@article {161132, title = {Immunofluorescence Staining and Microscopic Imaging of Plant Nuclei for Epigenetic Modifications.}, journal = {Methods Mol Biol}, volume = {2988}, year = {2026}, month = {2026}, pages = {83-92}, abstract = {

Histone posttranslational modifications (PTMs) and DNA methylation are the predominant epigenetic modifications on the chromatin that regulate gene expression. These modifications can be spatially resolved using microscopic examination of the nuclei with the help of commercially available antibodies. Here, we describe a detailed method to obtain intact nuclei from plant tissues and reproducibly immunostain the nuclei for specific chromatin marks and microscopic examination. This method can be readily extended to multiple plant species as the antibodies are raised against conserved epigenetic marks.

}, keywords = {Cell Nucleus, Chromatin, DNA Methylation, Epigenesis, Genetic, Epigenomics, Fluorescent Antibody Technique, Histones, Microscopy, Fluorescence, Plants, Protein Processing, Post-Translational}, issn = {1940-6029}, doi = {10.1007/978-1-0716-4981-7_8}, author = {Gandhivel, Vivek Hari-Sundar and Raju, Steffi and Shivaprasad, P V} } @article {160480, title = {OsWRKY53 dictates wound responses in rice through fine-tuning crosstalk between PEP and PSK-mediated signalling.}, journal = {Plant J}, volume = {123}, year = {2025}, month = {2025 Sep}, pages = {e70481}, abstract = {

Wounding is a major event during the multitude of stresses that plants face in their natural environment. Wound response is very dynamic and involves the integration of various regulatory networks culminating in successful wound-induced downstream signalling. Plants depend on endogenous molecular signals predominantly, small peptides, to initiate wound responses. Transcriptional response is paramount in dictating a successful wound response. Here we show the involvement of a WRKY transcription factor (TF) named OsWRKY53 that is upregulated upon wounding as well as wound-derived plant elicitor peptide (PEP), OsPep2 treatments. OsWRKY53 is involved in the positive regulation of gene expression of OsPep2-responsive genes. OsWRKY53 displays altered DNA occupancy in response to OsPep2 treatment over time, correlating with the altered gene expression. OsWRKY53 undergoes phosphorylation upon OsPep2 perception, and this contributes to the enhanced stability of the protein. Further, OsWRKY53 is involved in the simultaneous activation and suppression of OsPep2-responsive and phytosulfokine (PSK)-responsive genes, respectively. In agreement with these findings, perturbed expression of OsWRKY53 leads to compromised transcriptional responses post-wounding. Collectively, we establish that OsWRKY53 acts at the intersection of PEP and PSK-mediated transition of wound responses.

}, keywords = {Gene Expression Regulation, Plant, Oryza, Phosphorylation, Plant Proteins, Signal Transduction, Transcription Factors}, issn = {1365-313X}, doi = {10.1111/tpj.70481}, author = {Harshith, Chitthavalli Y and Palchaudhuri, Dipasmit and Dey, Riju and Raju, Steffi and Shivaprasad, Padubidri V} } @article {147285, title = {Wound-induced small-peptide-mediated signaling cascade, regulated by OsPSKR, dictates balance between growth and defense in rice.}, journal = {Cell Rep}, volume = {43}, year = {2024}, month = {2024 Jul 23}, pages = {114515}, abstract = {

Wounding is a general stress in plants that results from various pest and pathogenic infections in addition to environment-induced mechanical damages. Plants have sophisticated molecular mechanisms to recognize and respond to wounding, with those of monocots being distinct from dicots. Here, we show the involvement of two distinct categories of temporally separated, endogenously derived peptides, namely, plant elicitor peptides (PEPs) and phytosulfokine (PSK), mediating wound responses in rice. These peptides trigger a dynamic signal relay in which a receptor kinase involved in PSK perception named OsPSKR plays a major role. Perturbation of OsPSKR expression in rice leads to compromised development and constitutive autoimmune phenotypes. OsPSKR regulates the transitioning of defense to growth signals upon wounding. OsPSKR displays mutual antagonism with the OsPEPR1 receptor involved in PEP perception. Collectively, our work indicates the presence of a stepwise peptide-mediated signal relay that regulates the transition from defense to growth upon wounding in monocots.

}, keywords = {Gene Expression Regulation, Plant, Oryza, Peptides, Plant Diseases, Plant Proteins, Signal Transduction}, issn = {2211-1247}, doi = {10.1016/j.celrep.2024.114515}, author = {Harshith, Chitthavalli Y and Pal, Avik and Chakraborty, Monoswi and Nair, Ashwin and Raju, Steffi and Shivaprasad, Padubidri V} } @article {129504, title = {Plant polymerase IV sensitizes chromatin through histone modifications to preclude spread of silencing into protein-coding domains.}, journal = {Genome Res}, year = {2023}, month = {2023 Jun 05}, abstract = {

Across eukaryotes, gene regulation is manifested via chromatin states roughly distinguished as heterochromatin and euchromatin. The establishment, maintenance, and modulation of the chromatin states is mediated using several factors including chromatin modifiers. However, factors that avoid the intrusion of silencing signals into protein-coding genes are poorly understood. Here we show that a plant specific paralog of RNA polymerase (Pol) II, named Pol IV, is involved in avoidance of facultative heterochromatic marks in protein-coding genes, in addition to its well-established functions in silencing repeats and transposons. In its absence, H3K27 trimethylation (me3) mark intruded the protein-coding genes, more profoundly in genes embedded with repeats. In a subset of genes, spurious transcriptional activity resulted in small(s) RNA production, leading to post-transcriptional gene silencing. We show that such effects are significantly pronounced in rice, a plant with a larger genome with distributed heterochromatin compared with Our results indicate the division of labor among plant-specific polymerases, not just in establishing effective silencing via sRNAs and DNA methylation but also in influencing chromatin boundaries.

}, issn = {1549-5469}, doi = {10.1101/gr.277353.122}, author = {Vivek Hari Sundar G and Swetha, Chenna and Basu, Debjani and Pachamuthu, Kannan and Raju, Steffi and Chakraborty, Tania and Mosher, Rebecca A and Shivaprasad, P V} }