Browsing by Author "Badmi, Raghuram"

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Dynamic phosphorylation of miRNA biogenesis factor HYL1 by MPK3 involving nuclear–cytoplasmic shuttling and protein stability in Arabidopsis
(MDPI AG, 2022) Bhagat, Prakash Kumar; Verma, Deepanjali; Singh, Kirti; Badmi, Raghuram; Sharma, Deepika; Sinha, Alok Krishna
MicroRNAs (miRNAs) are one of the prime regulators of gene expression. The recruitment of hyponastic leaves 1 (HYL1), a double-stranded RNA binding protein also termed as DRB1, to the microprocessor complex is crucial for accurate primary-miRNA (pri-miRNA) processing and the accumulation of mature miRNA in Arabidopsis thaliana. In the present study, we investigated the role of the MAP kinase-mediated phosphorylation of AtHYL1 and its sub-cellular activity. AtMPK3 specifically phosphorylates AtHYL1 at the evolutionarily conserved serine-42 present at the Nterminal regions and plays an important role in its nuclear–cytosolic shuttling. Additionally, we identified that AtHYL1 is cleaved by trypsin-like proteases into an N-terminal fragment, which renders its subcellular activities. We, for the first time, report that the dimerization of AtHYL1 not only takes place in the nucleus, but also in the cytosol, and the C-terminal of AtHYL1 has a role in regulating its stability, as well as its subcellular localization. AtHYL1 is hyper-phosphorylated in mpk3 mutants, leading to higher stability and reduced degradation. Our data show that AtMPK3 is a negative regulator of AtHYL1 protein stability and that the AtMPK3-induced phosphorylation of AtHYL1 leads to its protein degradation.
Possible role of plant MAP kinases in the biogenesis and transcription regulation of rice microRNA pathway factors
(Elsevier B.V., 2018) Badmi, Raghuram; Sheikh, Arsheed Hussain; Bhagat, Prakash Kumar; Verma, Deepanjali; Noryang, Stanzin; Sinha, Alok Krishna
Signalling pathways play vital roles as determinants of almost all the molecular processes inside a eukaryotic cell. They are more often considered to be the link between extracellular and intracellular environmental cues. Gene silencing pathways have emerged to be involved in regulation of stress responses and developmental processes. However, very little is known about the crosstalk between signalling and silencing pathways and their influence on each other. The present work describes the effects of general protein kinase inhibitors and specific mitogen activated protein kinase (MAPK) pathway inhibitors on the components of microRNA pathway in rice. The kinase inhibitors significantly reduced the activities of miRNA biogenesis complex and changed the transcript expression of miRNA pathway factors. This study suggests a possible regulation of microRNA machinery by plant kinases and MAP kinases in particular.
Possible role of WRKY transcription factors in regulating immunity in Oryza sativa ssp. indica
(Elsevier B.V., 2021) Sheikh, Arsheed H.; Hussain, Rana M. Fraz; Tabassum, Naheed; Badmi, Raghuram; Marillonnet, Sylvestre; Scheel, Dierk; Lee, Justin; Sinha, Alok Krishna
Plants have developed a robust transcription machinery to combat potential pathogenic organisms. One of the hallmarks of early immune responses is the activation of the WRKY transcription factors post infection. Specific WRKYs proteins from Arabidopsis are known substrates of MAPK pathway to mediate the flg22 elicited early immunity. In the current study, using the Golden Gate cloning strategy, we aim to clone the entire WRKY transcription factor family from Oryza sativa ssp. indica consisting of more than 100 members and study their MAPK interaction and subsequent role in PTI. Using a reporter LUC assay in protoplasts we investigated the early defense responses in a few interesting OsWRKY candidates. Interestingly, we observed stringent regulation of WRKY expression in cells and their transcriptional expression only under specific stress responses. The phenomenon of gene expression regulation by intron retention (IR) was prevalently observed in rice WRKY transcripts. We could show the role of WRKY8, 24, and 77 in early defense responses. It was observed that WRKY24 enhanced the expression of early defense response marker genes like NHL10 while WRKY8 and WRKY77 supressed their expression. This study highlights the complicated mechanism by which OsWRKYs expression is possibly regulated and the distinctive roles of some individual members in plant immunity. At the same time this study serves as a cautionary warning for plant researchers to be mindful of the intron retention mechanism while cloning OsWRKYs.