@article {128071, title = {Begomoviral beta C1 orchestrates organellar genomic instability to augment viral infection}, journal = {PLANT JOURNAL }, year = {2023}, month = {05/2023}, abstract = {

Chloroplast is the site for transforming light energy to chemical energy. It also acts as a production unit for a variety of defense-related molecules. These defense moieties are necessary to mount a successful counter defense against pathogens, including viruses. Previous studies indicated disruption of chloroplast homeostasis as a basic strategy of Begomovirus for its successful infection leading to the production of vein-clearing, mosaic, and chlorotic symptoms in infected plants. Although begomoviral pathogenicity determinant protein Beta C1 (beta C1) was implicated for pathogenicity, the underlying mechanism was unclear. Here we show that, begomoviral beta C1 directly interferes with the host plastid homeostasis. beta C1 induced DPD1, an organelle-specific nuclease, implicated in nutrient salvage and senescence, as well as modulated the function of a major plastid genome maintainer protein RecA1, to subvert plastid genome. We show that beta C1 was able to physically interact with bacterial RecA and its plant homolog RecA1, resulting in its altered activity. We observed that knocking-down DPD1 during virus infection significantly reduced virus-induced necrosis. These results indicate the presence of a strategy in which a viral protein alters host defense by targeting modulators of chloroplast DNA. We predict that the mechanism identified here might have similarities in other plant-pathogen interactions.

}, doi = {10.1111/tpj.16186}, author = {Nair, Ashwin and Harshith, Chitthavalli Y. Y. and Narjala, Anushree and Shivaprasad, P V} } @article {126670, title = {Begomoviral βC1 orchestrates organellar genomic instability to augment viral infection.}, journal = {Plant J}, year = {2023}, month = {2023 Mar 14}, abstract = {

Chloroplast is the site for transforming light energy to chemical energy. It also acts as a production unit for a variety of defense-related molecules. These defense moieties are necessary to mount a successful counter defense against pathogens, including viruses. Previous studies indicated disruption of chloroplast homeostasis as a basic strategy of Begomovirus for its successful infection leading to the production of vein-clearing, mosaic, and chlorotic symptoms in infected plants. Although begomoviral pathogenicity determinant protein Beta C1 (βC1) was implicated for pathogenicity, the underlying mechanism was unclear. Here we show that, begomoviral βC1 directly interferes with the host plastid homeostasis. βC1 induced DPD1, an organelle-specific nuclease, implicated in nutrient salvage and senescence, as well as modulated the function of a major plastid genome maintainer protein RecA1, to subvert plastid genome. We show that βC1 was able to physically interact with bacterial RecA and its plant homolog RecA1, resulting in its altered activity. We observed that knocking-down DPD1 during virus infection significantly reduced virus-induced necrosis. These results indicate the presence of a strategy in which a viral protein alters host defense by targeting modulators of chloroplast DNA. We predict that the mechanism identified here might have similarities in other plant-pathogen interactions.

}, issn = {1365-313X}, doi = {10.1111/tpj.16186}, author = {Nair, Ashwin and Harshith, Chitthavalli Y and Narjala, Anushree and Shivaprasad, Padubidri V} } @article {112955, title = {Cultivar-specific miRNA-mediated RNA silencing in grapes.}, journal = {Planta}, volume = {256}, year = {2022}, month = {2022 Jun 23}, pages = {17}, abstract = {

MAIN CONCLUSION: In-depth comparative degradome analysis of two domesticated grape cultivars with diverse secondary metabolite accumulation reveals differential miRNA-mediated targeting. Small (s)RNAs such as micro(mi)RNAs and secondary small interfering (si) often work as negative switches of gene expression. In plants, it is well known that miRNAs target and cleave mRNAs that have high sequence complementarity. However, it is not known if there are variations in miRNA-mediated targeting between subspecies and cultivars that have been subjected to vast genetic modifications through breeding and other selections. Here, we have used PAREsnip2 tool for analysis of degradome datasets derived from two contrasting domesticated grape cultivars having varied fruit color, habit and leaf shape. We identified several interesting variations in sRNA targeting using degradome and 5{\textquoteright}RACE analysis between two contrasting grape cultivars that was further correlated using RNA-seq analysis. Several of the differences we identified are associated with secondary metabolic pathways. We propose possible means by which sRNAs might contribute to diversity in secondary metabolites and other development pathways between two domesticated cultivars of grapes.

}, keywords = {Gene Expression Regulation, Plant, MicroRNAs, Plant Breeding, RNA Interference, RNA, Plant, Sequence Analysis, RNA, Vitis}, issn = {1432-2048}, doi = {10.1007/s00425-022-03934-y}, author = {Tirumalai, Varsha and Narjala, Anushree and Swetha, Chenna and G Vivek Hari Sundar and Sujith, T N and Shivaprasad, P V} } @article {105482, title = {Degradome comparison between wild and cultivated rice identifies differential targeting by miRNAs.}, journal = {BMC Genomics}, volume = {23}, year = {2022}, month = {2022 Jan 14}, pages = {53}, abstract = {

BACKGROUND: Small non-coding (s)RNAs are involved in the negative regulation of gene expression, playing critical roles in genome integrity, development and metabolic pathways. Targeting of RNAs by ribonucleoprotein complexes of sRNAs bound to Argonaute (AGO) proteins results in cleaved RNAs having precise and predictable\ 5{\textquoteleft} ends. While tools to study sliced bits of RNAs to confirm the efficiency of sRNA-mediated regulation are available, they are sub-optimal. In this study, we provide an improvised version of a tool with better efficiency to accurately validate sRNA targets.

RESULTS: Here, we improvised the CleaveLand tool to identify additional micro (mi)RNA targets that belong to the same family and also other targets within a specified free energy cut-off. These additional targets were otherwise excluded during the default run. We employed these tools to understand the sRNA targeting efficiency in wild and cultivated rice, sequenced degradome from two rice lines, O. nivara and O. sativa indica Pusa Basmati-1 and analyzed variations in sRNA targeting. Our results indicate the existence of multiple miRNA-mediated targeting differences between domesticated and wild species. For example, Os5NG4 was targeted only in wild rice that might be responsible for the poor secondary wall formation when compared to cultivated rice. We also identified differential mRNA targets of secondary sRNAs that were generated after miRNA-mediated cleavage of primary targets.

CONCLUSIONS: We identified many differentially targeted mRNAs between wild and domesticated rice lines. In addition to providing a step-wise guide to generate and analyze degradome datasets, we showed how domestication altered sRNA-mediated cascade silencing during the evolution of indica rice.

}, keywords = {Argonaute Proteins, Base Sequence, MicroRNAs, Oryza, RNA, Messenger}, issn = {1471-2164}, doi = {10.1186/s12864-021-08288-5}, author = {Swetha, Chenna and Narjala, Anushree and Pandit, Awadhesh and Tirumalai, Varsha and Shivaprasad, P V} } @article {108050, title = {Nitrate-dependent regulation of miR444-OsMADS27 signaling cascade controls root development in rice.}, journal = {J Exp Bot}, year = {2022}, month = {2022 Mar 04}, abstract = {

Nitrate is an important nutrient and a key signaling molecule for plant development. A number of transcription factors involved in the process of nitrate response and their regulatory mechanisms have been identified. However, little is known about the nitrate sensor transcription factors and their regulatory mechanisms among crop plants. In this study, we identified functions of a nitrate-responsive miR444:MADS-box transcription factor OsMADS27 module and its downstream targets mediating rice root growth and stress responses. Transgenic rice plants expressing miR444 target mimic improved rice root growth. Although miR444 has the potential to target multiple genes, we identified OsMADS27 as the major miR444 target that regulates the expression of nitrate transporters, as well as several key genes including expansins and those associated with auxin signaling promote root growth. In agreement with this, overexpression of miRNA-resistant OsMADS27 improved root development and tolerance to stresses, while its silencing suppressed root growth. OsMADS27-mediated a robust stress tolerance in plants through its ability to bind to the promoters of specific stress regulators as observed in ChIP-Seq analysis. Our results provide evidence of nitrate-dependent miR444-OsMADS27 signaling cascade involved in the regulation of rice root growth as well as its surprising role in stress responses.

}, issn = {1460-2431}, doi = {10.1093/jxb/erac083}, author = {Pachamuthu, Kannan and Vivek Hari Sundar and Narjala, Anushree and Singh, Rahul R and Das, Soumita and Avik Pal, Harshith C Y and Shivaprasad, Padubidri V} } @article {99103, title = {Essential role of γ-clade RNA-dependent RNA polymerases in rice development and yield-related traits is linked to their atypical polymerase activities regulating specific genomic regions.}, journal = {New Phytol}, year = {2021}, month = {2021 Aug 27}, abstract = {

RNA-dependent RNA polymerases (RDR) generate double-stranded (ds)RNA triggers for RNA silencing across eukaryotes. Among the three clades, α- and β-clade members are key components of RNA silencing and mediators of stress responses across eukaryotes. However, γ-clade members are unusual in that they are represented in phylogenetically distant plants and fungi, and their functions are unknown. Using genetic, bioinformatic and biochemical methods, we show that γ-clade RDRs from Oryza sativa L. are involved in plant development as well as regulation of expression of coding and non-coding RNAs. Overexpression of γ-clade RDRs in transgenic rice and tobacco plants resulted in robust growth phenotype, whereas their silencing in rice displayed strong inhibition of growth. Small (s)RNA and RNA-seq analysis of OsRDR3 mis-expression lines suggested that it is specifically involved in the regulation of repeat-rich regions in the genome. Biochemical analysis confirmed that OsRDR3 has robust polymerase activities on both single stranded (ss)RNA and ssDNA templates similar to the activities reported for α-clade RDRs such as AtRDR6. Our results provide the first evidence of the importance of γ-clade RDRs in plant development, their atypical biochemical activities, and their contribution to the regulation of gene expression.

}, issn = {1469-8137}, doi = {10.1111/nph.17700}, author = {Jha, Vikram and Narjala, Anushree and Basu, Debjani and Sujith, T N and Pachamuthu, Kannan and Chenna, Swetha and Nair, Ashwin and Shivaprasad, Padubidri V} } @article {73040, title = {A conserved sequence signature is essential for robust plant miRNA biogenesis.}, journal = {Nucleic Acids Res}, year = {2020}, month = {2020 Feb 06}, abstract = {

Micro (mi)RNAs are 20-22nt long non-coding RNA molecules involved in post-transcriptional silencing of targets having high base-pair complementarity. Plant miRNAs are processed from long Pol II-transcripts with specific stem-loop structures by Dicer-like (DCL) 1 protein. Although there were reports indicating how a specific region is selected for miRNA biogenesis, molecular details were unclear. Here, we show that the presence of specific GC-rich sequence signature within miRNA/miRNA* region is required for the precise miRNA biogenesis. The involvement of GC-rich signatures in precise processing and abundance of miRNAs was confirmed through detailed molecular and functional analysis. Consistent with the presence of the miRNA-specific GC signature, target RNAs of miRNAs also possess conserved complementary sequence signatures in their miRNA binding motifs. The selection of these GC signatures was dependent on an RNA binding protein partner of DCL1 named HYL1. Finally, we demonstrate a direct application of this discovery for enhancing the abundance and efficiency of artificial miRNAs that are popular in plant functional genomic studies.

}, issn = {1362-4962}, doi = {10.1093/nar/gkaa077}, author = {Narjala, Anushree and Nair, Ashwin and Tirumalai, Varsha and G Vivek Hari Sundar and Shivaprasad, Padubidri V} }