10 results on '"Kambakam, Sekhar"'
Search Results
2. Rbbp4 loss disrupts neural progenitor cell cycle regulation independent of Rb and leads to Tp53 acetylation and apoptosis.
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Schultz‐Rogers, Laura E., Thayer, Michelle L., Kambakam, Sekhar, Wierson, Wesley A., Helmer, Jordan A., Wishman, Mark D., Wall, Kristen A., Greig, Jessica L., Forsman, Jaimie L., Puchhalapalli, Kavya, Nair, Siddharth, Weiss, Trevor J., Luiken, Jon M., Blackburn, Patrick R., Ekker, Stephen C., Kool, Marcel, and McGrail, Maura
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CELL cycle regulation ,PROGENITOR cells ,CELL cycle ,DEACETYLATION ,ACETYLATION ,APOPTOSIS ,RETINOBLASTOMA protein - Abstract
Background: Retinoblastoma binding protein 4 (Rbbp4) is a component of transcription regulatory complexes that control cell cycle gene expression. Previous work indicated that Rbbp4 cooperates with the Rb tumor suppressor to block cell cycle entry. Here, we use genetic analysis to examine the interactions of Rbbp4, Rb, and Tp53 in zebrafish neural progenitor cell cycle regulation and survival. Results: Rbbp4 is upregulated across the spectrum of human embryonal and glial brain cancers. Transgenic rescue of rbbp4 mutant embryos shows Rbbp4 is essential for zebrafish neurogenesis. Rbbp4 loss leads to apoptosis and γ‐H2AX in the developing brain that is suppressed by tp53 knockdown or maternal zygotic deletion. Mutant retinal neural precursors accumulate in M phase and fail to initiate G0 gene expression. rbbp4; rb1 mutants show an additive effect on the number of M phase cells. In rbbp4 mutants, Tp53 acetylation is detected; however, Rbbp4 overexpression did not rescue DNA damage‐induced apoptosis. Conclusion: Rbbp4 is necessary for neural progenitor cell cycle progression and initiation of G0 independent of Rb. Tp53‐dependent apoptosis in the absence of Rbpb4 correlates with Tp53 acetylation. Together these results suggest that Rbbp4 is required for cell cycle exit and contributes to neural progenitor survival through the regulation of Tp53 acetylation. Key Findings: Rbbp4 regulates neural progenitor cell cycle independent of RbRbbp4 loss leads to Tp53 acetylation, gamma‐H2AX labeling and apoptosis in neural progenitorsSuppression of gamma‐H2AX labeling and apoptosis by Tp53 knockdown in Rbbp4 mutants indicates Rbpp4 may induce apoptosis by transcriptional activation of Tp53 independent of the DNA damage response pathway [ABSTRACT FROM AUTHOR]
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- 2022
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3. Cre/lox regulated conditional rescue and inactivation with zebrafish UFlip alleles generated by CRISPR-Cas9 targeted integration.
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Fang Liu, Kambakam, Sekhar, Almeida, Maira P., Zhitao Ming, Welker, Jordan M., Wierson, Wesley A., Schultz-Rogers, Laura E., Ekker, Stephen C., Clark, Karl J., Essner, Jeffrey J., and McGrail, Maura
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ALLELES , *CRISPRS , *BRACHYDANIO , *GENE expression , *GENE targeting , *GENETIC vectors , *PHENOTYPES , *CHROMOSOME inversions - Abstract
The ability to regulate gene activity spatially and temporally is essential to investigate cell-type- specific gene function during development and in postembryonic processes and disease models. The Cre/lox system has been widely used for performing cell and tissue-specific conditional analysis of gene function in zebrafish. However, simple and efficient methods for isolation of stable, Cre/lox regulated zebrafish alleles are lacking. Here, we applied our GeneWeld CRISPR-Cas9 targeted integration strategy to generate floxed alleles that provide robust conditional inactivation and rescue. A universal targeting vector, UFlip, with sites for cloning short homology arms flanking a floxed 2A-mRFP gene trap, was integrated into an intron in rbbp4 and rb1. rbbp4off and rb1off integration alleles resulted in strong mRFP expression,>99% reduction of endogenous gene expression, and recapitulated known indel loss-of- function phenotypes. Introduction of Cre led to stable inversion of the floxed cassette, loss of mRFP expression, and phenotypic rescue. rbbp4on and rb1on integration alleles did not cause phenotypes in combination with a loss-of- function mutation. Addition of Cre led to conditional inactivation by stable inversion of the cassette, gene trapping and mRFP expression, and the expected mutant phenotype. Neural progenitor Cre drivers were used for conditional inactivation and phenotypic rescue to showcase how this approach can be used in specific cell populations. Together these results validate a simplified approach for efficient isolation of Cre/lox-responsive conditional alleles in zebrafish. Our strategy provides a new toolkit for generating genetic mosaics and represents a significant advance in zebrafish genetics. [ABSTRACT FROM AUTHOR]
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- 2022
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4. Arabidopsis non‐host resistance PSS30 gene enhances broad‐spectrum disease resistance in the soybean cultivar Williams 82.
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Kambakam, Sekhar, Ngaki, Micheline N., Sahu, Binod B., Kandel, Devi R., Singh, Prashant, Sumit, Rishi, Swaminathan, Sivakumar, Muliyar‐Krishna, Rajesh, and Bhattacharyya, Madan K.
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SOYBEAN diseases & pests , *DISEASE resistance of plants , *FOLIC acid , *PHYTOPATHOGENIC microorganisms , *PHYTOPHTHORA sojae , *ARABIDOPSIS , *SOYBEAN cyst nematode - Abstract
Summary: Non‐host resistance (NHR), which protects all members of a plant species from non‐adapted or non‐host plant pathogens, is the most common form of plant immunity. NHR provides the most durable and robust form of broad‐spectrum immunity against non‐adaptive pathogens pathogenic to other crop species. In a mutant screen for loss of Arabidopsis (Arabidopsis thaliana) NHR against the soybean (Glycine max (L.) Merr.) pathogen Phytophthora sojae, the Phytophthora sojae‐susceptible 30 (pss30) mutant was identified. The pss30 mutant is also susceptible to the soybean pathogen Fusarium virguliforme. PSS30 encodes a folate transporter, AtFOLT1, which was previously localized to chloroplasts and implicated in the transport of folate from the cytosol to plastids. We show that two Arabidopsis folate biosynthesis mutants with reduced folate levels exhibit a loss of non‐host immunity against P. sojae. As compared to the wild‐type Col‐0 ecotype, the steady‐state folate levels are reduced in the pss1, atfolt1 and two folate biosynthesis mutants, suggesting that folate is required for non‐host immunity. Overexpression of AtFOLT1 enhances immunity of transgenic soybean lines against two serious soybean pathogens, the fungal pathogen F. virguliforme and the soybean cyst nematode (SCN) Heterodera glycines. Transgenic lines showing enhanced SCN resistance also showed increased levels of folate accumulation. This study thus suggests that folate contributes to non‐host plant immunity and that overexpression of a non‐host resistance gene could be a suitable strategy for generating broad‐spectrum disease resistance in crop plants. Significance Statement: Plant diseases are chronic deterrents of crop production, causing crop losses of billions of dollars annually. Here we identified the Arabidopsis non‐host resistance gene PSS30, which encodes a folate transporter. Expression of PSS30 in soybean (Glycine max) generates broad‐spectrum disease resistance against its two most serious pathogens, Fusarium virguliforme and Heterodera glycines, which together cause annual soybean yield losses valued at close to 2 billion dollars. [ABSTRACT FROM AUTHOR]
- Published
- 2021
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5. Tanscriptomic Study of the Soybean-Fusarium virguliforme Interaction Revealed a Novel Ankyrin-Repeat Containing Defense Gene, Expression of Whose during Infection Led to Enhanced Resistance to the Fungal Pathogen in Transgenic Soybean Plants.
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Ngaki, Micheline N., Wang, Bing, Sahu, Binod B., Srivastava, Subodh K., Farooqi, Mohammad S., Kambakam, Sekhar, Swaminathan, Sivakumar, and Bhattacharyya, Madan K.
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SOYBEAN disease & pest resistance ,FUSARIUM ,GENE expression in plants ,FUNGAL diseases of plants ,SOYBEAN ,PLANT breeding - Abstract
Fusarium virguliforme causes the serious disease sudden death syndrome (SDS) in soybean. Host resistance to this pathogen is partial and is encoded by a large number of quantitative trait loci, each conditioning small effects. Breeding SDS resistance is therefore challenging and identification of single-gene encoded novel resistance mechanisms is becoming a priority to fight this devastating this fungal pathogen. In this transcriptomic study we identified a few putative soybean defense genes, expression of which is suppressed during F. virguliforme infection. The F. virguliforme infection-suppressed genes were broadly classified into four major classes. The steady state transcript levels of many of these genes were suppressed to undetectable levels immediately following F. virguliforme infection. One of these classes contains two novel genes encoding ankyrin repeat-containing proteins. Expression of one of these genes, GmARP1, during F. virguliforme infection enhances SDS resistance among the transgenic soybean plants. Our data suggest that GmARP1 is a novel defense gene and the pathogen presumably suppress its expression to establish compatible interaction. [ABSTRACT FROM AUTHOR]
- Published
- 2016
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6. PTOX Mediates Novel Pathways of Electron Transport in Etioplasts of Arabidopsis.
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Kambakam, Sekhar, Bhattacharjee, Ujjal, Petrich, Jacob, and Rodermel, Steve
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ARABIDOPSIS , *ELECTRON transport , *PLASTOQUINONES , *PLANTS - Abstract
The immutans ( im ) variegation mutant of Arabidopsis defines the gene for PTOX (plastid terminal oxidase), a versatile plastoquinol oxidase in chloroplast membranes. In this report we used im to gain insight into the function of PTOX in etioplasts of dark-grown seedlings. We discovered that PTOX helps control the redox state of the plastoquinone (PQ) pool in these organelles, and that it plays an essential role in etioplast metabolism by participating in the desaturation reactions of carotenogenesis and in one or more redox pathways mediated by PGR5 (PROTON GRADIENT REGULATION 5) and NDH (NAD(P)H dehydrogenase), both of which are central players in cyclic electron transport. We propose that these elements couple PTOX with electron flow from NAD(P)H to oxygen, and by analogy to chlororespiration (in chloroplasts) and chromorespiration (in chromoplasts), we suggest that they define a respiratory process in etioplasts that we have termed “etiorespiration”. We further show that the redox state of the PQ pool in etioplasts might control chlorophyll biosynthesis, perhaps by participating in mechanisms of retrograde (plastid-to-nucleus) signaling that coordinate biosynthetic and photoprotective activities required to poise the etioplast for light development. We conclude that PTOX is an important component of metabolism and redox sensing in etioplasts. [ABSTRACT FROM AUTHOR]
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- 2016
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7. Humidity assay for studying plant-pathogen interactions in miniature controlled discrete humidity environments with good throughput.
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Zhen Xu, Huawei Jiang, Sahu, Binod Bihari, Kambakam, Sekhar, Singh, Prashant, Xinran Wang, Qiugu Wang, Bhattacharyya, Madan K., and Liang Dong
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PLANT-pathogen relationships ,HUMIDITY ,DRYING agents ,EVAPORATION (Meteorology) ,CONVECTION (Meteorology) ,PHENOTYPES - Abstract
This paper reports a highly economical and accessible approach to generate different discrete relative humidity conditions in spatially separated wells of a modified multi-well plate for humidity assay of plant-pathogen interactions with good throughput. We demonstrated that a discrete humidity gradient could be formed within a few minutes and maintained over a period of a few days inside the device. The device consisted of a freeway channel in the top layer, multiple compartmented wells in the bottom layer, a water source, and a drying agent source. The combinational effects of evaporation, diffusion, and convection were synergized to establish the stable discrete humidity gradient. The device was employed to study visible and molecular disease phenotypes of soybean in responses to infection by Phytophthora sojae, an oomycete pathogen, under a set of humidity conditions, with two near-isogenic soybean lines, Williams and Williams 82, that differ for a Phytophthora resistance gene (Rps1-k). Our result showed that at 63% relative humidity, the transcript level of the defense gene GmPR1 was at minimum in the susceptible soybean line Williams and at maximal level in the resistant line Williams 82 following P. sojae CC5C infection. In addition, we investigated the effects of environmental temperature, dimensional and geometrical parameters, and other configurational factors on the ability of the device to generate miniature humidity environments. This work represents an exploratory effort to economically and efficiently manipulate humidity environments in a space-limited device and shows a great potential to facilitate humidity assay of plant seed germination and development, pathogen growth, and plant-pathogen interactions. Since the proposed device can be easily made, modified, and operated, it is believed that this present humidity manipulation technology will benefit many laboratories in the area of seed science, plant pathology, and plant-microbe biology, where humidity is an important factor that influences plant disease infection, establishment, and development. [ABSTRACT FROM AUTHOR]
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- 2016
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8. Impaired Chloroplast Biogenesis in Immutans, an Arabidopsis Variegation Mutant, Modifies Developmental Programming, Cell Wall Composition and Resistance to Pseudomonas syringae.
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Pogorelko, Gennady V., Kambakam, Sekhar, Nolan, Trevor, Foudree, Andrew, Zabotina, Olga A., and Rodermel, Steven R.
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CHLOROPLASTS , *PLANT mutation , *ARABIDOPSIS , *PLANT cell walls , *PSEUDOMONAS syringae , *DISEASE resistance of plants - Abstract
The immutans (im) variegation mutation of Arabidopsis has green- and white- sectored leaves due to action of a nuclear recessive gene. IM codes for PTOX, a plastoquinol oxidase in plastid membranes. Previous studies have revealed that the green and white sectors develop into sources (green tissues) and sinks (white tissues) early in leaf development. In this report we focus on white sectors, and show that their transformation into effective sinks involves a sharp reduction in plastid number and size. Despite these reductions, cells in the white sectors have near-normal amounts of plastid RNA and protein, and surprisingly, a marked amplification of chloroplast DNA. The maintenance of protein synthesis capacity in the white sectors might poise plastids for their development into other plastid types. The green and white im sectors have different cell wall compositions: whereas cell walls in the green sectors resemble those in wild type, cell walls in the white sectors have reduced lignin and cellulose microfibrils, as well as alterations in galactomannans and the decoration of xyloglucan. These changes promote susceptibility to the pathogen Pseudomonas syringae. Enhanced susceptibility can also be explained by repressed expression of some, but not all, defense genes. We suggest that differences in morphology, physiology and biochemistry between the green and white sectors is caused by a reprogramming of leaf development that is coordinated, in part, by mechanisms of retrograde (plastid-to-nucleus) signaling, perhaps mediated by ROS. We conclude that variegation mutants offer a novel system to study leaf developmental programming, cell wall metabolism and host-pathogen interactions. [ABSTRACT FROM AUTHOR]
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- 2016
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9. Cover Image.
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Schultz‐Rogers, Laura E., Thayer, Michelle L., Kambakam, Sekhar, Wierson, Wesley A., Helmer, Jordan A., Wishman, Mark D., Wall, Kristen A., Greig, Jessica L., Forsman, Jaimie L., Puchhalapalli, Kavya, Nair, Siddharth, Weiss, Trevor J., Luiken, Jon M., Blackburn, Patrick R., Ekker, Stephen C., Kool, Marcel, and McGrail, Maura
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CELL cycle regulation ,APOPTOSIS ,PROGENITOR cells - Published
- 2022
- Full Text
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10. The mechanism of variegation in immutans provides insight into chloroplast biogenesis.
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Foudree, Andrew, Putarjunan, Aarthi, Kambakam, Sekhar, Nolan, Trevor, Fussell, Jenna, Pogorelko, Gennady, and Rodermel, Steve
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VARIEGATION ,ARABIDOPSIS thaliana ,PLASTIDS ,THYLAKOIDS ,CHLOROPLASTS ,CAROTENOIDS - Abstract
The immutans (im) variegation mutant of Arabidopsis has green and white-sectored leaves due to the absence of fully functional plastid terminal oxidase (PTOX), a plastoquinol oxidase in thylakoid membranes. PTOX appears to be at the nexus of a growing number of biochemical pathways in the plastid, including carotenoid biosynthesis, PSI cyclic electron flow, and chlororespiration. During the early steps of chloroplast biogenesis, PTOX serves as an alternate electron sink and is a prime determinant of the redox poise of the developing photosynthetic apparatus. Whereas a lack of PTOX causes the formation of photooxidized plastids in the white sectors of im, compensating mechanisms allow the green sectors to escape the effects of the mutation. This manuscript provides an update on PTOX, the mechanism of im variegation, and findings about im compensatory mechanisms. [ABSTRACT FROM AUTHOR]
- Published
- 2012
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