About

Diwaker Tripathi is a molecular plant pathologist with over 10 years of experience in host-pathogen biology. His research focuses on mitochondrial DNA damage caused by various stressors during development. He approaches his scientific investigations from the perspective of molecular biology, utilizing a variety of experimental techniques to analyze and develop solutions at the molecular level. Diwaker has gained extensive experience in experimental methods and has developed team-oriented communication skills through collaborative projects with other laboratories. His goal is to use his education and research background to analyze and establish biological solutions to a variety of disease-related problems in economically important crops.

Research topics

• Biochemistry
• Biology
• Botany
• Cell biology
• Microbiology
• Chemistry
• Genetics
• Immunology

Selected publications

• Oxidative and Glycation Damage to Mitochondrial DNA and Plastid DNA during Plant Development

  Antioxidants · 2023-04-06 · 14 citations

  reviewOpen access1st author

  Oxidative damage to plant proteins, lipids, and DNA caused by reactive oxygen species (ROS) has long been studied. The damaging effects of reactive carbonyl groups (glycation damage) to plant proteins and lipids have also been extensively studied, but only recently has glycation damage to the DNA in plant mitochondria and plastids been reported. Here, we review data on organellar DNA maintenance after damage from ROS and glycation. Our focus is maize, where tissues representing the entire range of leaf development are readily obtained, from slow-growing cells in the basal meristem, containing immature organelles with pristine DNA, to fast-growing leaf cells, containing mature organelles with highly-fragmented DNA. The relative contributions to DNA damage from oxidation and glycation are not known. However, the changing patterns of damage and damage-defense during leaf development indicate tight coordination of responses to oxidation and glycation events. Future efforts should be directed at the mechanism by which this coordination is achieved.

  PublisherOA PDFDOI

• The intrinsically disordered C‐terminus of purinoceptor <scp>P2K1</scp> fine‐tunes plant responses to extracellular <scp>ATP</scp>

  FEBS Letters · 2023-07-19 · 13 citations

  articleOpen access

  P2K1 is a plant‐specific purinoceptor that perceives extracellular ATP (eATP), a signaling molecular implicated in various physiological processes. Interestingly, P2K1 harbors a C‐terminal intrinsically disordered region (IDR). When we overexpressed a truncated P2K1 (P2K1 t ) lacking the IDR, primary root growth completely ceased in response to eATP. We investigated the functional roles of the IDR in P2K1 using a combination of molecular genetics, calcium imaging, gene expression analysis, and histochemical approaches. We found that the P2K1 t variant gave rise to an amplified response to eATP, through accumulation of superoxide, altered cell wall integrity, and ultimate cell death in the primary root tip. Together, these observations underscore the significant involvement of the C‐terminal tail of P2K1 in root growth regulation.

  PublisherOA PDFDOI

• Ribonucleotide and R-Loop Damage in Plastid DNA and Mitochondrial DNA during Maize Development

  Plants · 2023-09-02 · 2 citations

  articleOpen access1st author

  Although the temporary presence of ribonucleotides in DNA is normal, their persistence represents a form of DNA damage. Here, we assess such damage and damage defense to DNA in plastids and mitochondria of maize. Shoot development proceeds from meristematic, non-pigmented cells containing proplastids and promitochondria at the leaf base to non-dividing green cells in the leaf blade containing mature organelles. The organellar DNAs (orgDNAs) become fragmented during this transition. Previously, orgDNA damage and damage defense of two types, oxidative and glycation, was described in maize, and now a third type, ribonucleotide damage, is reported. We hypothesized that ribonucleotide damage changes during leaf development and could contribute to the demise of orgDNAs. The levels of ribonucleotides and R-loops in orgDNAs and of RNase H proteins in organelles were measured throughout leaf development and in leaves grown in light and dark conditions. The data reveal that ribonucleotide damage to orgDNAs increased by about 2- to 5-fold during normal maize development from basal meristem to green leaf and when leaves were grown in normal light conditions compared to in the dark. During this developmental transition, the levels of the major agent of defense, RNase H, declined. The decline in organellar genome integrity during maize development may be attributed to oxidative, glycation, and ribonucleotide damages that are not repaired.

  PublisherOA PDFDOI

• Analysis of the Plastid Genome Sequence During Maize Seedling Development

  Frontiers in Genetics · 2022-04-26

  articleOpen access1st authorCorresponding

  Shoot development in maize progresses from small, non-pigmented meristematic cells to expanded cells in the green leaf. During this transition, large plastid DNA (ptDNA) molecules in proplastids become fragmented in the photosynthetically-active chloroplasts. The genome sequences were determined for ptDNA obtained from Zea mays B73 plastids isolated from four tissues: base of the stalk (the meristem region); fully-developed first green leaf; first three leaves from light-grown seedlings; and first three leaves from dark-grown (etiolated) seedlings. These genome sequences were then compared to the Z. mays B73 plastid reference genome sequence that was previously obtained from green leaves. The assembled plastid genome was identical among these four tissues to the reference genome. Furthermore, there was no difference among these tissues in the sequence at and around the previously documented 27 RNA editing sites. There were, however, more sequence variants (insertions/deletions and single-nucleotide polymorphisms) for leaves grown in the dark than in the light. These variants were tightly clustered into two areas within the inverted repeat regions of the plastid genome. We propose a model for how these variant clusters could be generated by replication-transcription conflict.

  PublisherDOI

• Glycation damage to organelles and their DNA increases during maize seedling development

  Scientific Reports · 2022-02-17 · 7 citations

  articleOpen access1st authorCorresponding

  Shoot development in maize begins when meristematic, non-pigmented cells at leaf base stop dividing and proceeds toward the expanded green cells of the leaf blade. During this transition, promitochondria and proplastids develop into mature organelles and their DNA becomes fragmented. Changes in glycation damage during organelle development were measured for protein and DNA, as well as the glycating agent methyl glyoxal and the glycation-defense protein DJ-1 (known as Park7 in humans). Maize seedlings were grown under normal, non-stressful conditions. Nonetheless, we found that glycation damage, as well as defenses against glycation, follow the same developmental pattern we found previously for reactive oxygen species (ROS): as damage increases, damage-defense measures decrease. In addition, light-grown leaves had more glycation and less DJ-1 compared to dark-grown leaves. The demise of maize organellar DNA during development may therefore be attributed to both oxidative and glycation damage that is not repaired. The coordination between oxidative and glycation damage, as well as damage-response from the nucleus is also discussed.

  PublisherOA PDFDOI

• Activation of indolic glucosinolate pathway by extracellular ATP in Arabidopsis

  PLANT PHYSIOLOGY · 2022 · 24 citations

  • Biology
  • Cell biology
  • Botany

  Dear Editor, Extracellular ATP (eATP) is a purinergic signal recognized by plasma membrane-localized transmembrane receptors, such as P2X or P2Y receptors found in mammals (Verkhratsky and Burnstock, 2014), and the P2K receptors found in plants (Choi et al., 2014; Pham et al., 2020). In mammals, eATP and purinoceptors are the basis of intercellular signaling used to regulate diverse processes including neuronal signaling, apoptosis, and inflammation (Verkhratsky and Burnstock, 2014). In plants, eATP and P2K1 appear to influence root growth (Weerasinghe et al., 2009; Zhu et al., 2017), but are currently best characterized as a damage associated molecular pattern signal and recognition system contributing to plant defense (Tanaka et al., 2014). In this study, we found that the indolic glucosinolate pathway is induced by eATP in Arabidopsis (Arabidopsis thaliana) and may play a role as a biochemical defense when plants are damaged by pathogens and herbivores. Accumulating evidence from our group and others suggests that eATP is released to trigger plant responses to various biotic stresses and touch/wounding (Dark et al., 2011; Ramachandran et al., 2019). Recently, the P2K1 overexpression line OxP2K1 was reported to enhance plant resistance against various foliar pathogens, such as Phytophthora brassicae (biotrophic oomycete), Pseudomonas syringae (hemibiotrophic bacterium), Botrytis cinerea (necrotrophic fungus), and Rhizoctonia solani (necrotrophic root fungus), whereas a knockout mutant line of P2K1, dorn1-3, showed increased susceptibility (Bouwmeester et al., 2011; Wang et al., 2014, 2016; Balagué et al., 2017; Chen et al., 2017; Tripathi et al., 2018; Kumar et al., 2020). As shown in Table 1 and Supplemental Figures S1–S4, a similar trend was observed with Sclerotinia sclerotiorum (necrotrophic fungus) and Phytophthora capsici (hemi-biotrophic oomycete), overexpression of P2K1 made the plants more resistant to the parasitic nematode Meloidogyne javanica, and, interestingly, pre-treatment of Arabidopsis with eATP made the plants more resistant to the turnip mosaic virus (TuMV). These data suggest that eATP plays an important role in plant defense against a broad range of pathogens. Furthermore, OxP2K1 showed enhanced resistance against Pieris rapae (specialist) and Spodoptera exigua (generalist) (Supplemental Figure S5), suggesting that eATP also plays an important role in the plant defense response against wounding caused by herbivorous insects. How eATP signaling contributes to defense against varied pests is not well understood.

  PublisherOA PDFDOI

• Reactive Oxygen Species, Antioxidant Agents, and DNA Damage in Developing Maize Mitochondria and Plastids

  Frontiers in Plant Science · 2020 · 43 citations

  1st authorCorresponding
  • Botany
  • Biology
  • Chemistry

  Maize shoot development progresses from non-pigmented meristematic cells at the base of the leaf to expanded and non-dividing green cells of the leaf blade. This transition is accompanied by the conversion of promitochondria and proplastids to their mature forms and massive fragmentation of both mitochondrial DNA (mtDNA) and plastid DNA (ptDNA), collectively termed organellar DNA (orgDNA). We measured developmental changes in reactive oxygen species (ROS), which at high concentrations can lead to oxidative stress and DNA damage, as well as antioxidant agents and oxidative damage in orgDNA. Our plants were grown under normal, non-stressful conditions. Nonetheless, we found more oxidative damage in orgDNA from leaf than stalk tissues and higher levels of hydrogen peroxide, superoxide, and superoxide dismutase in leaf than stalk tissues and in light-grown compared to dark-grown leaves. In both mitochondria and plastids, activities of the antioxidant enzyme peroxidase were higher in stalk than in leaves and in dark-grown than light-grown leaves. In protoplasts, the amount of the small-molecule antioxidants, glutathione and ascorbic acid, and catalase activity were also higher in the stalk than in leaf tissue. The data suggest that the degree of oxidative stress in the organelles is lower in stalk than leaf and lower in dark than light growth conditions. We speculate that the damaged/fragmented orgDNA in leaves (but not the basal meristem) results from ROS signaling to the nucleus to stop delivering DNA repair proteins to mature organelles producing large amounts of ROS.

  PublisherDOI

• Purinoceptor P2K1/DORN1 Enhances Plant Resistance Against a Soilborne Fungal Pathogen, Rhizoctonia solani

  Frontiers in Plant Science · 2020 · 34 citations

  • Biology
  • Microbiology
  • Botany

  relative to wild type, indicating that the salicylate and jasmonate defense signaling pathways functioned in resistance. These results indicated that a DAMP-mediated defense system confers basal resistance against an important root necrotrophic fungal pathogen.

  PublisherDOI

• In silico Prediction and Validations of Domains Involved in Gossypium hirsutum SnRK1 Protein Interaction With Cotton Leaf Curl Multan Betasatellite Encoded βC1

  Frontiers in Plant Science · 2019-05-28 · 26 citations

  articleOpen access

  Cotton leaf curl disease (CLCuD) caused by viruses of genus Begomovirus is a major constraint to cotton (Gossypium hirsutum) production in many cotton-growing regions of the world. Symptoms of the disease are caused by Cotton leaf curl Multan betasatellite (CLCuMB) that encodes a pathogenicity determinant protein, βC1. Here, we report the identification of interacting regions in βC1 protein first by using computational approaches including sequence recognition, and binding site and interface prediction methods. We show the domain-level interactions based on the structural analysis of G. hirsutum SnRK1 protein and its domains with CLCuMB-βC1. To verify and validate the in-silico predictions, three different experimental approaches, yeast two hybrid, bimolecular fluorescence complementation and pull down assay were used. These results showed that ubiquitin-associated domain (UBA) and autoinhibitory sequence (AIS) domains of G. hirsutum-encoded SnRK1 are involved in CLCuMB-βC1 interaction. This is the first comprehensive investigation that combined in-silico interaction prediction followed by experimental validation of interaction between CLCuMB-βC1 and a host protein. We demonstrated that data from computational biology could provide binding site information between CLCuD-associated viruses/satellites and new hosts that lack known binding site information for protein-protein interaction studies. Implications of these findings are discussed.

  PublisherDOI

• βC1, pathogenicity determinant encoded by Cotton leaf curl Multan betasatellite, interacts with calmodulin-like protein 11 (Gh-CML11) in Gossypium hirsutum

  PLoS ONE · 2019-12-03 · 16 citations

  articleOpen accessCorresponding

  Begomoviruses interfere with host plant machinery to evade host defense mechanism by interacting with plant proteins. In the old world, this group of viruses are usually associated with betasatellite that induces severe disease symptoms by encoding a protein, βC1, which is a pathogenicity determinant. Here, we show that βC1 encoded by Cotton leaf curl Multan betasatellite (CLCuMB) requires Gossypium hirsutum calmodulin-like protein 11 (Gh-CML11) to infect cotton. First, we used the in silico approach to predict the interaction of CLCuMB-βC1 with Gh-CML11. A number of sequence- and structure-based in-silico interaction prediction techniques suggested a strong putative binding of CLCuMB-βC1 with Gh-CML11 in a Ca+2-dependent manner. In-silico interaction prediction was then confirmed by three different experimental approaches: The Gh-CML11 interaction was confirmed using CLCuMB-βC1 in a yeast two hybrid system and pull down assay. These results were further validated using bimolecular fluorescence complementation system showing the interaction in cytoplasmic veins of Nicotiana benthamiana. Bioinformatics and molecular studies suggested that CLCuMB-βC1 induces the overexpression of Gh-CML11 protein and ultimately provides calcium as a nutrient source for virus movement and transmission. This is the first comprehensive study on the interaction between CLCuMB-βC1 and Gh-CML11 proteins which provided insights into our understating of the role of βC1 in cotton leaf curl disease.

  PublisherOA PDFDOI

Frequent coauthors

• N. G. P. Rao

  10 shared

• H. R. Pappu

  Washington State University

  7 shared

• B. S. Rana

  Panjab University

  7 shared

• Kiwamu Tanaka

  Washington State University

  6 shared

• N. G. P. Rao

  International Maize and Wheat Improvement Center

  6 shared

• Hira Kamal

  6 shared

• Delene J. Oldenburg

  University of Washington

  5 shared

• Arnold J. Bendich

  University of Washington

  5 shared

Education

• PhD, Plant Pathology

  Washington State University

  2014

• MS, Biology

  East Tennessee State University

  2010