. Hyperprogressive disease (HPD) in genitourinary (GU) cancer patients treated with PD-1/PD-L1 inhibitors
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Inhwan Hwang, Inkeun Park, Jae-Lyun Lee, and Shinkyo Yoon
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Oncology ,Cancer Research ,medicine.medical_specialty ,biology ,business.industry ,Genitourinary system ,medicine.medical_treatment ,Disease progression ,Cancer ,Immunotherapy ,Disease ,medicine.disease ,Internal medicine ,PD-L1 ,biology.protein ,Medicine ,business - Abstract
369 Background: Post immunotherapy response and disease progression pattern are still under investigation. In terms of progression, rapid progression pattern called HPD has been observed during the early cycles of PD-1/PD-L1 inhibitors therapy in patients with various cancers. Data of HPD in patients with GU cancer are limited. Methods: This study included 203 patients with GU cancer treated with PD-1/PD-L1 inhibitors between February 2015 and June 2018. HPD was defined as greater than 50% increase in tumor burden compared with before-treatment computed tomography (CT) scan and greater than 2-fold increase in tumor growth rate (TGR) compared with previous TGR. Development of extensive new lesions were also regarded as HPD. Response was evaluated by RECIST v1.1 on at least two consecutive CT scan before the initiation of PD-1/PD-L1 inhibitors and one CT scan within 2 months after initiation of PD-1/PD-L1 inhibitors. Results: 102 patients with renal cell carcinoma (RCC) and 101 patients with urothelial cell carcinoma (UCC) were included. Median age was 64 years (range 56-71) and 141 patients (70%) were male. The most common metastatic site was lymph node (66%), followed by lung (51%) and bone (34%). HPD was observed in 13 out of 230 patients (5.7%; 95% CI, 5.0-6.3). Median OS for patients with progressive disease and HPD were 7.3 months and 3.5 months, respectively ( p=0.09). HPD was more frequently found in patients with UCC than RCC (11% vs. 0.9%, p=0.01). In multivariate analysis, patients of UCC ( p upper normal limit, p=0.01), and hypoalbumimemia (< 3.5 gm/dL, p
- Published
- 2019
126. Generation of transgenic Arabidopsis plants expressing mcherry-fused organelle marker proteins
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Dae Heon Kim, Zheng-Yi Xu, and Inhwan Hwang
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biology ,Live cell imaging ,Arabidopsis ,Endoplasmic reticulum ,Organelle ,food and beverages ,Plant Science ,Lytic vacuole ,Cell fractionation ,biology.organism_classification ,mCherry ,Protein subcellular localization prediction ,Cell biology - Abstract
In eukaryotic cells, a major proportion of the cellular proteins localize to various subcellular organelles where they are involved in organelle-specific cellular processes. Thus, the localization of a particular protein in the cell is an important part of understanding the physiological role of the protein in the cell. Various approaches such as subcellular fractionation, immunolocalization and live imaging have been used to define the localization of organellar proteins. Of these various approaches, the most powerful one is the live imaging because it can show in vivo dynamics of protein localization depending on cellular and environmental conditions without disturbing cellular structures. However, the live imaging requires the ability to detect the organelles in live cells. In this study, we report generation of a new set of transgenic Arabidopsis plants using various organelle marker proteins fused to a fluorescence protein, monomeric Cherry (mCherry). All these markers representing different subcellular organelles such as chloroplasts, mitochondria, peroxisomes, endoplasmic reticulum (ER) and lytic vacuole showed clear and specific signals regardless of the cell types and tissues. These marker lines can be used to determine localization of organellar proteins by colocalization and also to study the dynamics of organelles under various developmental and environmental conditions.
- Published
- 2013
127. In Vivo Removal of N-Terminal Fusion Domains From Recombinant Target Proteins Produced in Nicotiana benthamiana.
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Islam, Md Reyazul, Seoyoung Choi, Muthamilselvan, Thangarasu, Kunyoo Shin, and Inhwan Hwang
- Abstract
Plants show great potential for producing recombinant proteins in a cost-effective manner. Many strategies have therefore been employed to express high levels of recombinant proteins in plants. Although foreign domains are fused to target proteins for high expression or as an affinity tag for purification, the retention of foreign domains on a target protein may be undesirable, especially for biomedical purposes. Thus, their removal is often crucial at a certain time point after translation. Here, we developed a new strategy to produce target proteins without foreign domains. This involved in vivo removal of foreign domains fused to the N-terminus by the small ubiquitin-related modifier (SUMO) domain/SUMO-specific protease system. This strategy was tested successfully by generating a recombinant gene, BiP:p38:bdSUMO : His:hLIF, that produced human leukemia inhibitory factor (hLIF) fused to p38, a coat protein of the Turnip crinkle virus; the inclusion of p38 increased levels of protein expression. The recombinant protein was expressed at high levels in the leaf tissue of Nicotiana benthamiana. Coexpression of bdSENP1, a SUMO-specific protease, proteolytically released His:hLIF from the full-length recombinant protein in the endoplasmic reticulum of N. benthamiana leaf cells. His:hLIF was purified from leaf extracts via Ni
2+ –NTA affinity purification resulting in a yield of 32.49 mg/kg, and the N-terminal 5-residues were verified by amino acid sequencing. Plant-produced His:hLIF was able to maintain the pluripotency of mouse embryonic stem cells. This technique thus provides a novel method of removing foreign domains from a target protein in planta. [ABSTRACT FROM AUTHOR]- Published
- 2020
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128. Clinical Outcomes of Second-Line Chemotherapy after Progression on Nab-Paclitaxel Plus Gemcitabine in Patients with Metastatic Pancreatic Adenocarcinoma.
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Kyoungmin Lee, Kyunghye Bang, Changhoon Yoo, Inhwan Hwang, Jae Ho Jeong, Heung-Moon Chang, Dongwook Oh, Tae Jun Song, Do Hyun Park, Sang Soo Lee, Sung Koo Lee, Myung-Hwan Kim, Jin-hong Park, Kyu-pyo Kim, and Baek-Yeol Ryoo
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CANCER chemotherapy ,ADENOCARCINOMA ,PROGRESSION-free survival ,TREATMENT effectiveness ,FACTOR analysis - Abstract
Purpose Since the introduction of nab-paclitaxel plus gemcitabine (nab-P+GEM) as first-line (1L) treatment for metastatic pancreatic adenocarcinoma (mPDAC), optimal second-line (2L) chemotherapy after progression is unclear. We assessed clinical outcomes of 2L chemotherapy for disease that progressed on 1L nab-P+GEM. Materials and Methods Among the 203 patients previously treated with 1L nab-P+GEM for mPDAC at Asan Medical Center, between February and December 2016, records of 120 patients receiving 2L chemotherapy after progression on nab-P+GEM were retrospectively reviewed. The response rate and survival were evaluated along with analysis of prognostic factors. Results Fluoropyrimidine-oxaliplatin doublets (FOLFOX or XELOX) were used in 78 patients (65.0%), fluoropyrimidine monotherapy in 37 (30.8%), and liposomal irinotecan plus fluorouracil in two (1.7%). The median progression-free survival (PFS) and overall survival (OS) were 3.29 months and 7.33 months from the start of 2L therapy. Fluoropyrimidine-oxaliplatin regimens and fluoropyrimidine monotherapy did not yield significantly different median PFS (2.89 months vs. 3.81 months, p=0.40) or OS (7.04 months vs. 7.43 months, p=0.86). A high neutrophil-lymphocyte ratio (> 2.2) and a short time to progression with 1L nab-P+GEM (< 6.4 months) were independent prognostic factors of poor OS with 2L therapy. Conclusion 2L fluoropyrimidine-oxaliplatin doublets and fluoropyrimidine monotherapy after failure of 1L nab-P+GEM had modest efficacy, with no differences in treatment outcomes between them. Further investigation is warranted for the optimal 2L chemo-regimens and sequencing of systemic chemotherapy for patients with mPDAC. [ABSTRACT FROM AUTHOR]
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- 2020
- Full Text
- View/download PDF
129. The Clathrin Adaptor Complex AP-2 Mediates Endocytosis of BRASSINOSTEROID INSENSITIVE1 inArabidopsis
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Dominique Eeckhout, Isabelle Vanhoutte, Zheng-Yi Xu, Geert De Jaeger, Eugenia Russinova, Jürgen Kleine-Vehn, Sibu Simon, Kyungyoung Song, Jiří Friml, Wim Dejonghe, Soo Youn Kim, Astrid Gadeyne, Geert Persiau, Niloufer G. Irani, Daniël Van Damme, Inhwan Hwang, and Simone Di Rubbo
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0106 biological sciences ,Clathrin adaptor complex ,education ,Adaptor Protein Complex 2 ,Arabidopsis ,Plant Science ,Endocytosis ,Plant Roots ,01 natural sciences ,Clathrin ,03 medical and health sciences ,chemistry.chemical_compound ,Brassinosteroid ,Research Articles ,030304 developmental biology ,0303 health sciences ,biology ,Arabidopsis Proteins ,Cell Membrane ,fungi ,Signal transducing adaptor protein ,Cell Biology ,Receptor-mediated endocytosis ,biology.organism_classification ,Cell biology ,Protein Transport ,chemistry ,biology.protein ,Clathrin adaptor proteins ,Protein Kinases ,Protein Binding ,Signal Transduction ,010606 plant biology & botany - Abstract
Clathrin-mediated endocytosis (CME) regulates many aspects of plant development, including hormone signaling and responses to environmental stresses. Despite the importance of this process, the machinery that regulates CME in plants is largely unknown. In mammals, the heterotetrameric ADAPTOR PROTEIN COMPLEX-2 (AP-2) is required for the formation of clathrin-coated vesicles at the plasma membrane (PM). Although the existence of AP-2 has been predicted in Arabidopsis thaliana, the biochemistry and functionality of the complex is still uncharacterized. Here, we identified all the subunits of the Arabidopsis AP-2 by tandem affinity purification and found that one of the large AP-2 subunits, AP2A1, localized at the PM and interacted with clathrin. Furthermore, endocytosis of the leucine-rich repeat receptor kinase, BRASSINOSTEROID INSENSITIVE1 (BRI1), was shown to depend on AP-2. Knockdown of the two Arabidopsis AP2A genes or overexpression of a dominant-negative version of the medium AP-2 subunit, AP2M, impaired BRI1 endocytosis and enhanced the brassinosteroid signaling. Our data reveal that the CME machinery in Arabidopsis is evolutionarily conserved and that AP-2 functions in receptor-mediated endocytosis.
- Published
- 2013
130. Arabidopsis μ-adaptin subunit AP1M of adaptor protein complex 1 mediates late secretory and vacuolar traffic and is required for growth
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Kyungyoung Song, York-Dieter Stierhof, Misoon Park, Inhwan Hwang, Ulrike Mayer, Ilka Reichardt, Gerd Jürgens, and Hyeran Kim
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Cell division ,Protein subunit ,Adaptor Protein Complex 1 ,Arabidopsis ,Golgi Apparatus ,Biology ,Adaptor Protein Complex alpha Subunits ,Microscopy, Electron, Transmission ,Adaptor Protein Complex gamma Subunits ,Interphase ,Secretory pathway ,Cytokinesis ,Multidisciplinary ,Arabidopsis Proteins ,Secretory Vesicles ,Cell Membrane ,Signal transducing adaptor protein ,Biological Sciences ,Adaptor Protein Complex mu Subunits ,Cell biology ,Mutagenesis, Insertional ,Protein Transport ,Vacuoles ,trans-Golgi Network - Abstract
Adaptor protein (AP) complexes are the predominant coat proteins of membrane vesicles in post-Golgi trafficking of mammalian cells. Each AP complex contains a specific medium subunit, μ-adaptin, that selects cargo proteins bearing sequence-specific sorting motifs. Much less is known about the AP complexes and their μ subunits in plants. Because of uncertain homology, the μ-adaptins of Arabidopsis have been designated muA through muD [Happel et al. (2004) Plant J 37(5):678–693]. Furthermore, only muD has been assigned to a specific AP complex, AP-3, involved in Golgi-vacuolar trafficking [Niihama et al. (2009) Plant Cell Physiol 50(12):2057–2068, Zwiewka et al. (2011) Cell Res 21(12):1711–1722, and Wolfenstetter et al. (2012) Plant Cell 24(1):215–232]. In contrast, the μ subunit of neither the post-Golgi trafficking AP-1 complex nor the endocytic AP-2 complex has been identified. Here, we report the functional analysis of redundant AP-1 μ-adaptins AP1M1 (also known as muB1) and AP1M2 (also known as muB2). Coimmunoprecipitation revealed that both AP1M2 and its less strongly expressed isoform AP1M1 are complexed with the large subunit γ-adaptin of AP-1. In addition, AP1M2 was localized at or near the trans -Golgi network. Knockout mutations of AP1M2 impaired pollen function and arrested plant growth whereas the ap1m1 ap1m2 double mutant was nearly pollen-lethal. At the cellular level, the absence of AP1M2 entailed inhibition of multiple trafficking pathways from the trans -Golgi network to the vacuole and to the plasma membrane in interphase and to the plane of cell division in cytokinesis. Thus, AP-1 is crucial in post-Golgi trafficking in plant cells and required for cell division and plant growth.
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- 2013
131. Direct Targeting of Proteins from the Cytosol to Organelles: The ER versus Endosymbiotic Organelles
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Dae Heon Kim and Inhwan Hwang
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Signal peptide ,Endoplasmic reticulum ,food and beverages ,Cell Biology ,Biology ,medicine.disease_cause ,Biochemistry ,Membrane contact site ,Transport protein ,Cell biology ,Structural Biology ,Cytoplasm ,Organelle ,Protein targeting ,Genetics ,medicine ,Molecular Biology ,Peroxisomal targeting signal - Abstract
In eukaryotic cells consisting of many different types of organelles, targeting of organellar proteins is one of the most fundamental cellular processes. Proteins belonging to the endoplasmic reticulum (ER), chloroplasts and mitochondria are targeted individually from the cytosol to their cognate organelles. As the targeting to these organelles occurs in the cytosol during or after translation, the most crucial aspect is how specific targeting to these three organelles can be achieved without interfering with other targeting pathways. For these organelles, multiple mechanisms are used for targeting proteins, but the exact mechanism used depends on the type of protein and organelle, the location of targeting signals in the protein and the location of the protein in the organelle. In this review, we discuss the various mechanisms involved in protein targeting to the ER, chloroplasts and mitochondria, and how the targeting specificity is determined for these organelles in plant cells.
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- 2013
132. Recruitment of Arf1-GDP to Golgi by Glo3p-Type ArfGAPs Is Crucial for Golgi Maintenance and Plant Growth
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Kwan Yong Choi, David Robinson, Mihue Jang, Inhwan Hwang, Yongjik Lee, Myounghui Lee, Kyungyoung Song, Myung Ki Min, and Junho Lee
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Physiology ,Blotting, Western ,Green Fluorescent Proteins ,Molecular Sequence Data ,Arabidopsis ,Golgi Apparatus ,Plant Science ,Plasma protein binding ,Biology ,Guanosine Diphosphate ,symbols.namesake ,Microscopy, Electron, Transmission ,RNA interference ,Organelle ,Fluorescence Resonance Energy Transfer ,Genetics ,Arabidopsis thaliana ,Amino Acid Sequence ,Peptide sequence ,Phylogeny ,Microscopy, Confocal ,Sequence Homology, Amino Acid ,Arabidopsis Proteins ,Protoplasts ,GTPase-Activating Proteins ,Cell Biology ,Golgi apparatus ,Plants, Genetically Modified ,biology.organism_classification ,Cell biology ,Protein Transport ,Cytosol ,Biochemistry ,Mutation ,Vacuoles ,symbols ,ADP-Ribosylation Factor 1 ,RNA Interference ,Protein Binding - Abstract
ADP-ribosylation factor1 (Arf1), a member of the small GTP-binding proteins, plays a pivotal role in protein trafficking to multiple organelles. In its GDP-bound form, Arf1 is recruited from the cytosol to organelle membranes, where it functions in vesicle-mediated protein trafficking. However, the mechanism of Arf1-GDP recruitment remains unknown. Here, we provide evidence that two Glo3p-type Arf GTPase-activating proteins (ArfGAPs), ArfGAP domain8 (AGD8) and AGD9, are involved in the recruitment of Arf1-GDP to the Golgi apparatus in Arabidopsis (Arabidopsis thaliana). RNA interference plants expressing low levels of AGD8 and AGD9 exhibited abnormal Golgi morphology, inhibition of protein trafficking, and arrest of plant growth and development. In RNA interference plants, Arf1 was poorly recruited to the Golgi apparatus. Conversely, high levels of AGD8 and AGD9 induced Arf1 accumulation at the Golgi and suppressed Golgi disruption and inhibition of vacuolar trafficking that was caused by overexpression of AGD7. Based on these results, we propose that the Glo3p-type ArfGAPs AGD8 and AGD9 recruit Arf1-GDP from the cytosol to the Golgi for Arf1-mediated protein trafficking, which is essential for plant development and growth.
- Published
- 2012
133. Mitochondrial Targeting of the Arabidopsis F1-ATPase γ-Subunit via Multiple Compensatory and Synergistic Presequence Motifs
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Goeun Lee, Yun-Joo Yoo, Sumin Lee, Young Jun Oh, Inhwan Hwang, Yong Jik Lee, Dong-Wook Lee, Owen Duncan, and James Whelan
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Genetics ,Nuclear gene ,biology ,Arabidopsis Proteins ,C-terminus ,Immunoblotting ,Molecular Sequence Data ,Arabidopsis ,Cell Biology ,Plant Science ,Mitochondrion ,biology.organism_classification ,Immunohistochemistry ,Polymerase Chain Reaction ,Mitochondria ,Cell biology ,Green fluorescent protein ,Proton-Translocating ATPases ,Cytosol ,Arabidopsis thaliana ,Sequence motif ,Research Articles - Abstract
The majority of mitochondrial proteins are encoded in the nuclear genome and imported into mitochondria posttranslationally from the cytosol. An N-terminal presequence functions as the signal for the import of mitochondrial proteins. However, the functional information in the presequence remains elusive. This study reports the identification of critical sequence motifs from the presequence of Arabidopsis thaliana F1-ATPase γ-subunit (pFAγ). pFAγ was divided into six 10–amino acid segments, designated P1 to P6 from the N to the C terminus, each of which was further divided into two 5–amino acid subdivisions. These P segments and their subdivisions were substituted with Ala residues and fused to green fluorescent protein (GFP). Protoplast targeting experiments using these GFP constructs revealed that pFAγ contains several functional sequence motifs that are dispersed throughout the presequence. The sequence motifs DQEEG (P4a) and VVRNR (P5b) were involved in translocation across the mitochondrial membranes. The sequence motifs IAARP (P2b) and IAAIR (P3a) participated in binding to mitochondria. The sequence motifs RLLPS (P2a) and SISTQ (P5a) assisted in pulling proteins into the matrix, and the sequence motif IAARP (P2b) functioned in Tom20-dependent import. In addition, these sequence motifs exhibit complex relationships, including synergistic functions. Thus, multiple sequence motifs dispersed throughout the presequence are proposed to function cooperatively during protein import into mitochondria.
- Published
- 2012
134. Characterization of Glycosylphosphatidylinositol-Anchored Lipid Transfer Protein 2 (LTPG2) and Overlapping Function between LTPG/LTPG1 and LTPG2 in Cuticular Wax Export or Accumulation in Arabidopsis thaliana
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Mi Chung Suh, Myung Ki Min, Saet Buyl Lee, Hyojin Kim, Hae Jin Kim, and Inhwan Hwang
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Physiology ,Cuticle ,Molecular Sequence Data ,Mutant ,Arabidopsis ,Plant Science ,Biology ,Fatty Acid-Binding Proteins ,Plant Epidermis ,Cell wall ,Gene Knockout Techniques ,Gene Expression Regulation, Plant ,Arabidopsis thaliana ,Amino Acid Sequence ,Wax ,integumentary system ,Epidermis (botany) ,Arabidopsis Proteins ,Cell Membrane ,Cell Biology ,General Medicine ,biology.organism_classification ,Cell biology ,DNA-Binding Proteins ,Plant Leaves ,Biochemistry ,Waxes ,visual_art ,Mutation ,visual_art.visual_art_medium ,Silique ,Carrier Proteins ,Transcription Factors - Abstract
Cuticular waxes are synthesized by the extensive export of intracellular lipids from epidermal cells. However, it is still not known how hydrophobic cuticular lipids are exported to the plant surface through the hydrophilic cell wall. The LTPG2 gene was isolated based on Arabidopsis microarray analysis; this gene is predominantly expressed in stem epidermal peels as compared with in stems. The expression of LTPG2 transcripts was observed in various organs, including stem epidermis and silique walls. The composition of the cuticular wax was significantly altered in the stems and siliques of the ltpg2 mutant and ltpg1 ltpg2 double mutant. In particular, the reduced level of the C29 alkane, which is the major component of cuticular waxes in ltpg1 ltpg2 stems and siliques, was similar to the sum of reduced values of either parent. The total cuticular wax load was reduced by approximately 13% and 20% in both ltpg2 and ltpg1 ltpg2 siliques, respectively, and by approximately 14% in ltpg1 ltpg2 stems when compared with the wild-type. Similarly, severe alterations in the cuticular layer structure of epidermal cells of ltpg2 and ltpg1 ltpg2 stems and silique walls were observed. In tobacco epidermal cells, intracellular trafficking of the fluorescent LTPG/LTPG1 and LTPG2 to the plasma membrane was prevented by a dominant-negative mutant form of ADP-ribosylation factor 1, ARF1(T31N). Taken together, these results indicate that LTPG2 is functionally overlapped with LTPG/LTPG1 during cuticular wax export or accumulation and LTPG/LTPG1 and LTPG2 are targeted to the plasma membrane via the vesicular trafficking system.
- Published
- 2012
135. An A/ENTH Domain-Containing Protein Functions as an Adaptor for Clathrin-Coated Vesicles on the Growing Cell Plate in Arabidopsis Root Cells
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Dae Heon Kim, Wonhwa Cho, Mihue Jang, Soo Youn Kim, Inhwan Hwang, Goeun Lee, Kyungyoung Song, Yongjik Lee, and Gil Je Lee
- Subjects
Epsin ,biology ,Physiology ,Plant Science ,Cell plate ,Phragmoplast ,Clathrin ,Cell biology ,Genetics ,biology.protein ,Ap180 ,Clathrin adaptor proteins ,ENTH domain ,Cytokinesis - Abstract
Cytokinesis is the process of partitioning the cytoplasm of a dividing cell, thereby completing mitosis. Cytokinesis in the plant cell is achieved by the formation of a new cell wall between daughter nuclei using components carried in Golgi-derived vesicles that accumulate at the midplane of the phragmoplast and fuse to form the cell plate. Proteins that play major roles in the development of the cell plate in plant cells are not well defined. Here, we report that an AP180 amino-terminal homology/epsin amino-terminal homology domain-containing protein from Arabidopsis (Arabidopsis thaliana) is involved in clathrin-coated vesicle formation from the cell plate. Arabidopsis Epsin-like Clathrin Adaptor1 (AtECA1; At2g01600) and its homologous proteins AtECA2 and AtECA4 localize to the growing cell plate in cells undergoing cytokinesis and also to the plasma membrane and endosomes in nondividing cells. AtECA1 (At2g01600) does not localize to nascent cell plates but localizes at higher levels to expanding cell plates even after the cell plate fuses with the parental plasma membrane. The temporal and spatial localization patterns of AtECA1 overlap most closely with those of the clathrin light chain. In vitro protein interaction assays revealed that AtECA1 binds to the clathrin H chain via its carboxyl-terminal domain. These results suggest that these AP180 amino-terminal homology/epsin amino-terminal homology domain-containing proteins, AtECA1, AtECA2, and AtECA4, may function as adaptors of clathrin-coated vesicles budding from the cell plate.
- Published
- 2012
136. Overexpression of an Arabidopsis β-glucosidase gene enhances drought resistance with dwarf phenotype in creeping bentgrass
- Author
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Yun-Jeong Han, Ok-Jin Hwang, Kyu-Chang Cho, Inhwan Hwang, Ah-Young Shin, Jeong-Il Kim, and Yun Sung Choi
- Subjects
Transgene ,Drought tolerance ,Arabidopsis ,Plant Science ,Genetically modified crops ,Genes, Plant ,Agrostis ,chemistry.chemical_compound ,Transformation, Genetic ,Botany ,Northern blot ,Abscisic acid ,biology ,Arabidopsis Proteins ,Abiotic stress ,beta-Glucosidase ,fungi ,food and beverages ,General Medicine ,Plants, Genetically Modified ,biology.organism_classification ,Adaptation, Physiological ,Droughts ,Plant Leaves ,Transformation (genetics) ,Phenotype ,chemistry ,Agronomy and Crop Science ,Abscisic Acid ,Herbicide Resistance - Abstract
An Arabidopsis β-glucosidase, AtBG1 is known to hydrolyze glucose-conjugated, biologically inactive abscisic acid (ABA) to produce active ABA, which increases the level of ABA in plants. Since an increase of ABA in plants confers tolerance against abiotic stress such as drought, we introduced the pCAMBIA3301 vector harboring the AtBG1 gene into creeping bentgrass through Agrobacterium-mediated transformation. After transformation, putative transgenic plants were selected using the BASTA resistance assay at a concentration of 0.8 %. Genomic integration of the AtBG1 gene was confirmed by genomic PCR and Southern blot analysis, and gene expression was validated by Northern blot and Western blot analyses. Interestingly, the transgenic bentgrass plants overexpressing AtBG1 had a dwarf phenotype with reduced growth rates when compared to wild-type creeping bentgrass. In addition, the transgenic plants accumulated higher ABA levels and displayed enhanced drought tolerance. These results suggest that the expression of AtBG1 in plants induces the accumulation of higher ABA levels, which results in the formation of dwarf creeping bentgrass and enhances the survival in water-limiting environments. Key message We used an Arabidopsis β-glucosidase AtBG1 to engineer a crop with elevated active ABA levels, and developed transgenic creeping bentgrass with enhanced drought tolerance and dwarf phenotype.
- Published
- 2012
137. A Vacuolar β-Glucosidase Homolog That Possesses Glucose-Conjugated Abscisic Acid Hydrolyzing Activity Plays an Important Role in Osmotic Stress Responses in Arabidopsis
- Author
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Mitsunori Seo, Dae Heon Kim, Yuri Kanno, Soo Youn Kim, Jing Bo Jin, Dae-Jin Yun, Zheng-Yi Xu, Ray A. Bressan, Jae Cheol Jeong, Ting Dong, Inhwan Hwang, and Kwang Hee Lee
- Subjects
Osmosis ,Osmotic shock ,Mutant ,Arabidopsis ,Plant Science ,Vacuole ,Sodium Chloride ,chemistry.chemical_compound ,Biosynthesis ,Gene Expression Regulation, Plant ,Cellulases ,Arabidopsis thaliana ,Desiccation ,Abscisic acid ,Research Articles ,Regulation of gene expression ,biology ,Arabidopsis Proteins ,organic chemicals ,fungi ,food and beverages ,Cell Biology ,biology.organism_classification ,chemistry ,Biochemistry ,Vacuoles ,Abscisic Acid - Abstract
The phytohormone abscisic acid (ABA) plays a critical role in various physiological processes, including adaptation to abiotic stresses. In Arabidopsis thaliana, ABA levels are increased both through de novo biosynthesis and via β-glucosidase homolog1 (BG1)-mediated hydrolysis of Glc-conjugated ABA (ABA-GE). However, it is not known how many different β-glucosidase proteins produce ABA from ABA-GE and how the multiple ABA production pathways are coordinated to increase ABA levels. Here, we report that a previously undiscovered β-glucosidase homolog, BG2, produced ABA by hydrolyzing ABA-GE and plays a role in osmotic stress response. BG2 localized to the vacuole as a high molecular weight complex and accumulated to high levels under dehydration stress. BG2 hydrolyzed ABA-GE to ABA in vitro. In addition, BG2 increased ABA levels in protoplasts upon application of exogenous ABA-GE. Overexpression of BG2 rescued the bg1 mutant phenotype, as observed for the overexpression of NCED3 in bg1 mutants. Multiple Arabidopsis bg2 alleles with a T-DNA insertion in BG2 were more sensitive to dehydration and NaCl stress, whereas BG2 overexpression resulted in enhanced resistance to dehydration and NaCl stress. Based on these observations, we propose that, in addition to the de novo biosynthesis, ABA is produced in multiple organelles by organelle-specific β-glucosidases in response to abiotic stresses.
- Published
- 2012
138. Secretory Pathway Research: The More Experimental Systems the Better
- Author
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Jaideep Mathur, Jean-Marc Neuhaus, Fernando Aniento, Lorenzo Frigerio, Jürgen Denecke, David G. Robinson, Chris Hawes, and Inhwan Hwang
- Subjects
Protein function ,Molecular cell biology ,Secretory Pathway ,Protoplasts ,Research ,fungi ,Mutant ,food and beverages ,Biological Transport ,Cell Biology ,Plant Science ,Biology ,Subcellular localization ,Cell biology ,Plant Leaves ,Perspective ,Gene expression ,Ectopic expression ,Gene ,Secretory pathway ,Fluorescent Dyes - Abstract
Transient gene expression, in plant protoplasts or specific plant tissues, is a key technique in plant molecular cell biology, aimed at exploring gene products and their modifications to examine functional subdomains, their interactions with other biomolecules, and their subcellular localization. Here, we highlight some of the major advantages and potential pitfalls of the most commonly used transient gene expression models and illustrate how ectopic expression and the use of dominant mutants can provide insights into protein function.
- Published
- 2012
139. An Arabidopsis Prenylated Rab Acceptor 1 Isoform, AtPRA1.B6, Displays Differential Inhibitory Effects on Anterograde Trafficking of Proteins at the Endoplasmic Reticulum
- Author
-
Myoung Hui Lee, Chanjin Jung, Yongjik Lee, Junho Lee, Inhwan Hwang, and Soo Youn Kim
- Subjects
Physiology ,Endoplasmic reticulum ,Plant Science ,Biology ,Cell biology ,Green fluorescent protein ,Transport protein ,Secretory protein ,Membrane protein ,Biochemistry ,Genetics ,Endomembrane system ,Rab ,Secretory pathway - Abstract
Prenylated Rab acceptors (PRAs), members of the Ypt-interacting protein family of small membrane proteins, are thought to aid the targeting of prenylated Rabs to their respective endomembrane compartments. In plants, the Arabidopsis (Arabidopsis thaliana) PRA1 family contains 19 members that display varying degrees of sequence homology to animal PRA1 and localize to the endoplasmic reticulum (ER) and/or endosomes. However, the exact role of these proteins remains to be fully characterized. In this study, the effect of AtPRA1.B6, a member of the AtPRA1 family, on the anterograde trafficking of proteins targeted to various endomembrane compartments was investigated. High levels of AtPRA1.B6 resulted in differential inhibition of coat protein complex II vesicle-mediated anterograde trafficking. The trafficking of the vacuolar proteins sporamin:GFP (for green fluorescent protein) and AALP:GFP, the secretory protein invertase:GFP, and the plasma membrane proteins PMP:GFP and H+-ATPase:GFP was inhibited in a dose-dependent manner, while the trafficking of the Golgi-localized proteins ST:GFP and KAM1(ƊC):mRFP was not affected. Conversely, in RNA interference plants displaying lower levels of AtPRA1.B6 transcripts, the trafficking efficiency of sporamin:GFP and AALP:GFP to the vacuole was increased. Localization and N-glycan pattern analyses of cargo proteins revealed that AtPRA1.B6-mediated inhibition of anterograde trafficking occurs at the ER. In addition, AtPRA1.B6 levels were controlled by cellular processes, including 26S proteasome-mediated proteolysis. Based on these results, we propose that AtPRA1.B6 is a negative regulator of coat protein complex II vesicle-mediated anterograde trafficking for a subset of proteins at the ER.
- Published
- 2011
140. Small Heat Shock Protein Hsp17.8 Functions as an AKR2A Cofactor in the Targeting of Chloroplast Outer Membrane Proteins in Arabidopsis
- Author
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Junho Lee, Zheng-Yi Xu, Inhwan Hwang, Yun Jeong Na, Eun-Ju Sohn, Dae Heon Kim, and Yun-Joo Yoo
- Subjects
HSPA12A ,Physiology ,fungi ,food and beverages ,Plant Science ,Chloroplast outer membrane ,Biology ,medicine.disease_cause ,Fusion protein ,Cell biology ,Membrane protein ,Biochemistry ,Heat shock protein ,Protein targeting ,Genetics ,medicine ,Ankyrin repeat ,Plastid - Abstract
Plastid proteins that are encoded by the nuclear genome and synthesized in the cytosol undergo posttranslational targeting to plastids. Ankyrin repeat protein 2A (AKR2A) and AKR2B were recently shown to be involved in the targeting of proteins to the plastid outer envelope. However, it remains unknown whether other factors are involved in this process. In this study, we investigated a factor involved in AKR2A-mediated protein targeting to chloroplasts in Arabidopsis (Arabidopsis thaliana). Hsp17.8, a member of the class I (CI) cytosolic small heat shock proteins (sHsps), was identified in interactions with AKR2A. The interaction between Hsp17.8 and AKR2A was further confirmed by coimmunoprecipitation experiments. The carboxyl-terminal ankyrin repeat domain of AKR2A was responsible for AKR2A binding to Hsp17.8. Other CI cytosolic sHsps also interact with AKR2A to varying degrees. Additionally, Hsp17.8 binds to chloroplasts in vitro and enhances AKR2A binding to chloroplasts. HSP17.8 was expressed under normal growth conditions, and its expression increased after heat shock. Hsp17.8 exists as a dimer under normal physiological conditions, and it is converted to high oligomeric complexes, ranging from 240 kD to greater than 480 kD, after heat shock. High levels of Hsp17.8 together with AKR2A resulted in increased plastid targeting of Outer Envelope Protein7 (OEP7), a plastid outer envelope protein expressed as a green fluorescent protein fusion protein. In contrast, artificial microRNA suppression of HSP17.8 and closely related CI cytosolic sHSPs in protoplasts resulted in a reduction of OEP7:green fluorescent protein targeting to plastids. Based on these data, we propose that Hsp17.8 functions as an AKR2A cofactor in targeting membrane proteins to plastid outer membranes under normal physiological conditions.
- Published
- 2011
141. Both the Hydrophobicity and a Positively Charged Region Flanking the C-Terminal Region of the Transmembrane Domain of Signal-Anchored Proteins Play Critical Roles in Determining Their Targeting Specificity to the Endoplasmic Reticulum or Endosymbiotic Organelles in Arabidopsis Cells
- Author
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Jin-Ho Kim, Sumin Lee, Hyun-Kyung Lee, Sanguk Kim, Dae Heon Kim, Inhwan Hwang, and Junho Lee
- Subjects
Chloroplasts ,Recombinant Fusion Proteins ,Green Fluorescent Proteins ,Molecular Sequence Data ,Arabidopsis ,Plant Science ,Protein Sorting Signals ,Endoplasmic Reticulum ,Organelle ,Amino Acid Sequence ,Symbiosis ,Research Articles ,biology ,Arabidopsis Proteins ,Protoplasts ,Endoplasmic reticulum ,Cell Biology ,Plant cell ,biology.organism_classification ,Mitochondria ,Protein Structure, Tertiary ,Cell biology ,Protein Transport ,Transmembrane domain ,Membrane protein ,Mutation ,Cell fractionation ,Hydrophobic and Hydrophilic Interactions ,Biogenesis ,Protein Binding ,Subcellular Fractions - Abstract
Proteins localized to various cellular and subcellular membranes play pivotal roles in numerous cellular activities. Accordingly, in eukaryotic cells, the biogenesis of organellar proteins is an essential process requiring their correct localization among various cellular and subcellular membranes. Localization of these proteins is determined by either cotranslational or posttranslational mechanisms, depending on the final destination. However, it is not fully understood how the targeting specificity of membrane proteins is determined in plant cells. Here, we investigate the mechanism by which signal-anchored (SA) proteins are differentially targeted to the endoplasmic reticulum (ER) or endosymbiotic organelles using in vivo targeting, subcellular fractionation, and bioinformatics approaches. For targeting SA proteins to endosymbiotic organelles, the C-terminal positively charged region (CPR) flanking the transmembrane domain (TMD) is necessary but not sufficient. The hydrophobicity of the TMD in CPR-containing proteins also plays a critical role in determining targeting specificity; TMDs with a hydrophobicity value >0.4 on the Wimley and White scale are targeted primarily to the ER, whereas TMDs with lower values are targeted to endosymbiotic organelles. Based on these data, we propose that the CPR and the hydrophobicity of the TMD play a critical role in determining the targeting specificity between the ER and endosymbiotic organelles.
- Published
- 2011
142. Homomeric Interaction of AtVSR1 Is Essential for Its Function as a Vacuolar Sorting Receptor
- Author
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Yansong Miao, Mihue Jang, Liwen Jiang, Hyangju Kang, Hyeran Kim, Inhwan Hwang, and Jeong Ho Chang
- Subjects
Physiology ,Green Fluorescent Proteins ,Molecular Sequence Data ,Arabidopsis ,Receptors, Cell Surface ,Plant Science ,Vacuole ,Plasma protein binding ,Structure-Activity Relationship ,Protein structure ,Genetics ,Protein Isoforms ,Homomeric ,Amino Acid Sequence ,biology ,Arabidopsis Proteins ,Clathrin-Coated Vesicles ,Plants, Genetically Modified ,biology.organism_classification ,Clathrin ,Transmembrane protein ,Protein Structure, Tertiary ,Transport protein ,Cell biology ,Molecular Weight ,Protein Transport ,Transmembrane domain ,Amino Acid Substitution ,Cell Biology and Signal Transduction ,Vacuoles ,Mutant Proteins ,Protein Multimerization ,Protein Binding ,trans-Golgi Network - Abstract
Vacuolar sorting receptors, BP80/VSRs, play a critical role in vacuolar trafficking of soluble proteins in plant cells. However, the mechanism of action of BP80 is not well understood. Here, we investigate the action mechanism of AtVSR1, a member of BP80 proteins in Arabidopsis (Arabidopsis thaliana), in vacuolar trafficking. AtVSR1 exists as multiple forms, including a high molecular mass homomeric complex in vivo. Both the transmembrane and carboxyl-terminal cytoplasmic domains of AtVSR1 are necessary for the homomeric interaction. The carboxyl-terminal cytoplasmic domain contains specific sequence information, whereas the transmembrane domain has a structural role in the homomeric interaction. In protoplasts, an AtVSR1 mutant, C2A, that contained alanine substitution of the region involved in the homomeric interaction, was defective in trafficking to the prevacuolar compartment and localized primarily to the trans-Golgi network. In addition, overexpression of C2A, but not wild-type AtVSR1, inhibited trafficking of soluble proteins to the vacuole and caused their secretion into the medium. Furthermore, C2A:hemagglutinin in transgenic plants interfered with the homomeric interaction of endogenous AtVSR1 and inhibited vacuolar trafficking of sporamin:green fluorescent protein. These data suggest that homomeric interaction of AtVSR1 is critical for its function as a vacuolar sorting receptor.
- Published
- 2010
143. Activation of glucosidase via stress-induced polymerization rapidly increases active pools of abscisic acid
- Author
-
Kwang Hee Lee, Hai Lan Piao, Iidoo Hwang, June M. Kwak, In-Jung Lee, and Inhwan Hwang
- Subjects
Abscisic acid -- Research ,Polymerization -- Research ,Biological sciences - Abstract
A study report about AtBG1, a [beta]-glucosidase, which hydrolyzes glucose-conjugated, biologically inactive abscisic acid (ABA) to produce active ABA, is presented. The report suggests that the activation of inactive ABA pools by polymerized AtBG1 is a mechanism by which plants rapidly adjust ABA levels and respond to changing environmental cues.
- Published
- 2006
144. Silencing of NbCEP1 encoding a chloroplast envelope protein containing 15 leucine-rich-repeats disrupts chloroplast biogenesis in Nicotiana benthamiana
- Author
-
Young Eun Jeon, A-Reum Hwang, Hyun Sook Pai, and Inhwan Hwang
- Subjects
Chloroplasts ,Molecular Sequence Data ,Nicotiana benthamiana ,Leucine-Rich Repeat Proteins ,Chloroplast membrane ,Gene Expression Regulation, Plant ,Transit Peptide ,Tobacco ,Amino Acid Sequence ,Gene Silencing ,RNA, Messenger ,Molecular Biology ,Plant Proteins ,biology ,fungi ,Proteins ,food and beverages ,Intracellular Membranes ,Cell Biology ,General Medicine ,biology.organism_classification ,Fusion protein ,Molecular biology ,Mitochondria ,Cell biology ,N-terminus ,Chloroplast ,Protein Transport ,Chloroplast stroma ,Phenotype ,Chloroplast DNA ,Sequence Alignment ,Subcellular Fractions - Abstract
We characterized the physiological functions of Nicotiana benthamiana Chloroplast Envelope Protein 1 (NbCEP1) in Nicotiana benthamiana. NbCEP1 contains a chloroplast transit peptide and a single transmembrane domain at the N terminus, and most of its protein coding region is comprised of 15 leucine-rich-repeats (LRRs). The NbCEP1 gene is expressed in both aerial and underground plant tissues, and is induced by light. A GFP fusion protein of full length NbCEP1 was targeted to the chloroplast envelope and co-localized with OEP7:RFP, a marker protein for the chloroplast envelope. A fusion protein consisting of GFP and the NbCEP1 transit peptide mainly localized in the chloroplast stroma. Reduction of NbCEP1 expression by virus-induced gene silencing resulted in a leaf yellowing phenotype without much affecting overall plant growth. At the cellular level, depletion of NbCEP1 severely influenced chloroplast development, reducing both the number and size of the chloroplasts. Interestingly, mitochondrial development was also impaired, possibly an indirect effect of chloroplast ablation. A deficiency in NbCEP1 activity decreased the chlorophyll and carotenoid levels. Our results suggest that NbCEP1 plays a critical function, possibly through protein-protein interactions mediated by its LRRs, in chloroplast development in N. benthamiana.
- Published
- 2009
145. Heat Shock Protein Cognate 70-4 and an E3 Ubiquitin Ligase, CHIP, Mediate Plastid-Destined Precursor Degradation through the Ubiquitin-26S Proteasome System inArabidopsis
- Author
-
Gerd Jürgens, Sookjin Lee, Yongjik Lee, York-Dieter Stierhof, Ulrike Mayer, Dong-Wook Lee, Sumin Lee, and Inhwan Hwang
- Subjects
DNA, Bacterial ,Proteasome Endopeptidase Complex ,Protein Folding ,Chloroplasts ,animal structures ,Ubiquitin-Protein Ligases ,Arabidopsis ,macromolecular substances ,Plant Science ,Protein degradation ,Ubiquitin ,Heat shock protein ,Protein Precursors ,Plastid ,Phylogeny ,Research Articles ,Oligonucleotide Array Sequence Analysis ,biology ,Arabidopsis Proteins ,fungi ,HSC70 Heat-Shock Proteins ,food and beverages ,Cell Biology ,Plants, Genetically Modified ,biology.organism_classification ,Ubiquitin ligase ,Mutagenesis, Insertional ,Cytosol ,Biochemistry ,Proteasome ,RNA, Plant ,embryonic structures ,biology.protein ,Protein Processing, Post-Translational - Abstract
Plastid-targeted proteins pass through the cytosol as unfolded precursors. If proteins accumulate in the cytosol, they can form nonspecific aggregates that cause severe cellular damage. Here, we demonstrate that high levels of plastid precursors are degraded through the ubiquitin-proteasome system (UPS) in Arabidopsis thaliana cells. The cytosolic heat shock protein cognate 70-4 (Hsc70-4) and E3 ligase carboxy terminus of Hsc70-interacting protein (CHIP) were highly induced in plastid protein import2 plants, which had a T-DNA insertion at Toc159 and showed an albino phenotype and a severe defect in protein import into chloroplasts. Hsc70-4 and CHIP together mediated plastid precursor degradation when import-defective chloroplast-targeted reporter proteins were transiently expressed in protoplasts. Hsc70-4 recognized specific sequence motifs in transit peptides and thereby led to precursor degradation through the UPS. CHIP, which interacted with Hsc70-4, functioned as an E3 ligase in the Hsc70-4–mediated protein degradation. The physiological role of Hsc70-4 was confirmed by analyzing Hsc70-4 RNA interfernce plants in an hsc70-1 mutant background. Plants with lower Hsc70 levels exhibited abnormal embryogenesis, resulting in defective seedlings that displayed high levels of reactive oxygen species and monoubiquitinated Lhcb4 precursors. We propose that Hsc70-4 and CHIP mediate plastid-destined precursor degradation to prevent cytosolic precursor accumulation and thereby play a critical role in embryogenesis.
- Published
- 2009
146. Multiple Sequence Motifs in the Rubisco Small Subunit Transit Peptide Independently Contribute to Toc159-Dependent Import of Proteins into Chloroplasts
- Author
-
Dong-Wook Lee, Sumin Lee, Inhwan Hwang, and Young Jun Oh
- Subjects
chemistry.chemical_classification ,Nuclear gene ,Physiology ,Mutant ,food and beverages ,Plant Science ,Biology ,Amino acid ,Serine ,Chloroplast ,chemistry ,Biochemistry ,Transit Peptide ,Genetics ,Plastid ,Sequence motif - Abstract
A large number of plastid proteins encoded by the nuclear genome are posttranslationally imported into plastids by at least two distinct mechanisms: the Toc159-dependent and Toc132/Toc120-dependent pathways. Light-induced photosynthetic proteins are imported through the Toc159-dependent pathway, whereas constitutive housekeeping plastid proteins are imported into plastids through the Toc132/Toc120 pathway. However, it remains unknown which features of the plastid protein transit peptide (TP) determine the import pathway. We have discovered sequence elements of the Rubisco small subunit TP (RbcS-tp) that play a role in determining import through the Toc159-dependent pathway in vivo. We generated multiple hybrid mutants using the RbcS-tp and the E1α-subunit of pyruvate dehydrogenase TP (E1α-tp) as representative peptides mediating import through the Toc159-dependent and Toc159-independent pathways, respectively. Import experiments using these hybrid mutants in wild-type and ppi2 mutant protoplasts revealed that multiple sequence motifs in the RbcS-tp independently contribute to Toc159-dependent protein import into chloroplasts. One of these motifs is the group of serine residues located in the N-terminal 12-amino acid segment and the other is the C-terminal T5 region of the RbcS-tp ranging from amino acid positions 41 to 49. Based on these findings, we propose that multiple sequence elements in the RbcS-tp contribute independently to Toc159-dependent import of proteins into chloroplasts.
- Published
- 2009
147. Disruption of Glycosylphosphatidylinositol-Anchored Lipid Transfer Protein Gene Altered Cuticular Lipid Composition, Increased Plastoglobules, and Enhanced Susceptibility to Infection by the Fungal Pathogen Alternaria brassicicola
- Author
-
Young Sam Go, Hyun Jong Bae, Sung Ho Cho, Ohkmae K. Park, Hong Joo Cho, Jong Ho Park, Inhwan Hwang, Dong Sook Lee, Saet Buyl Lee, and Mi Chung Suh
- Subjects
Alternaria brassicicola ,Epidermis (botany) ,Physiology ,Cuticle ,fungi ,Plant Science ,Cutin ,Vacuole ,Biology ,Vascular bundle ,biology.organism_classification ,Microbiology ,Cell biology ,Genetics ,Plant lipid transfer proteins ,Lipid Transport - Abstract
All aerial parts of vascular plants are covered with cuticular waxes, which are synthesized by extensive export of intracellular lipids from epidermal cells to the surface. Although it has been suggested that plant lipid transfer proteins (LTPs) are involved in cuticular lipid transport, the in planta evidence is still not clear. In this study, a glycosylphosphatidylinositol-anchored LTP (LTPG1) showing higher expression in epidermal peels of stems than in stems was identified from an Arabidopsis (Arabidopsis thaliana) genome-wide microarray analysis. The expression of LTPG1 was observed in various tissues, including the epidermis, stem cortex, vascular bundles, mesophyll cells, root tips, pollen, and early-developing seeds. LTPG1 was found to be localized in the plasma membrane. Disruption of the LTPG1 gene caused alterations of cuticular lipid composition, but no significant changes on total wax and cutin monomer loads were seen. The largest reduction (10 mass %) in the ltpg1 mutant was observed in the C29 alkane, which is the major component of cuticular waxes in the stems and siliques. The reduced content was overcome by increases of the C29 secondary alcohols and C29 ketone wax loads. The ultrastructure analysis of ltpg1 showed a more diffuse cuticular layer structure, protrusions of the cytoplasm into the vacuole in the epidermis, and an increase of plastoglobules in the stem cortex and leaf mesophyll cells. Furthermore, the ltpg1 mutant was more susceptible to infection by the fungus Alternaria brassicicola than the wild type. Taken together, these results indicated that LTPG1 contributed either directly or indirectly to cuticular lipid accumulation.
- Published
- 2009
148. Drought Stress-Induced Rma1H1, a RING Membrane-Anchor E3 Ubiquitin Ligase Homolog, Regulates Aquaporin Levels via Ubiquitination in TransgenicArabidopsisPlants
- Author
-
Zheng-Yi Xu, Woo Taek Kim, Seok Keun Cho, Ora Son, Inhwan Hwang, and Hyun-Kyung Lee
- Subjects
Proteasome Endopeptidase Complex ,Recombinant Fusion Proteins ,Ubiquitin-Protein Ligases ,Green Fluorescent Proteins ,Molecular Sequence Data ,Arabidopsis ,Aquaporin ,Plant Science ,Aquaporins ,Endoplasmic Reticulum ,Ubiquitin ,Stress, Physiological ,Arabidopsis thaliana ,Amino Acid Sequence ,Ligase activity ,Research Articles ,Plant Proteins ,biology ,Arabidopsis Proteins ,Endoplasmic reticulum ,fungi ,Cell Membrane ,Ubiquitination ,food and beverages ,Intracellular Membranes ,Cell Biology ,Plants, Genetically Modified ,biology.organism_classification ,Droughts ,Ubiquitin ligase ,Protein Transport ,Biochemistry ,Proteasome ,biology.protein ,lipids (amino acids, peptides, and proteins) ,Capsicum ,Sequence Alignment - Abstract
Ubiquitination is involved in a variety of biological processes, but the exact role of ubiquitination in abiotic responses is not clearly understood in higher plants. Here, we investigated Rma1H1, a hot pepper (Capsicum annuum) homolog of a human RING membrane-anchor 1 E3 ubiquitin (Ub) ligase. Bacterially expressed Rma1H1 displayed E3 Ub ligase activity in vitro. Rma1H1 was rapidly induced by various abiotic stresses, including dehydration, and its overexpression in transgenic Arabidopsis thaliana plants conferred strongly enhanced tolerance to drought stress. Colocalization experiments with marker proteins revealed that Rma1H1 resides in the endoplasmic reticulum (ER) membrane. Overexpression of Rma1H1 in Arabidopsis inhibited trafficking of an aquaporin isoform PIP2;1 from the ER to the plasma membrane and reduced PIP2;1 levels in protoplasts and transgenic plants. This Rma1H1-induced reduction of PIP2;1 was inhibited by MG132, an inhibitor of the 26S proteasome. Furthermore, Rma1H1 interacted with PIP2;1 in vitro and ubiquitinated it in vivo. Similar to Rma1H1, Rma1, an Arabidopsis homolog of Rma1H1, localized to the ER, and its overexpression reduced the PIP2;1 protein level and inhibited trafficking of PIP2;1 from the ER to the plasma membrane in protoplasts. In addition, reduced expression of Rma homologs resulted in the increased level of PIP2;1 in protoplasts. We propose that Rma1H1 and Rma1 play a critical role in the downregulation of plasma membrane aquaporin levels by inhibiting aquaporin trafficking to the plasma membrane and subsequent proteasomal degradation as a response to dehydration in transgenic Arabidopsis plants.
- Published
- 2009
149. Sorting and Anterograde Trafficking at the Golgi Apparatus: Figure 1
- Author
-
Inhwan Hwang
- Subjects
symbols.namesake ,Protein Sorting Signals ,Physiology ,Genetics ,symbols ,Signal transducing adaptor protein ,Cell migration ,Plant Science ,Golgi apparatus ,Biology ,Transport protein ,Cell biology - Abstract
In general, the basic principles of trafficking systems in plant cells appear to be similar to those in animal and yeast cells, indicating that trafficking mechanisms are highly conserved throughout all eukaryotes ([Jurgens, 2004][1]; [Vitale and Hinz, 2005][2]). However, there are still
- Published
- 2008
150. Isolation and characterization of multiple abundant lipid transfer protein isoforms in developing sesame (Sesamum indicum L.) seeds
- Author
-
Ah Mi Choi, Mi Chung Suh, Cheol-Goo Hur, Inhwan Hwang, Sung Ho Cho, and Saet Buyl Lee
- Subjects
Physiology ,Molecular Sequence Data ,Arabidopsis ,Plant Science ,Cutin ,Sesamum ,chemistry.chemical_compound ,Gene Expression Regulation, Plant ,Genetics ,Protein Isoforms ,Abscisic acid ,Phospholipids ,Plant Proteins ,Expressed Sequence Tags ,chemistry.chemical_classification ,Expressed sequence tag ,Base Sequence ,biology ,Protoplasts ,Fatty Acids ,food and beverages ,Blotting, Northern ,biology.organism_classification ,Amino acid ,chemistry ,Biochemistry ,Pedaliaceae ,Seeds ,Carrier Proteins ,Sequence Alignment ,Plant lipid transfer proteins - Abstract
Sesame ( Sesamum indicum ) is an important oilseed crop; approximately 50% of the seed dry weight is storage oil. In a previous report, developing sesame seed expressed sequence tags (ESTs) revealed that ESTs encoding lipid transfer protein (LTPs) were one of the most abundant groups of sesame ESTs. LTP functions in the transfer of wax or cutin monomers and in the defense response against pathogen attack. To study the biological role of the abundant LTP isoforms in developing seeds, 122 ESTs out of 3328 sesame ESTs were analyzed against Arabidopsis and rice proteome databases. LTP fraction, which was partially purified from developing sesame seeds, actively transferred fluorescent phospholipids and bound to fatty acids. Full-length cDNAs of five out of 21 LTP isoforms were isolated and named SiLTP1 – SiLTP5 . The predicted amino acid sequences of the five SiLTPs harbor typical characteristics of LTPs, including conserved arrangement of cysteine residues. Northern blot analysis revealed that the five SiLTP isoforms were most abundantly expressed in developing seeds, but were also detected in flower tissues. Also, SiLTP3 and SiLTP4 transcripts were expressed in leaves and seed-pot walls, respectively. In addition, SiLTP2 and SiLTP4 transcripts were significantly induced in 6-day-old sesame seedlings by application of NaCl, mannitol, and abscisic acid (ABA). Transient expression of green fluorescent protein (GFP)-fusion constructs in Arabidopsis protoplasts revealed that SiLTP1 and SiLTP2 were secreted by different pathways. Taken together, the abundant LTPs in developing sesame seeds are involved in lipid transfer into the extracellular matrix. Possible biological roles of SiLTPs related to organ-specific expression and abiotic stresses are discussed.
- Published
- 2008
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