Publications of NIPGR Scientists
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Item The Mediator complex subunit, OsMED26_2, modulates plant growth, seed set and seed traits related to starch quality in rice(Elsevier B.V., 2026) Prusty, Ankita; Malik, Naveen; Ranjan, Rajeev; Agarwal, Pinky; Parida, Swarup K.; Kapoor, Sanjay; Tyagi, Akhilesh K.The Mediator (MED) complex is a multi-subunit structure crucial for RNA polymerase II-dependent transcription in eukaryotes. In this study, we investigated the function of a seed-preferential subunit of the rice Mediator complex, namely, OsMED26_2, for the first time. Knockdown of OsMED26_2 in rice reduced plant height and altered panicle morphology with shorter panicles, lesser branching, and fewer seeds per panicle. OsMED26_2 knockdown also led to shorter grains with shorter length and chalky endosperm. A significantly higher percentage of grains with chalkiness (PGWC) and degree of chalky endosperm (DCE) was observed in OsMED26_2 knockdown lines. OsMED26_2-knockdown seeds contained lower starch levels and altered proportions of amylose and amylopectin. Scanning electron microscopy further showed that these changes caused irregular, round, and loosely packed starch granules in the endosperm, contributing to the chalky phenotype. Decreased amylose content and increased grain chalkiness were corroborated by the downregulation of the Waxy (Wx) gene, which is involved in amylose synthesis, and altered expression of AMY3A, CHALK5, FLO4, GPA3, and SUSY3 genes, which regulate grain chalkiness. Our findings demonstrate that OsMED26_2 is critical in regulating panicle architecture, impacting yield, and modulating starch level and composition to control grain chalkiness and thereby suggesting its functional significance especially in manipulating yield attributing grain cooking quality traits of rice.Item Cytological, transcriptome and miRNome temporal landscapes decode enhancement of rice grain size(BioMed Central Ltd, 2023) Mahto, Arunima; Yadav, Antima; Aswathi, P. V.; Parida, Swarup K.; Tyagi, Akhilesh K.; Agarwal, PinkyBackground Rice grain size (GS) is an essential agronomic trait. Though several genes and miRNA modules influencing GS are known and seed development transcriptomes analyzed, a comprehensive compendium connecting all possible players is lacking. This study utilizes two contrasting GS indica rice genotypes (small-grained SN and large-grained LGR). Rice seed development involves five stages (S1–S5). Comparative transcriptome and miRNome atlases, substantiated with morphological and cytological studies, from S1–S5 stages and flag leaf have been analyzed to identify GS proponents. Results Histology shows prolonged endosperm development and cell enlargement in LGR. Stand-alone and comparative RNAseq analyses manifest S3 (5–10 days after pollination) stage as crucial for GS enhancement, coherently with cell cycle, endoreduplication, and programmed cell death participating genes. Seed storage protein and carbohydrate accumulation, cytologically and by RNAseq, is shown to be delayed in LGR. Fourteen transcription factor families influence GS. Pathway genes for four phytohormones display opposite patterns of higher expression. A total of 186 genes generated from the transcriptome analyses are located within GS trait-related QTLs deciphered by a cross between SN and LGR. Fourteen miRNA families express specifically in SN or LGR seeds. Eight miRNA-target modules display contrasting expressions amongst SN and LGR, while 26 (SN) and 43 (LGR) modules are differentially expressed in all stages. Conclusions Integration of all analyses concludes in a “Domino effect” model for GS regulation highlighting chronology and fruition of each event. This study delineates the essence of GS regulation, providing scope for future exploits. The rice grain development database (RGDD) ( www.nipgr.ac.in/RGDD/index.php; https://doi.org/10.5281/zenodo.7762870) has been developed for easy access of data generated in this paper.Item OsCPK29 interacts with MADS68 to regulate pollen development in rice(Elsevier B.V., 2022) Ranjan, Rajeev; Malik, Naveen; Sharma, Shivam; Agarwal, Pinky; Kapoor, Sanjay; Tyagi, Akhilesh K.Pollen development and its germination are obligatory for the reproductive success of flowering plants. Calcium-dependent protein kinases (CPKs, also known as CDPKs) regulate diverse signaling pathways controlling plant growth and development. Here, we report the functional characterization of a novel OsCPK29 from rice, which is mainly expressed during pollen maturation stages of the anther. OsCPK29 exclusively localizes in the nucleus, and its N-terminal variable domain is responsible for retaining it in the nucleus. OsCPK29 knockdown rice plants exhibit reduced fertility, set fewer seeds, and produce collapsed non-viable pollen grains that do not germinate. Cytological analysis of anther semi-thin sections during different developmental stages suggested that pollen abnormalities appear after the vacuolated pollen stage. Detailed microscopic study of pollen grains further revealed that they were lacking the functional intine layer although exine layer was present. Consistent with that, downregulation of known intine development-related rice genes was also observed in OsCPK29 silenced anthers. Furthermore, it has been demonstrated that OsCPK29 interacts in vitro as well as in vivo with the MADS68 transcription factor which is a known regulator of pollen development. Therefore, phenotypic observations and molecular studies suggest that OsCPK29 is an important regulator of pollen development in rice.Item Genome-wide analysis of polymorphisms identified domestication-associated long low diversity region carrying important rice grain size/weight QTL(John Wiley & Sons, 2020) Kumar, Angad; Daware, Anurag; Kumar, Arvind; Kumar, Vinay; Krishnan S, Gopala; Mondal, Subhasish; Patra, Bhaskar Chandra; Singh, Ashok. K.; Tyagi, Akhilesh K.; Parida, Swarup K.; Thakur, Jitendra K.Rice grain size and weight are major determinants of grain quality and yield and so have been under rigorous selection since domestication. However, genetic basis for contrasting grain size/weight trait among Indian germplasms and their association with domestication‐driven evolution is not well understood. In this study, two long (LGG) and two short grain (SGG) genotypes were resequenced. LGG (LGR and PB 1121) differentiated from SGG (Sonasal and Bindli) by 504,439 SNPs and 78,166 InDels. The LRK gene cluster was different and a truncation mutation in the LRK8 kinase domain was associated with LGG. Phylogeny with 3000 diverse rice accessions revealed that the four sequenced genotypes belonged to japonica group and were at the edge of the clades indicating them to be the potential source of genetic diversity available in Indian rice germplasm. Six SNPs were significantly associated with grain size/weight and top four of them could be validated in mapping population, suggesting this study as a valuable resource for high‐throughput genotyping. A contiguous ~ 6 Mb long low diversity region (LDR) carrying a major grain weight QTL (harbouring OsTOR gene) was identified on chromosome 5. This LDR was identified as an evolutionary important site with significant positive selection and multiple selection sweeps, and showed association with many domestication‐related traits including grain size/weight. The aus population retained more allelic variations in the LDR than japonica and indica populations, suggesting it to be one of the divergence loci. All the data and analyses can be accessed from RiceSzWtBase database.Item Transcription factor OsNF-YB9 regulates reproductive growth and development in rice(Springer Nature Publishing AG, 2019) Das, Sweta; Parida, Swarup K.; Agarwal, Pinky; Tyagi, Akhilesh K.Nuclear Factor-Y (NF-Y) family of transcription factors takes part in many aspects of growth and development in eukaryotes. They have been classifed into three subunit classes, namely, NF-YA, NF-YB and NF-YC. In plants, this transcription factor family is much diverged and takes part in several developmental processes and stress. We investigated NF-Y subunit genes of rice (Oryza sativa) and found OsNF-YB9 as the closest homologue of LEAFY COTYLEDON1. OsNF-YB9 delayed the heading date when ectopically expressed in rice. Expression of several heading date regulating genes such as Hd1, Ehd1, Hd3a and RFT1 were altered. OsNF-YB9 overexpression also resulted in morphological defects in the reproductive organs and led to pseudovivipary. OsNF-YB9 interacted with MADS1, a key regulator of foral development. This NF-Y subunit acted upstream to several transcription factors as well as signalling proteins involved in brassinosteroid and gibberellic acid metabolism and cell cycle. OsNF-YB9 and OsNF-YC12 interacted in planta and the latter also delayed heading in rice upon overexpression suggesting its involvement in a similar pathway. Our data provide new insights into the rice heading date pathway integrating these OsNF-Y subunit members to the network. These features can be exploited to improve vegetative growth and yield of rice plants in future.Item Rice stress associated protein 1 (OsSAP1) interacts with aminotransferase (OsAMTR1) and pathogenesis-related 1a protein (OsSCP) and regulates abiotic stress responses(Frontiers Media S.A., 2016) Kothari, Kamakshi S.; Dansana, Prasant K.; Giri, Jitender; Tyagi, Akhilesh K.Stress associated proteins (SAPs) are the A20/AN1 zinc-finger containing proteins which can regulate the stress signaling in plants. The rice SAP protein, OsSAP1 has been shown to confer abiotic stress tolerance to plants, when overexpressed, by modulating the expression of endogenous stress-related genes. To further understand the mechanism of OsSAP1-mediated stress signaling, OsSAP1 interacting proteins were identified using yeast two-hybrid analysis. Two novel proteins, aminotransferase (OsAMTR1) and a SCP/TAPS or pathogenesis-related 1 class of protein (OsSCP) were found to interact with OsSAP1. The genes encoding OsAMTR1 and OsSCP were stress-responsive and showed higher expression upon abiotic stress treatments. The role of OsAMTR1 and OsSCP under stress was analyzed by overexpressing them constitutively in Arabidopsis and responses of transgenic plants were assessed under salt and water-deficit stress. The OsAMTR1 and OsSCP overexpressing plants showed higher seed germination, root growth and fresh weight than wild-type plants under stress conditions. Overexpression of OsAMTR1 and OsSCP affected the expression of many known stress-responsive genes which were not affected by the overexpression of OsSAP1. Moreover, the transcript levels of OsSCP and OsAMTR1 were also unaffected by the overexpression of OsSAP1. Hence, it was concluded that OsSAP1 regulates the stress responsive signaling by interacting with these proteins which further regulate the downstream stress responsive gene expression.Item Rice improvement through genome-based functional analysis and molecular breeding in India(Springer, 2016) Agarwal, Pinky; Parida, Swarup K.; Raghuvanshi, Saurabh; Kapoor, Sanjay; Khurana, Paramjit; Khurana, Jitendra P.; Tyagi, Akhilesh K.Rice is one of the main pillars of food security in India. Its improvement for higher yield in sustainable agriculture system is also vital to provide energy and nutritional needs of growing world population, expected to reach more than 9 billion by 2050. The high quality genome sequence of rice has provided a rich resource to mine information about diversity of genes and alleles which can contribute to improvement of useful agronomic traits. Defining the function of each gene and regulatory element of rice remains a challenge for the rice community in the coming years. Subsequent to participation in IRGSP, India has continued to contribute in the areas of diversity analysis, transcriptomics, functional genomics, marker development, QTL mapping and molecular breeding, through national and multi-national research programs. These efforts have helped generate resources for rice improvement, some of which have already been deployed to mitigate loss due to environmental stress and pathogens. With renewed efforts, Indian researchers are making new strides, along with the international scientific community, in both basic research and realization of its translational impact.Item OsiSAP1 overexpression improves water-deficit stress tolerance in transgenic rice by affecting expression of endogenous stress-related genes(Springer, 2014) Dansana, Prasant K.; Kothari, Kamakshi S.; Vij, Shubha; Tyagi, Akhilesh K.OsiSAP1, an A20/AN1 zinc-finger protein, confers water-deficit stress tolerance at different stages of growth by affecting expression of several endogenous genes in transgenic rice. Transgenic lines have been generated from rice constitutively expressing OsiSAP1, an A20/AN1 zinc-finger containing stress-associated protein gene from rice, driven by maize UBIQUITIN gene promoter and evaluated for water-deficit stress tolerance at different stages of growth. Their seeds show early germination and seedlings grow better under water-deficit stress compared to non-transgenic (NT) rice. Leaves from transgenic seedlings showed lesser membrane damage and lipid peroxidation under water-deficit stress. Relatively lower rate of leaf water loss has been observed in detached intact leaves from transgenic plants during late vegetative stage. Delayed leaf rolling and higher relative water content were also observed in transgenic plants under progressive water-deficit stress during reproductive developmental stage. Although reduction in grain yield is observed under unstressed condition, the relative water-deficit stress-induced yield losses are lower in transgenic rice vis-à-vis NT plants thereby resulting in yield loss protection. Transcriptome analysis suggests that overexpression of OsiSAP1 in transgenic rice results in altered expression of several endogenous genes including those coding for transcription factors, membrane transporters, signaling components and genes involved in metabolism, growth and development. A total of 150 genes were found to be more than twofold up-regulated in transgenic rice of which 43 genes are known to be involved in stress response. Our results suggest that OsiSAP1 is a positive regulator of water-deficit stress tolerance in rice.Item Comprehensive genomic analysis and expression profiling of phospholipase C gene family during abiotic stresses and development in rice(PLOS, 2013) Singh, Amarjeet; Kanwar, Poonam; Pandey, Amita; Tyagi, Akhilesh K.; Sopory, Sudhir K.; Kapoor, Sanjay; Pandey, Girdhar K.BACKGROUND: Phospholipase C (PLC) is one of the major lipid hydrolysing enzymes, implicated in lipid mediated signaling. PLCs have been found to play a significant role in abiotic stress triggered signaling and developmental processes in various plant species. Genome wide identification and expression analysis have been carried out for this gene family in Arabidopsis, yet not much has been accomplished in crop plant rice. METHODOLOGY/PRINCIPAL FINDINGS: An exhaustive in-silico exploration of rice genome using various online databases and tools resulted in the identification of nine PLC encoding genes. Based on sequence, motif and phylogenetic analysis rice PLC gene family could be divided into phosphatidylinositol-specific PLCs (PI-PLCs) and phosphatidylcholine- PLCs (PC-PLC or NPC) classes with four and five members, respectively. A comparative analysis revealed that PLCs are conserved in Arabidopsis (dicots) and rice (monocot) at gene structure and protein level but they might have evolved through a separate evolutionary path. Transcript profiling using gene chip microarray and quantitative RT-PCR showed that most of the PLC members expressed significantly and differentially under abiotic stresses (salt, cold and drought) and during various developmental stages with condition/stage specific and overlapping expression. This finding suggested an important role of different rice PLC members in abiotic stress triggered signaling and plant development, which was also supported by the presence of relevant cis-regulatory elements in their promoters. Sub-cellular localization of few selected PLC members in Nicotiana benthamiana and onion epidermal cells has provided a clue about their site of action and functional behaviour. CONCLUSION/SIGNIFICANCE: The genome wide identification, structural and expression analysis and knowledge of sub-cellular localization of PLC gene family envisage the functional characterization of these genes in crop plants in near future.Item A 286 bp upstream regulatory region of a rice anther-specific gene, OSIPP3, confers pollen-specific expression in Arabidopsis(Springer, 2013) Khurana, Reema; Kathuria, Hitesh; Mukhopadhyay, Arnab; Kapoor, Sanjay; Tyagi, Akhilesh K.OSIPP3 gene (coding for pectin methylesterase inhibitor protein) was isolated from a pre-pollinated inflorescence-specific cDNA library by differential screening of stage-specific libraries from Oryza sativa. OSIPP3 is present in the genome of rice as a single copy gene. OSIPP3 gene was expressed exclusively in the pre-pollinated spikelets of rice. Upstream regulatory region (URR) of OSIPP3 was isolated and a series of 5'-deletions were cloned upstream of GUS reporter gene and were used to transform Arabidopsis. OSIPP3_del1 and del2 transgenic plants showed GUS expression in root, anther and silique, while OSIPP3_del3 showed GUS activity only in anthers and siliques. Pollen-specific expression was observed in case of plants harboring OSIPP3_del4 construct. It can, therefore, be concluded that the OSIPP3 URR between -178 and +108 bp is necessary for conferring pollen-specific expression in Arabidopsis.