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The neurotransmitter serotonin plays a pivotal role in mood and depression. It also acts as a vasoconstrictor within blood vessels and is the main neurotransmitter in the gastrointestinal system. In neurotransmission, released serotonin is taken up by serotonin transporters, which are principal targets of antidepressants and the psychostimulant, ecstasy. The investigation of serotonin transporters have relied almost exclusively on the use of radiolabeled serotonin in heterogenous end-point assays. Here we adapt the genetically encoded fluorescent biosensor, iSeroSnFR, to establish and validate the Serotonin (5-HT) Fluorescence Assay for Transport and Release (5-HT_FAsTR) for functional and pharmacological studies of serotonin transport and release. We demonstrate the applicability of the method for the study of a neuronal, high-affinity, low-capacity serotonin transporter (SERT) as well as an extraneuronal low-affinity, high-capacity organic cation transporter and mutants thereof. 5HT_FAsTR offers an accessible, versatile and reliable semi-homogenous assay format that only relies on a fluorescence plate reader for repeated, real-time measurements of serotonin influx and efflux. 5HT_FAsTR accelerates and democratizes functional characterization and pharmacological studies of serotonin transporters and genetic variants thereof in disease states such as depression, anxiety and ADHD.

• Renal proximal tubular epithelial cells (RPTECs) are the main targets of drug-induced kidney injury; however, no useful evaluation system is currently available. • We evaluated kidney injuries induced by 32 nephrotoxic drugs for up to 28 days using three-dimensional cultured RPTECs (3D-RPTECs) with enhanced expression of drug transporters. • The predictive performance of 3D-RPTECs for drug-induced kidney injury was better than conventional 2D-cultured RPTECs and HK-2 cells. • 3D-RPTECs are useful for in vitro evaluation of RPTEC injury by measuring intracellular ATP levels. Drug-induced kidney injury (DIKI) is the major cause of acute kidney injury (AKI). Renal proximal tubular epithelial cells (RPTECs) are the primary target sites of DIKI and express transporters involved in renal drug disposition. In the present study, we focused on three-dimensionally cultured human RPTECs (3D-RPTECs) with elevated expression of drug transporters to investigate their utility in DIKI evaluation. Intracellular ATP levels in 3D-RPTECs are reduced by tenofovir and cisplatin that are substrates of an organic anion transporter 1 and an organic cation transporter 2, respectively. In addition, 3D-RPTECs were exposed to 17 and 15 drugs that are positive and negative to RPTEC toxicity, respectively, for up to 28 d. The 20 % decreasing concentration of drugs for ATP amount (EC 20) was obtained, and the ratio of EC 20 values and clinical maximum concentration (C max) ≤100 were used as cut-off value to evaluate potential of DIKI. The sensitivities of 3D-RPTECs were 82.4 % and 88.2 % after 7 d and 28 d of drug exposure, respectively, and the specificities were 100 % and 93.3 %, respectively. The predictive performance of 3D-RPTECs was higher than that of two-dimensional cultured RPTECs and the kidney cell line HK-2. In conclusion, 3D-RPTECs are useful for in vitro evaluation of RPTEC injury by measuring intracellular ATP levels. [ABSTRACT FROM AUTHOR]

Amino acids are important for cellular metabolism. Their uptake across the plasma membrane is mediated by transport proteins. Despite the fact that the organic anion transporting polypeptide 4C1 (OATP4C1, Uniprot: Q6ZQN7) mediates transport of l‐arginine and l‐arginine derivatives, other members of the OATP family have not been characterized as amino acid transporters. The OATP family member OATP3A1 (gene symbol SLCO3A1, Uniprot: Q9UIG8) is ubiquitously expressed in human cells and highly expressed in many cancer tissues and cell lines. However, only a few substrates are known for OATP3A1. Accordingly, knowledge about its biological relevance is restricted. Our aim was to identify new substrates of OATP3A1 to gain insights into its (patho‐)physiological function. In an LC‐MS‐based untargeted metabolomics assay using untreated OATP3A1‐overexpressing HEK293 cells and control cells, we identified several amino acids as potential substrates of OATP3A1. Subsequent uptake experiments using exogenously added substrates revealed OATP3A1‐mediated transport of l‐tryptophan, l‐tyrosine, and l‐phenylalanine with 194.8 ± 28.7% (P < 0.05), 226.2 ± 18.7% (P < 0.001), and 235.2 ± 13.5% (P < 0.001), respectively, in OATP3A1‐overexpressing cells compared to control cells. Furthermore, kinetic transport parameters (Km values) were determined (Trp = 61.5 ± 14.2 μm, Tyr = 220.8 ± 54.5 μm, Phe = 234.7 ± 20.6 μm). In summary, we identified the amino acids l‐tryptophan, l‐tyrosine, and l‐phenylalanine as new substrates of OATP3A1. These findings could be used for a better understanding of (patho‐)physiological processes involving increased demand of amino acids, where OATP3A1 should be considered as an important uptake transporter of l‐tryptophan, l‐tyrosine, and l‐phenylalanine. [ABSTRACT FROM AUTHOR]

The four-carbon non-proteinogenic amino acid γ-aminobutyric acid (GABA) accumulates to high levels in plants in response to various abiotic and biotic stress stimuli, and plays a role in C:N balance, signaling, and as a transport regulator. Expression in Xenopus oocytes and voltage-clamping allowed the characterization of Arabidopsis GAT2 (At5g41800) as a low affinity GABA transporter with a K 0.5 G A B A ~8 mM. l -Alanine and butylamine represented additional substrates. GABA-induced currents were strongly dependent on the membrane potential, reaching the highest affinity and highest transport rates at strongly negative membrane potentials. Mutation of Ser17, previously reported to be phosphorylated in planta, did not result in altered affinity. In a short-term stress experiment, AtGAT2 mRNA levels were up-regulated at low water potential and under osmotic stress (polyethylene glycol and mannitol). Furthermore, AtGAT2 promoter activity was detected in vascular tissues, maturating pollen, and the phloem unloading region of young seeds. Even though this suggested a role for AtGAT2 in long-distance transport and loading of sink organs, under the conditions tested neither AtGAT2 -overexpressing plants, atgat2 or atgat1 T-DNA insertion lines, nor atgat1 atgat2 doubleknockout mutants differed from wild-type plants in growth on GABA, amino acid levels, or resistance to salt and osmotic stress. [ABSTRACT FROM AUTHOR]

137Cs diffused into the environment due to a nuclear power plant accident has caused serious problems for safe crop production. In plants, Cs+ is similar in its ionic form to K+. Cs+ is absorbed and transported mainly by the K+ transport mechanism. However, the full picture of the genes contributing to Cs+ transport and the transport mechanism of rice is still unclear. This study focused on OsHKT2;1, a candidate Cs+ transporter under low K+ conditions. To verify the ability of OsHKT2;1 to transport Cs+, the OsHKT2;1 mutant (hkt2;1) was grown in a 137Cs-contaminated paddy field in Fukushima. The 137Cs concentration in hkt2;1 aboveground was higher than in the wild type (WT), and the K concentration in these samples did not change between WT and hkt2;1, whereas the Na concentration was lower in hkt2;1. Uptake experiments with radioactive tracers (22Na+, 43K+, and 137Cs+) in hydroponic systems with different elemental compositions showed a negative correlation between Na+ and Cs+ accumulation in rice shoot cultivated under low K+ conditions. These results indicated that OsHKT2;1 does not directly contribute to Cs+ uptake but is an important factor in regulating Cs+ translocation by controlling Na+ accumulation. This indicates the possibility of controlling rice Cs content by regulating the Na+ environment during cultivation. [ABSTRACT FROM AUTHOR]

Rice tocopherols, vitamin E compounds with antioxidant activity, play essential roles in human health. Even though the key genes involved in vitamin E biosynthetic pathways have been identified in plants, the genetic architecture of vitamin E content in rice grain remains unclear. A genome-wide association study (GWAS) on 179 genotypically diverse rice accessions with 34,323 SNP markers was conducted to detect QTLs that define total and a-tocopherol contents in rice grains. Total and a-tocopherol contents had a strong positive correlation and varied greatly across the accessions, ranging from 0.230-31.76 and 0.011-30.83 (mg/g), respectively. A total of 13 QTLs were identified, which were spread across five of the rice chromosomes. Among the 13 QTLs, 11 were considered major with phenotypic variation explained (PVE) greater than 10%. Twelve transcription factor (TF) genes, one microprotein (miP), and a transposon were found to be associated with the QTLs with putative roles in controlling tocopherol contents. Moreover, intracellular transport proteins, ABC transporters, nonaspanins, and SNARE, were identified as associated genes on chromosomes 1 and 8. In the vicinity of seven QTLs, protein kinases were identified as key signaling factors. Haplotype analysis revealed the QTLs qAlph1.1, qTot1.1, qAlph2.1, qAlph6.1, qTot6.1, and qTot8.3 to have significant haplogroups. Quantitative RT-PCR validated the expression direction and magnitude of WRKY39 (Os02g0265200), PIP5Ks (Os08g0450800), and MADS59 (Os06g0347700) in defining the major tocopherol contents. This study provides insights for ongoing biofortification efforts to breed and/or engineer vitamin E and antioxidant levels in rice and other cereals. [ABSTRACT FROM AUTHOR]

AbstractThe Zrt/Irt-like protein (ZIP) family consists of ubiquitously expressed divalent d-block metal transporters that play central roles in the uptake, secretion, excretion, and distribution of several essential and toxic metals in living organisms. The past few years has witnessed rapid progress in the molecular basis of these membrane transport proteins. In this critical review, we summarize the research progress at the molecular level of the ZIP family and discuss the future prospects. Furthermore, an evolutionary path for the unique ZIP fold and a new classification of the ZIP family are proposed based on the presented structural and sequence analyses. [ABSTRACT FROM AUTHOR]

Following the Goldilocks principle, mitochondria size must be "just right." Mitochondria balance division and fusion to avoid becoming too big or too small. Defects in this balance produce dysfunctional mitochondria in human diseases. Mitochondrial safeguard (MitoSafe) is a defense mechanism that protects mitochondria against extreme enlarging by suppressing fusion in mammalian cells. In MitoSafe, hyperfused mitochondria elicit flickering—short pulses of mitochondrial depolarization. Flickering activates an inner membrane protease, Oma1, which in turn proteolytically inactivates a mitochondrial fusion protein, Opa1. The mechanisms underlying flickering are unknown. Using a live-imaging screen, we identified Slc25a3 (a mitochondrial carrier transporting phosphate and copper) as necessary for flickering and Opa1 cleavage. Remarkably, copper, but not phosphate, is critical for flickering. Furthermore, we found that two copper-containing mitochondrial enzymes, superoxide dismutase 1 and cytochrome c oxidase, regulate flickering. Our data identify an unforeseen mechanism linking copper, redox homeostasis, and membrane flickering in mitochondrial defense against deleterious fusion. [Display omitted] • Flickering prevents harmful extreme mitochondrial fusion by inducing Opa1 cleavage • Mitochondrial copper, transported by Slc25a3, regulates flickering • Copper-containing SOD1 suppresses flickering • Copper-containing cytochrome c oxidase promotes flickering Murata et al. report that mitochondrial copper, transported by Slc25a3, regulates the balance between mitochondrial fusion and division by suppressing fusion when mitochondria become hyperfused. This function of copper is mediated by two copper-containing enzymes: superoxide dismutase 1 and cytochrome c oxidase. [ABSTRACT FROM AUTHOR]

Aims Methods Results Conclusion Dersimelagon is a novel, investigational, orally administered, selective agonist of the melanocortin‐1 receptor that has demonstrated efficacy at increasing symptom‐free light exposure and an acceptable safety profile in patients with protoporphyria. A phase 1 drug–drug interaction (DDI) study demonstrated that dersimelagon 300 mg has the potential for clinically relevant DDIs with drugs that are substrates for breast cancer resistance protein, such as atorvastatin and rosuvastatin. This study uses physiologically based pharmacokinetic (PBPK) modelling to further investigate the DDI effects at lower doses of dersimelagon with substrate drugs.The data from in silico, in vitro and in vivo studies were used to construct a PBPK model for dersimelagon to assess the DDI potential between dersimelagon and substrate drugs for cytochrome P450 3A, P‐glycoprotein, organic anion transporting polypeptide 1B1/1B3, organic anion transporter 3 and breast cancer resistance protein, including atorvastatin and rosuvastatin.The systemic exposure of atorvastatin based on the maximum plasma concentration and area under the plasma concentration–time curve was predicted to increase 1.21‐fold and 1.25‐fold, respectively, if coadministered with dersimelagon 100 mg, and 1.42‐fold and 1.45‐fold with dersimelagon 200 mg. The systemic exposure of rosuvastatin followed trends similar to atorvastatin (1.67‐fold and 1.34‐fold increase in maximum plasma concentration and area under the plasma concentration–time curve, respectively, with dersimelagon 100 mg, and 2.40‐fold and 1.69‐fold with dersimelagon 200 mg).Overall, PBPK modelling results indicate that the simulated changes in plasma exposure of atorvastatin and rosuvastatin following coadministration with dersimelagon 100 or 200 mg are not clinically significant, but caution and appropriate clinical monitoring should be recommended. [ABSTRACT FROM AUTHOR]

ATP‐binding cassette (ABC) transporters hydrolyse ATP to transport various substrates. Previous studies have shown that ABC transporters are responsible for transporting plant hormones and heavy metals, thus contributing to plant immunity. Herein, we identified a wheat G‐type ABC transporter, TaABCG2‐5B, that responds to salicylic acid (SA) treatment and is induced by Fusarium graminearum, the primary pathogen causing Fusarium head blight (FHB). The loss‐of‐function mutation of TaABCG2‐5B (ΔTaabcg2‐5B) reduced SA accumulation and increased susceptibility to F. graminearum. Conversely, overexpression of TaABCG2‐5B (OE‐TaABCG2‐5B) exerted the opposite effect. Quantification of intracellular SA in ΔTaabcg2‐5B and OE‐TaABCG2‐5B protoplasts revealed that TaABCG2‐5B acts as an importer, facilitating the transport of SA into the cytoplasm. This role was further confirmed by Cd2+ absorption experiments in wheat roots, indicating that TaABCG2‐5B also participates in Cd2+ transport. Thus, TaABCG2‐5B acts as an importer and is crucial for transporting multiple substrates. Notably, the homologous gene TaABCG2‐5A also facilitated Cd2+ uptake in wheat roots but did not significantly influence SA accumulation or FHB resistance. Therefore, TaABCG2 could be a valuable target for enhancing wheat tolerance to Cd2+ and improving FHB resistance. [ABSTRACT FROM AUTHOR]

We measured the intracellular zinc ion concentration of murine fetal neural stem/progenitor cells (NSPCs) and that in the differentiated cells. The NSPCs cultured with 1.5 μM Zn2+ proliferated slightly faster than that in the zinc-deficient medium and the intracellular zinc concentration of the NSPCs and that of their differentiated cells (DCs) cultured with 1.5 μM Zn2+ was 1.34-fold and 2.00-fold higher than those in the zinc-deficient medium, respectively. The zinc transporter genes upregulated over the 3.5-fold change were Zip1, Zip4, Zip12, Zip13, ZnT1, ZnT8, and ZnT10 whereas the only downregulated one was Zip8 during the differentiation of NSPCs to DCs. The cell morphologies of both NSPCs and DCs in the low oxygen culture condition consisting of 2%O2 and 5%CO2, the high carbon dioxide condition consisting of 21%O2 and 10%CO2, and the normal condition consisting of 21%O2 and 5%CO2 were essentially the same each other. The expression of Zip4, Zip8, Zip12, and Zip14 was not drastically changed depending on the O2 and CO2 concentrations. [ABSTRACT FROM AUTHOR]

Ovarian cancer (OC), particularly high-grade serous carcinoma (HGSC), is a leading cause of gynecological cancer mortality due to late diagnosis and chemoresistance. While studies on OC cell lines have shown that overexpression of the ATP7A membrane transporter correlates with resistance to platinum-based drugs (PtBMs) and cross-resistance to copper (Cu), clinical evidence is lacking. The functionality of ceruloplasmin (CP), the main Cu-transporting protein in the blood, is dependent on, among other things, ATP7A activity. This study investigated ATP7A expression and CP levels as potential biomarkers for predicting responses to PtBMs. We included 28 HGSC patients who underwent neoadjuvant chemotherapy (NACT). ATP7A expression in ovarian and peritoneal tissues before NACT and in peritoneal and omental tissues after NACT was analyzed via qPCR, and CP levels in ascites and plasma were measured via ELISA before and after NACT. In total, 54% of patients exhibited ATP7A expression in pretreatment tissue (ovary and/or peritoneum), while 43% of patients exhibited ATP7A expression in tissue after treatment (peritoneum and/or omentum). A significant association was found between higher ATP7A expression in the peritoneum before NACT and an unfavorable CA-125 elimination rate constant k (KELIM) score. Patients with omental ATP7A expression had significantly higher plasma mean CP levels before NACT. Plasma CP levels decreased significantly after NACT, and higher CP levels after NACT were associated with a shorter platinum-free interval (PFI). These findings suggest that the ATP7A transporter and CP have the potential to serve as predictive markers of chemoresistance, but further research is needed to validate their clinical utility. [ABSTRACT FROM AUTHOR]

The neuronal glycine transporter GlyT2 removes glycine from the synaptic cleft through active Na+, Cl−, and glycine cotransport contributing to the termination of the glycinergic signal as well as supplying substrate to the presynaptic terminal for the maintenance of the neurotransmitter content in synaptic vesicles. Patients with mutations in the human GlyT2 gene (SLC6A5), develop hyperekplexia or startle disease (OMIM 149400), characterized by hypertonia and exaggerated startle responses to trivial stimuli that may have lethal consequences in the neonates as a result of apnea episodes. Post‐translational modifications in cysteine residues of GlyT2 are an aspect of structural interest we analyzed. Our study is compatible with a reversible and short‐lived S‐acylation in spinal cord membranes, detectable by biochemical and proteomics methods (acyl‐Rac binding and IP‐ABE) confirmed with positive and negative controls (palmitoylated and non‐palmitoylated proteins). According to a short‐lived modification, direct labeling using click chemistry was faint but mostly consistent. We have analyzed the physiological properties of a GlyT2 mutant lacking the cysteines with high prediction of palmitoylation and the mutant is less prone to be included in lipid rafts, an effect also observed upon treatment with the palmitoylation inhibitor 2‐bromopalmitate. This work demonstrates there are determinants of lipid raft inclusion associated with the GlyT2 mutated cysteines, which are presumably modified by palmitoylation. [ABSTRACT FROM AUTHOR]

Zrt/Irt-like protein 8 (ZIP8), which is a Zn transporter, plays a pivotal role as a Mn transporter. Recent studies have shown that a ZIP8 SNP (rs13107325 C→T, A391T) is associated with multiple diseases, likely by causing systemic Mn deficiency. However, the underlying molecular mechanisms remain unclear. We attempted to address this issue in cell-based experiments using Madin-Darby canine kidney cells stably expressing ZIP8 WT or the A391T SNP mutant under the control of the Tet-regulatable promoter. We showed that the A391T mutant lost the property of Mn-responsive accumulation on the cell surface, which was observed in WT ZIP8. We also showed that the loss of Mn-responsive accumulation of A391T mutant was associated with its reduced Mn uptake, compared with WT ZIP8, in the Mn uptake assay using the radioisotope 54Mn. Our results potentially explain how the ZIP8 A391T substitution is associated with disease pathogenesis caused by Mn deficiency. [ABSTRACT FROM AUTHOR]

Developing recombinant Saccharomyces cerevisiae strains capable of transporting and fermenting cellobiose directly is a promising strategy for second-generation ethanol production from lignocellulosic biomass. In this study, we cloned and expressed in the S. cerevisiae CEN.PK2-1C strain an intracellular β-glucosidase (SpBGL7) from Spathaspora passalidarum and co-expressed the cellobiose transporter SiHXT2.4 from Scheffersomyces illinoinensis, and two putative transporters, one from Candida tropicalis (CtCBT1 gene), and one from Meyerozyma guilliermondii (MgCBT2 gene). While all three transporters allowed cell growth on cellobiose, only the MgCBT2 permease allowed cellobiose fermentation, although cellobiose consumption was incomplete. The analysis of the β-glucosidase and transport activities revealed that the cells stopped consuming cellobiose due to a drop in the transport activity. Since ubiquitinylation of lysine residues at the N- or C-terminal domains of the permease are involved in the endocytosis and degradation of sugar transporters, we constructed truncated versions of the permease lacking lysine residues at the C-terminal domain (MgCBT2ΔC), and at both the C- and N-terminal domain (MgCBT2ΔNΔC) and co-expressed these permeases with the SpBGL7 β-glucosidase in an industrial strain. While the strain harboring the MgCBT2ΔC transporter continued to produce incomplete cellobiose fermentations as the wild-type MgCBT2 permease, the strain with the MgCBT2ΔNΔC permease was able to consume and ferment all the cellobiose present in the medium. Thus, our results highlight the importance of expressing cellobiose transporters lacking lysine at the N- and C-terminal domains for efficient cellobiose fermentation by recombinant S. cerevisiae. [ABSTRACT FROM AUTHOR]

The ubiquitin-proteasomal pathway regulates the functional expression of many membrane transporters in a variety of cellular systems. Nothing is currently known about the role of ubiquitin E3 ligase, neural precursor cell-expressed developmentally down-regulated gene 4 (Nedd4-1) and the proteasomal degradation pathway in regulating human vitamin C transporter-2 (hSVCT2) in neuronal cells. hSVCT2 mediates the uptake of ascorbic acid (AA) and is the predominantly expressed vitamin C transporter isoform in neuronal systems. Therefore, we addressed this knowledge gap in our study. Analysis of mRNA revealed markedly higher expression of Nedd4-1 in neuronal samples than that of Nedd4-2. Interestingly, Nedd4-1 expression in the hippocampus was higher in patients with Alzheimer's disease (AD) and age-dependently increased in the J20 mouse model of AD. The interaction of Nedd4-1 and hSVCT2 was confirmed by coimmunoprecipitation and colocalization. While the coexpression of Nedd4-1 with hSVCT2 displayed a significant decrease in AA uptake, siRNA-mediated knockdown of Nedd4-1 expression up-regulated the AA uptake. Further, we mutated a classical Nedd4 protein interacting motif ("PPXY") within the hSVCT2 polypeptide and observed markedly decreased AA uptake due to the intracellular localization of the mutated hSVCT2. Also, we determined the role of the proteasomal degradation pathway in hSVCT2 functional expression in SH-SY5Y cells and the results indicated that the proteasomal inhibitor (MG132) significantly up-regulated the AA uptake and hSVCT2 protein expression level. Taken together, our findings show that the regulation of hSVCT2 functional expression is at least partly mediated by the Nedd4-1 dependent ubiquitination and proteasomal pathways.

Heavy metal (HM) pollution, specifically cadmium (Cd) contamination, is a worldwide concern for its consequences for plant health and ecosystem stability. This review sheds light on the intricate mechanisms underlying Cd toxicity in plants and the various strategies employed by these organisms to mitigate its adverse effects. From molecular responses to physiological adaptations, plants have evolved sophisticated defense mechanisms to counteract Cd stress. We highlighted the role of phytochelatins (PCn) in plant detoxification, which chelate and sequester Cd ions to prevent their accumulation and minimize toxicity. Additionally, we explored the involvement of glutathione (GSH) in mitigating oxidative damage caused by Cd exposure and discussed the regulatory mechanisms governing GSH biosynthesis. We highlighted the role of transporter proteins, such as ATP-binding cassette transporters (ABCs) and heavy metal ATPases (HMAs), in mediating the uptake, sequestration, and detoxification of Cd in plants. Overall, this work offered valuable insights into the physiological, molecular, and biochemical mechanisms underlying plant responses to Cd stress, providing a basis for strategies to alleviate the unfavorable effects of HM pollution on plant health and ecosystem resilience.

Abstract We investigated whether calorie restriction (CR) enhances metabolic adaptations to endurance training (ET). Ten-week-old male Institute of Cancer Research (ICR) mice were fed ad libitum or subjected to 30% CR. The mice were subdivided into sedentary and ET groups. The ET group performed treadmill running (20–25 m/min, 30 min, 5 days/week) for 5 weeks. We found that CR decreased glycolytic enzyme activity and monocarboxylate transporter (MCT) 4 protein content, while enhancing glucose transporter 4 protein content in the plantaris and soleus muscles. Although ET and CR individually increased citrate synthase activity in the plantaris muscle, the ET-induced increase in respiratory chain complex I protein content was counteracted by CR. In the soleus muscle, mitochondrial enzyme activity and protein levels were increased by ET, but decreased by CR. It has been suggested that CR partially interferes with skeletal muscle adaptation to ET.

The Mycobacterium tuberculosis (Mtb) cell envelope provides a protective barrier against the immune response and antibiotics. The mycobacterial membrane protein large (MmpL) family of proteins export cell envelope lipids and siderophores; therefore, these proteins are important for the basic biology and pathogenicity of Mtb. In particular, MmpL3 is essential and a known drug target. Despite interest in MmpL3, the structural data in the field are incomplete. Utilizing homology modeling, AlphaFold, and biophysical techniques, we characterized the cytoplasmic C‐terminal domain (CTD) of MmpL3 to better understand its structure and function. Our in silico models of the MmpL11TB and MmpL3TB CTD reveal notable features including a long unstructured linker that connects the globular domain to the last transmembrane (TM) in each transporter, charged lysine and arginine residues facing the membrane, and a C‐terminal alpha helix. Our predicted overall structure enables a better understanding of these transporters. [ABSTRACT FROM AUTHOR]

Pseudomonas aeruginosa is an opportunistic pathogen that requires iron to survive in the host; however, the host immune system limits the availability of iron. Pyochelin (PCH) is a major siderophore produced by P. aeruginosa during infection, which can help P. aeruginosa survive in an iron-restricted environment and cause infection. The infection activity of P. aeruginosa is regulated by the Pseudomonas quinolone signal (PQS) quorum-sensing system. The system uses 2-heptyl-3-hydroxy-4-quinolone (PQS) or its precursor, 2-heptyl-4-quinolone (HHQ), as the signal molecule. PQS can control specific life processes such as mediating quorum sensing, cytotoxicity, and iron acquisition. This review summarizes the biosynthesis of PCH and PQS, the shared transport system of PCH and PQS, and the regulatory relationship between PCH and PQS. The correlation between the PQS and PCH is emphasized to provide a new direction for future research. [ABSTRACT FROM AUTHOR]

This study investigated sex-specific differences in high-energy phosphate, glycolytic, and mitochondrial enzyme activities and also metabolite transporter protein levels in the skeletal muscles of adult (5 months old), middle-aged (12 months old), and advanced-aged (24 months old) mice. While gastrocnemius glycogen content increased with age regardless of sex, gastrocnemius triglyceride levels increased only in advanced-aged female mice. Aging decreased creatine kinase and adenylate kinase activities in the plantaris muscle of both sexes and in the soleus muscle of male mice but not in female mice. Irrespective of sex, phosphofructokinase and lactate dehydrogenase (LDH) activities decreased in the plantaris and soleus muscles. Additionally, hexokinase activity in the plantaris muscle and LDH activity in the soleus muscle decreased to a greater extent in aged male mice compared with those in aged female mice. Mitochondrial enzyme activities increased in the plantaris muscle of aged female mice but did not change in male mice. The protein content of the glucose transporter 4 in the aged plantaris muscle and fatty acid translocase/cluster of differentiation 36 increased in the aged plantaris and soleus muscles of both sexes, with a significantly higher content in female mice. These findings suggest that females possess a better ability to maintain metabolic enzyme activity and higher levels of metabolite transport proteins in skeletal muscle during aging, despite alterations in lipid metabolism. Our data provide a basis for studying muscle metabolism in the context of age-dependent metabolic perturbations and diseases that affect females and males differently. [ABSTRACT FROM AUTHOR]

Most medications undergo metabolism and elimination via CYP450 enzymes, while uptake and efflux transporters play vital roles in drug elimination from various organs. Interactions often occur when multiple drugs share CYP450-transporter-mediated metabolic pathways, necessitating a unique clinical care strategy to address the diverse types of CYP450 and transporter-mediated drug-drug interactions (DDI). The primary focus of this review is to record relevant mechanisms regarding DDI between COVID-19 and tuberculosis (TB) treatments, specifically through the influence of CYP450 enzymes and transporters on drug absorption, distribution, metabolism, elimination, and pharmacokinetics. This understanding empowers clinicians to prevent subtherapeutic and supratherapeutic drug levels of COVID medications when co-administered with TB drugs, thereby mitigating potential challenges and ensuring optimal treatment outcomes. A comprehensive analysis is presented, encompassing various illustrative instances of TB drugs that may impact COVID-19 clinical behavior, and vice versa. This review aims to provide valuable insights to healthcare providers, facilitating informed decision-making and enhancing patient safety while managing co-infections. Ultimately, this study contributes to the body of knowledge necessary to optimize therapeutic approaches and improve patient outcomes in the face of the growing challenges posed by infectious diseases. [ABSTRACT FROM AUTHOR]

Sugars Will Eventually be Exported Transporters (SWEETs) are the most recently discovered family of plant sugar transporters. By acting as uniporters, SWEETs facilitate the diffusion of sugars across cell membranes and play an important role in various physiological processes such as abiotic stress adaptation. At SWEET17, a vacuolar fructose facilitator, was shown to be involved in the modulation of the root system during drought. In addition, previous studies have shown that overexpression of an apple homolog leads to increased drought tolerance in tomato plants. Therefore, SWEET17 might be a molecular element involved in plant responses to drought. However, the role and function of SWEET17 in above-ground tissues of Arabidopsis under drought stress remain elusive. By combining gene expression analysis and stem architecture with the sugar profiles of different above-ground tissues, we uncovered a putative role for SWEET17 in carbohydrate supply and thus cauline branch elongation, especially during periods of carbon limitation, as occurs under drought stress. Thus, SWEET17 seems to be involved in maintaining efficient plant reproduction under drought stress conditions. [ABSTRACT FROM AUTHOR]

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In secondary active transporters, a relatively limited set of protein folds have evolved diverse solute transport functions. Because of the conformational changes inherent to transport, altering substrate specificity typically involves remodeling the entire structural landscape, limiting our understanding of how novel substrate specificities evolve. In the current work, we examine a structurally minimalist family of model transport proteins, the small multidrug resistance (SMR) transporters, to understand the molecular basis for the emergence of a novel substrate specificity. We engineer a selective SMR protein to promiscuously export quaternary ammonium antiseptics, similar to the activity of a clade of multidrug exporters in this family. Using combinatorial mutagenesis and deep sequencing, we identify the necessary and sufficient molecular determinants of this engineered activity. Using X-ray crystallography, solid-supported membrane electrophysiology, binding assays, and a proteoliposome-based quaternary ammonium antiseptic transport assay that we developed, we dissect the mechanistic contributions of these residues to substrate polyspecificity. We find that substrate preference changes not through modification of the residues that directly interact with the substrate but through mutations peripheral to the binding pocket. Our work provides molecular insight into substrate promiscuity among the SMRs and can be applied to understand multidrug export and the evolution of novel transport functions more generally. [ABSTRACT FROM AUTHOR]

SUMMARY: The basal region of maize (Zea mays) kernels, which includes the pedicel, placenta‐chalazal, and basal endosperm transfer layers, serves as the maternal/filial interface for nutrient transfer from the mother plant to the developing seed. However, transcriptome dynamics of this maternal/filial interface remain largely unexplored. To address this gap, we conducted high‐temporal‐resolution RNA sequencing of the basal and upper kernel regions between 4 and 32 days after pollination and deeply analyzed transcriptome dynamics of the maternal/filial interface. Utilizing 790 specifically and highly expressed genes in the basal region, we performed the gene ontology (GO) term and weighted gene co‐expression network analyses. In the early‐stage basal region, we identified five MADS‐box transcription factors (TFs) as hubs. Their homologs have been demonstrated as pivotal regulators at the maternal/filial interface of rice or Arabidopsis, suggesting their potential roles in maize kernel development. In the filling‐stage basal region, numerous GO terms associated with transcriptional regulation and transporters are significantly enriched. Furthermore, we investigated the molecular function of three hub TFs. Through genome‐wide DNA affinity purification sequencing combined with promoter transactivation assays, we suggested that these three TFs act as regulators of 10 basal‐specific transporter genes involved in the transfer of sugars, amino acids, and ions. This study provides insights into transcriptomic dynamic and regulatory modules of the maternal/filial interface. In the future, genetic investigation of these hub regulators must advance our understanding of maternal/filial interface development and function. Significance Statement: Our study is the first systematic effort to generate high‐temporal‐resolution transcriptomic dynamics of this maternal/filial interface in the basal kernel region of maize. We identified 790 highly expressed genes in the basal region, providing a resource for understanding the development and functioning of this interface. We analyzed three previously unknown transcription factors that potentially play a critical role in regulating transporter genes related to sugar, amino acids, and ions. [ABSTRACT FROM AUTHOR]

We investigated whether calorie restriction (CR) enhances metabolic adaptations to endurance training (ET). Ten-week-old male Institute of Cancer Research (ICR) mice were fed ad libitum or subjected to 30% CR. The mice were subdivided into sedentary and ET groups. The ET group performed treadmill running (20–25 m/min, 30 min, 5 days/week) for 5 weeks. We found that CR decreased glycolytic enzyme activity and monocarboxylate transporter (MCT) 4 protein content, while enhancing glucose transporter 4 protein content in the plantaris and soleus muscles. Although ET and CR individually increased citrate synthase activity in the plantaris muscle, the ET-induced increase in respiratory chain complex I protein content was counteracted by CR. In the soleus muscle, mitochondrial enzyme activity and protein levels were increased by ET, but decreased by CR. It has been suggested that CR partially interferes with skeletal muscle adaptation to ET. [ABSTRACT FROM AUTHOR]

Insufficient expression of steroidogenic acute regulatory-related lipid transfer protein 5 (StarD5) on liver cholesterol/lipid homeostasis is not clearly defined. The ablation of StarD5 was analyzed in mice on a normal or Western diet (WD) to determine its importance in hepatic lipid accumulation and fibrosis compared with wild-type (WT) mice. Rescue experiments in StarD5−/− mice and hepatocytes were performed. In addition to increased hepatic triglyceride (TG)-cholesterol levels, global StarD5−/− mice fed a normal diet displayed reduced plasma triglycerides and liver very low-density lipoprotein (VLDL) secretion as compared with WT counterparts. Insulin levels and homeostatic model assessment for insulin resistance (HOMA-IR) scoring were elevated, demonstrating developing insulin resistance (IR). WD-fed StarD5−/− mice upregulated WW domain containing transcription regulator 1 (TAZ or WWTR1) expression with accelerated liver fibrosis when compared with WD-fed WT mice. Suppression of oxysterol 7α-hydroxylase (CYP7B1) coupled with chronic accumulation of toxic oxysterol levels correlated with presentation of fibrosis. "Hepatocyte-selective" StarD5 overexpression in StarD5−/− mice restored expression, reduced hepatic triglycerides, and improved HOMA-IR. Observations in two additional mouse and one human metabolic dysfunction-associated steatotic liver disease (MASLD) model were supportive. The downregulation of StarD5 with hepatic lipid excess is a previously unappreciated physiological function appearing to promote lipid storage for future needs. Conversely, lingering downregulation of StarD5 with prolonged lipid-cholesterol excess accelerates fatty liver's transition to fibrosis; mediated via dysregulation in the oxysterol signaling pathway. NEW & NOTEWORTHY: We have found that deletion of the cholesterol transport protein StarD5 in mice leads to an increase in insulin resistance and lipid accumulation due to the upregulation of lipid synthesis and decrease VLDL secretion from the liver. In addition, deletion of StarD5 increased fibrosis when mice were fed a Western diet. This represents a novel pathway of fibrosis development in the liver. [ABSTRACT FROM AUTHOR]

Copper (Cu) is an essential micronutrient for all living organisms but is also highly toxic in excess. Cellular homoeostasis of Cu is maintained by various transporters and metallochaperones. Here, we investigated the biological function of OsCOPT7, a member of the copper transporters (COPT) family, in Cu homoeostasis in rice. OsCOPT7 was mainly expressed in the roots and the expression was upregulated by Cu deficiency. OsCOPT7 was localized at the tonoplast and the endoplasmic reticulum. Knockout of OsCOPT7 increased Cu accumulation in the roots but decreased Cu concentrations in the shoots and grain. The knockout mutants contained higher concentrations of Cu in the roots cell sap but markedly lower concentrations of Cu in the xylem sap than wild‐type plants. Seed setting and grain yield were reduced significantly in the knockout mutants grown in a low Cu soil. Knockout mutants were more tolerant to Cu toxicity. Yeast two‐hybrid and bimolecular fluorescence complementation assays showed that OsCOPT7 interacts physically with the rice Cu chaperone antioxidant protein 1 (OsATX1). Taken together, our results indicate that OsCOPT7 is a specific Cu transporter functioning to export Cu from the vacuoles and the ER and plays an important role in controlling the root‐to‐shoot Cu translocation in rice. Summary statement: Copper (Cu) is an essential micronutrient for all living organisms but is also highly toxic in excess. Cellular homoeostasis of Cu is maintained by various transporters and metallochaperones. We show that OsCOPT7 is a Cu transporter functioning to export Cu from the vacuoles and the endoplasmic reticulum and plays an important role in controlling the root‐to‐shoot Cu translocation in rice. [ABSTRACT FROM AUTHOR]

Grain filling is critical for determining yield and quality, raising the question of whether central coordinators exist to facilitate the uptake and storage of various substances from maternal to filial tissues. The duplicate NAC transcription factors ZmNAC128 and ZmNAC130 could potentially serve as central coordinators. By analyzing differentially expressed genes from zmnac128 zmnac130 mutants across different genetic backgrounds and growing years, we identified 243 highly and differentially expressed genes (hdEGs) as the core target genes. These 243 hdEGs were associated with storage metabolism and transporters. ZmNAC128 and ZmNAC130 play vital roles in storage metabolism, and this study revealed two additional starch metabolism-related genes, sugary enhancer1 and hexokinase1, as their direct targets. A key finding of this study was the inclusion of 17 transporter genes within the 243 hdEGs, with significant alterations in the levels of more than 10 elements/substances in mutant kernels. Among them, six out of the nine upregulated transporter genes were linked to the transport of heavy metals and metalloids (HMMs), which was consistent with the enrichment of cadmium, lead, and arsenic observed in mutant kernels. Interestingly, the levels of Mg and Zn, minerals important to biofortification efforts, were reduced in mutant kernels. In addition to their direct involvement in sugar transport, ZmNAC128 and ZmNAC130 also activate the expression of the endosperm-preferential nitrogen and phosphate transporters ZmNPF1.1 and ZmPHO1;2. This coordinated regulation limits the intake of HMMs, enhances biofortification, and facilitates the uptake and storage of essential nutrients. [ABSTRACT FROM AUTHOR]

Queuosine (Q) is a modification of the wobble base of tRNA harboring GUN anticodons with roles in decoding accuracy and efficiency. Its synthesis is complex with multiple enzymatic steps, and several pathway intermediates can be salvaged. The only two transporter families known to salvage Q precursors are QPTR/COG1738 and QrtT/QueT. Analyses of the distribution of known Q synthesis and salvage genes in human gut and oral microbiota genomes have suggested that more transporter families remain to be found and that Q precursor exchanges must occur within the structured microenvironments of the mammalian host. Using physical clustering and fusion-based association with Q salvage genes, candidate genes for missing transporters were identified and five were tested experimentally by complementation assays in Escherichia coli. Three genes encoding transporters from three different Pfam families, a ureide permease (PF07168) from Acidobacteriota bacterium, a hemolysin III family protein (PF03006) from Bifidobacterium breve, and a Major Facilitator Superfamily protein (PF07690) from Bartonella henselae, were found to allow the transport of both preQ0 and preQ1 in this heterologous system. This work suggests that many transporter families can evolve to transport Q precursors, reinforcing the concept of transporter plasticity. [ABSTRACT FROM AUTHOR]

137Cs diffused into the environment due to a nuclear power plant accident has caused serious problems for safe crop production. In plants, Cs+ is similar in its ionic form to K+. Cs+ is absorbed and transported mainly by the K+ transport mechanism. However, the full picture of the genes contributing to Cs+ transport and the transport mechanism of rice is still unclear. This study focused on OsHKT2;1, a candidate Cs+ transporter under low K+ conditions. To verify the ability of OsHKT2;1 to transport Cs+, the OsHKT2;1 mutant (hkt2;1) was grown in a 137Cs-contaminated paddy field in Fukushima. The 137Cs concentration in hkt2;1 aboveground was higher than in the wild type (WT), and the K concentration in these samples did not change between WT and hkt2;1, whereas the Na concentration was lower in hkt2;1. Uptake experiments with radioactive tracers (22Na+, 43K+, and 137Cs+) in hydroponic systems with different elemental compositions showed a negative correlation between Na+ and Cs+ accumulation in rice shoot cultivated under low K+ conditions. These results indicated that OsHKT2;1 does not directly contribute to Cs+ uptake but is an important factor in regulating Cs+ translocation by controlling Na+ accumulation. This indicates the possibility of controlling rice Cs content by regulating the Na+ environment during cultivation.

Rice tocopherols, vitamin E compounds with antioxidant activity, play essential roles in human health. Even though the key genes involved in vitamin E biosynthetic pathways have been identified in plants, the genetic architecture of vitamin E content in rice grain remains unclear. A genome-wide association study (GWAS) on 179 genotypically diverse rice accessions with 34,323 SNP markers was conducted to detect QTLs that define total and α- tocopherol contents in rice grains. Total and α-tocopherol contents had a strong positive correlation and varied greatly across the accessions, ranging from 0.230-31.76 and 0.011-30.83 (μg/g), respectively. A total of 13 QTLs were identified, which were spread across five of the rice chromosomes. Among the 13 QTLs, 11 were considered major with phenotypic variation explained (PVE) greater than 10%. Twelve transcription factor (TF) genes, one microprotein (miP), and a transposon were found to be associated with the QTLs with putative roles in controlling tocopherol contents. Moreover, intracellular transport proteins, ABC transporters, nonaspanins, and SNARE, were identified as associated genes on chromosomes 1 and 8. In the vicinity of seven QTLs, protein kinases were identified as key signaling factors. Haplotype analysis revealed the QTLs qAlph1.1, qTot1.1, qAlph2.1, qAlph6.1, qTot6.1, and qTot8.3 to have significant haplogroups. Quantitative RT-PCR validated the expression direction and magnitude of WRKY39 (Os02g0265200), PIP5Ks (Os08g0450800), and MADS59 (Os06g0347700) in defining the major tocopherol contents. This study provides insights for ongoing biofortification efforts to breed and/or engineer vitamin E and antioxidant levels in rice and other cereals.

Background Amino acids are not only the main form of N in rice, but also are vital for its growth and development. These processes are facilitated by amino acid transporters within the plant. Despite their significance, only a few AAP amino acid transporters have been reported. Results In this study, we observed that there were differences in the expression of amino acid transporter OsAAP7 among 521 wild cultivated rice varieties, and it directly negatively correlated with tillering and grain yield per plant. We revealed that OsAAP7 protein was localized to the endoplasmic reticulum and had absorption and transport affinity for amino acids such as phenylalanine (Phe), lysine (Lys), leucine (Leu), and arginine (Arg) using subcellular localization, yeast substrate testing, fluorescent amino acid uptake, and amino acid content determination. Further hydroponic studies showed that exogenous application of amino acids Phe, Lys and Arg inhibited the growth of axillary buds in the overexpression lines, and promoted the elongation of axillary buds in the mutant lines. Finally, RNA-seq analysis showed that the expression patterns of genes related to nitrogen, auxin and cytokinin pathways were changed in axillary buds of OsAAP7 transgenic plants. Conclusions This study revealed the gene function of OsAAP7, and found that blocking of amino acid transporter OsAAP7 with CRISPR/Cas9 technology promoted tillering and yield by determining basic and neutral amino acids accumulation in rice.

Low-affinity nitrate transporter genes have been identified in subfamilies 4–8 of the rice nitrate transporter 1 (NRT1)/peptide transporter family (NPF), but the OsNPF3 subfamily responsible for nitrate and phytohormone transport and rice growth and development remains unknown. In this study, we described OsNPF3.1 as an essential nitrate and phytohormone transporter gene for rice tillering and nitrogen utilization efficiency (NUtE). OsNPF3.1 possesses four major haplotypes of its promoter sequence in 517 cultivars, and its expression is positively associated with tiller number. Its expression was higher in the basal part, culm, and leaf blade than in other parts of the plant, and was strongly induced by nitrate, abscisic acid (ABA) and gibberellin 3 (GA3) in the root and shoot of rice. Electrophysiological experiments demonstrated that OsNPF3.1 is a pH-dependent low-affinity nitrate transporter, with rice protoplast uptake assays showing it to be an ABA and GA3 transporter. OsNPF3.1 overexpression significantly promoted ABA accumulation in the roots and GA accumulation in the basal part of the plant which inhibited axillary bud outgrowth and rice tillering, especially at high nitrate concentrations. The NUtE of OsNPF3.1-overexpressing plants was enhanced under low and medium nitrate concentrations, whereas the NUtE of OsNPF3.1 clustered regularly interspaced short palindromic repeats (CRISPR) plants was increased under high nitrate concentrations. The results indicate that OsNPF3.1 transports nitrate and phytohormones in different rice tissues under different nitrate concentrations. The altered OsNPF3.1 expression improves NUtE in the OsNPF3.1-overexpressing and CRISPR lines at low and high nitrate concentrations, respectively.

Abstract The neurotransmitter serotonin plays a pivotal role in mood and depression. It also acts as a vasoconstrictor within blood vessels and is the main neurotransmitter in the gastrointestinal system. In neurotransmission, released serotonin is taken up by serotonin transporters, which are principal targets of antidepressants and the psychostimulant, ecstasy. The investigation of serotonin transporters have relied almost exclusively on the use of radiolabeled serotonin in heterogenous end-point assays. Here we adapt the genetically encoded fluorescent biosensor, iSeroSnFR, to establish and validate the Serotonin (5-HT) Fluorescence Assay for Transport and Release (5-HT_FAsTR) for functional and pharmacological studies of serotonin transport and release. We demonstrate the applicability of the method for the study of a neuronal, high-affinity, low-capacity serotonin transporter (SERT) as well as an extraneuronal low-affinity, high-capacity organic cation transporter and mutants thereof. 5HT_FAsTR offers an accessible, versatile and reliable semi-homogenous assay format that only relies on a fluorescence plate reader for repeated, real-time measurements of serotonin influx and efflux. 5HT_FAsTR accelerates and democratizes functional characterization and pharmacological studies of serotonin transporters and genetic variants thereof in disease states such as depression, anxiety and ADHD.

In bacteria, the cell envelope is the key element surrounding and protecting the bacterial content from mechanical or osmotic damages. It allows the selective interchanges of solutes, ions, cellular debris, and drugs between the cellular compartments and the external environment, thanks to the presence of transmembrane proteins called transporters. The major component of the cell envelope is the peptidoglycan, consisting of long linear glycan strands cross-linked by short peptide stems. During cell growth or under stress conditions, peptidoglycan fragments, the muropeptides, are released by bacteria and recognized by the host Pattern Recognition Receptor, promoting the activation of their innate defense mechanisms. The review sums up the salient aspects of microbiota–host interaction with a focus on the NOD-dependent immune response to bacterial peptidoglycan and on the accountability of muropeptide transporters in the crosstalk with the host and in antibiotic resistance. Furthermore, it retraces the discoveries and applications of microorganisms-derived components such as vaccines or vaccine adjuvants.

Drug-metabolizing enzymes and transporters are major determinants of the absorption, disposition, metabolism, and excretion (ADME) of drugs, and changes in ADME gene expression or function may alter the pharmacokinetics/ pharmacodynamics (PK/PD) and further influence drug safety and therapeutic outcomes. ADME gene functions are controlled by diverse factors, such as genetic polymorphism, transcriptional regulation, and coadministered medications. MicroRNAs (miRNAs) are a superfamily of regulatory small noncoding RNAs that are transcribed from the genome to regulate target gene expression at the post-transcriptional level. The roles of miRNAs in controlling ADME gene expression have been demonstrated, and such miRNAs may consequently influence cellular drug metabolism and disposition capacity. Several types of miRNA mimics and small interfering RNA (siRNA) reagents have been developed and widely used for ADME research. In this review article, we first provide a brief introduction to the mechanistic actions of miRNAs in post-transcriptional gene regulation of drug-metabolizing enzymes, transporters, and transcription factors. After summarizing conventional small RNA production methods, we highlight the latest advances in novel recombinant RNA technologies and applications of the resultant bioengineered RNA (BioRNA) agents to ADME studies. BioRNAs produced in living cells are not only powerful tools for general biological and biomedical research but also potential therapeutic agents amenable to clinical investigations.

Plant root systems play a pivotal role in plant physiology and exhibit diverse phenotypic traits. Understanding the genetic mechanisms governing root growth and development in model plants like maize is crucial for enhancing crop resilience to drought and nutrient limitations. This study focused on identifying and characterizing ZmPILS6, an annotated auxin efflux carrier, as a key regulator of various crown root traits in maize. ZmPILS6-modified roots displayed reduced network area and suppressed lateral root formation, which are desirable traits for the "steep, cheap, and deep" ideotype. The research revealed that ZmPILS6 localizes to the endoplasmic reticulum and plays a vital role in controlling the spatial distribution of indole-3-acetic acid (IAA or "auxin") in primary roots. The study also demonstrated that ZmPILS6 can actively efflux IAA when expressed in yeast. Furthermore, the loss of ZmPILS6 resulted in significant proteome remodeling in maize roots, particularly affecting hormone signaling pathways. To identify potential interacting partners of ZmPILS6, a weighted gene coexpression analysis was performed. Altogether, this research contributes to the growing knowledge of essential genetic determinants governing maize root morphogenesis, which is crucial for guiding agricultural improvement strategies. [ABSTRACT FROM AUTHOR]

Non-alcoholic fatty liver disease (NAFLD) is one of the most common liver diseases, which can cause serious complications and gradually increase the mortality rate. However, the effects of NAFLD on drug-metabolizing enzymes and transporters remain unclear, which may cause some confusion regarding patient medication. In this study, a NAFLD rat model was constructed by feeding rats with methionine and choline deficiency diets for 6 weeks, and the mRNA and protein levels of drug-metabolizing enzymes and transporter were analyzed by real-time fluorescent quantitative PCR and Western blot, respectively. The activity of drug-metabolizing enzymes was detected by cocktail methods. In the NAFLD rat model, the mRNA expression of phase I enzymes, phase II enzymes, and transporters decreased. At the protein level, only CYP1A1, CYP1B1, CYP2C11, and CYP2J3 presented a decrease. In addition, the activities of CYP1A2, CYP2B1, CYP2C11, CYP2D1, CYP3A2, UGT1A1, UGT1A3, UGT1A6, and UGT1A9 decreased. These changes may be caused by the alteration of FXR, HNF4α, LXRα, LXRβ, PXR, and RXR. In conclusion, NAFLD changes the expression and activity of hepatic drug-metabolizing enzymes and transporters in rats, which may affect drug metabolism and pharmacokinetics. In clinical medication, drug monitoring should be strengthened to avoid potential risks. [Display omitted] • The expression of drug-metabolizing enzymes and transporters changed in non-alcoholic fatty liver disease (NAFLD) rats. • The activity of hepatic drug-metabolizing enzymes was reduced in NAFLD rats. • The expression of nuclear receptors was downregulated in NAFLD rats. [ABSTRACT FROM AUTHOR]

• This study revealed that Asn124, Asn135 and Asn492 in OATP1A2 and Asn176 and Asn538 in OATP2B1 are N -glycosylation sites. • The effect of deletion of N -glycans on the transport activity of OATPs was evaluated based on the kinetics parameters per single molecules using the expression level of OATPs on the plasma membrane quantified by LC-MS/MS. • Among the three N -glycans of OATP1A2, those at Asn124 and Asn135 are especially important for the expression on the plasma membrane and transport activity. • Deletion of each N -glycan of OATP2B1 reduces the expression on the plasma membrane and enhances substrate affinity, and these effects becomes stronger when both N -glycans are disrupted. Organic anion transporting polypeptide (OATP)1A2 and OATP2B1 have potential N -glycosylation sites, but their influence remains unclear. This study aimed to identify the N -glycosylation sites of OATP1A2/2B1 and investigate their impact on the expression and function of OATP1A2/2B1. Human embryonic kidney cells expressing OATP1A2 or OATP2B1 (HEK293-OATP1A2/2B1) were exposed to tunicamycin, an N -glycosylation inhibitor, and a plasma membrane fraction (PMF) Western blot assay and an estrone 3-sulfate (E3S) uptake study were conducted. HEK293-OATP1A2/OATP2B1 cell lines with mutation(s) at potential N -glycosylation sites were established, and the Western blotting and uptake study were repeated. Tunicamycin reduced the PMF levels and E3S uptake of OATP1A2/OATP2B1. The Asn124Gln, Asn135Gln, and Asn492Gln mutations in OATP1A2 and Asn176Gln and Asn538Gln mutations in OATP2B1 reduced the molecular weights of the OATP molecules and their PMF levels. The PMF levels of OATP1A2 Asn124/135Gln, OATP1A2 Asn124/135/492Gln, and OATP2B1 Asn176/538Gln were further reduced. The maximum transport velocities of OATP1A2 Asn124Gln, OATP1A2 Asn135Gln, and OATP2B1 Asn176/538Gln were markedly reduced to 10 %, 4 %, and 10 % of the wild-type level, respectively. In conclusion, the N -glycans at Asn124 and Asn135 of OATP1A2 and those at Asn176 and Asn538 of OATP2B1 are essential for the plasma membrane expression of these molecules and also affect their transport function. [ABSTRACT FROM AUTHOR]

Purpose: Olmesartan medoxomil (olmesartan-MX), an ester-type prodrug of the angiotensin II receptor blocker (ARB) olmesartan, is predominantly anionic at intestinal pH. Human organic anion transporting polypeptide 2B1 (OATP2B1) is expressed in the small intestine and is involved in the absorption of various acidic drugs. This study was designed to test the hypothesis that OATP2B1-mediated uptake contributes to the enhanced intestinal absorption of olmesartan-MX, even though olmesartan itself is not a substrate of OATP2B1. Methods: Tetracycline-inducible human OATP2B1- and rat Oatp2b1-overexpressing HEK 293 cell lines (hOATP2B1/T-REx-293 and rOatp2b1/T-REx-293, respectively) were established to characterize OATP2B1-mediated uptake. Rat jejunal permeability was measured using Ussing chambers. ARBs were quantified by liquid chromatography-tandem mass spectrometry. Results: Significant olmesartan-MX uptake was observed in hOATP2B1/T-REx-293 and rOatp2b1/T-REx-293 cells, whereas olmesartan uptake was undetectable or much lower than olmesartan-MX uptake, respectively. Furthermore, olmesartan-MX exhibited several-fold higher uptake in Caco-2 cells and greater permeability in rat jejunum compared to olmesartan. Olmesartan-MX uptake in hOATP2B1/T-REx-293 cells and in Caco-2 cells was significantly decreased by OATP2B1 substrates/inhibitors such as 1 mM estrone-3-sulfate, 100 µM rifamycin SV, and 100 µM fluvastatin. Rat Oatp2b1-mediated uptake and rat jejunal permeability of olmesartan-MX were significantly decreased by 50 µM naringin, an OATP2B1 inhibitor. Oral administration of olmesartan-MX with 50 µM naringin to rats significantly reduced the area under the plasma concentration–time curve of olmesartan to 76.9%. Conclusion: Olmesartan-MX is a substrate for OATP2B1, and the naringin-sensitive transport system contributes to the improved intestinal absorption of olmesartan-MX compared with its parent drug, olmesartan. [ABSTRACT FROM AUTHOR]

Understanding the homeostatic interactions among essential trace metals is important for explaining their roles in cellular systems. Recent studies in vertebrates suggest that cellular Mn metabolism is related to Zn metabolism in multifarious cellular processes. However, the underlying mechanism remains unclear. In this study, we examined the changes in the expression of proteins involved in cellular Zn and/or Mn homeostatic control and measured the Mn as well as Zn contents and Zn enzyme activities to elucidate the effects of Mn and Zn homeostasis on each other. Mn treatment decreased the expression of the Zn homeostatic proteins metallothionein (MT) and ZNT1 and reduced Zn enzyme activities, which were attributed to the decreased Zn content. Moreover, loss of Mn efflux transport protein decreased MT and ZNT1 expression and Zn enzyme activity without changing extracellular Mn content. This reduction was not observed when supplementing with the same Cu concentrations and in cells lacking Cu efflux proteins. Furthermore, cellular Zn homeostasis was oppositely regulated in cells expressing Zn and Mn importer ZIP8, depending on whether Zn or Mn concentration was elevated in the extracellular milieu. Our results provide novel insights into the intricate interactions between Mn and Zn homeostasis in mammalian cells and facilitate our understanding of the physiopathology of Mn, which may lead to the development of treatment strategies for Mn‐related diseases in the future. [ABSTRACT FROM AUTHOR]

Aspergillus oryzae , also known as the yellow koji mold, produces various hydrolytic enzymes that are widely used in different industries. Its high capacity to produce secretory proteins makes this filamentous fungus a suitable host for heterologous protein production. Amylolytic gene promoter is widely used to express heterologous genes in A. oryzae. The expression of this promoter is strictly regulated by several transcription factors, whose activation involves various factors. Furthermore, the expression levels of amylolytic and heterologous genes are post-transcriptionally regulated by mRNA degradation mechanisms in response to aberrant transcriptional termination or endoplasmic reticulum stress. This review discusses the transcriptional and post-transcriptional regulatory mechanisms controlling the expression of genes encoding secretory proteins in A. oryzae. [ABSTRACT FROM AUTHOR]

Diatoms are unicellular algae with morphologically diverse silica cell walls, which are called frustules. The mechanism of frustule morphogenesis has attracted attention in biology and nanomaterials engineering. However, the genetic regulation of the morphology remains unclear. We therefore used transcriptome sequencing to search for genes involved in frustule morphology in the centric diatom Pleurosira laevis, which exhibits morphological plasticity between flat and domed valve faces in salinity 2 and 7, respectively. We observed differential expression of transposable elements (TEs) and transporters, likely due to osmotic response. Up‐regulation of mechanosensitive ion channels and down‐regulation of Ca2+‐ATPases in cells with flat valves suggested that cytosolic Ca2+ levels were changed between the morphologies. Calcium signaling could be a mechanism for detecting osmotic pressure changes and triggering morphological shifts. We also observed an up‐regulation of ARPC1 and annexin, involved in the regulation of actin filament dynamics known to affect frustule morphology, as well as the up‐regulation of genes encoding frustule‐related proteins such as BacSETs and frustulin. Taken together, we propose a model in which salinity‐induced morphogenetic changes are driven by upstream responses, such as the regulation of cytosolic Ca2+ levels, and downstream responses, such as Ca2+‐dependent regulation of actin dynamics and frustule‐related proteins. This study highlights the sensitivity of euryhaline diatoms to environmental salinity and the role of active cellular processes in controlling gross valve morphology under different osmotic pressures. [ABSTRACT FROM AUTHOR]

(1) Background: In hereditary creatine transporter deficiency (CTD), there is an absence of creatine in the brain and neurological symptoms are present, including severe language impairment. However, the pathological changes caused by creatine deficiency that generate neuropsychological symptoms have been poorly studied. (2) Aims: To investigate if the language impairment in CTD is underpinned by possible pathological changes. (3) Methods: We used MRI tractography to investigate the trophism of the left arcuate fasciculus, a white matter bundle connecting Wernicke's and Broca's language areas that is specifically relevant for language establishment and maintenance, in two patients (28 and 18 y.o.). (4) Results: The T1 and T2 MRI imaging results were unremarkable, but the left arcuate fasciculus showed a marked decrease in mean fractional anisotropy (FA) compared to healthy controls. In contrast, the FA values in the corticospinal tract were similar to those of healthy controls. Although white matter atrophy has been reported in CTD, this is the first report to show a selective abnormality of the language-relevant arcuate fasciculus, suggesting a possible region-specific impact of creatine deficiency. [ABSTRACT FROM AUTHOR]