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Plant virus satellites are maintained by their associated helper viruses, and satellites influence viral pathogenesis. Diseases caused by geminivirus-betasatellite complexes can become epidemics and therefore have become a threat to economically important crops across the world. Here, we identified a novel molecular function of the betasatellite-encoded pathogenicity determinant βC1. The tomato leaf curl Patna betasatellite (ToLCPaB)-encoded βC1 protein was found to exhibit novel ATPase activity in the presence of the divalent metal ion cofactor MgCl

Plant virus satellites are maintained by their associated helper viruses, and satellites influence viral pathogenesis. Diseases caused by geminivirus-betasatellite complexes can become epidemics and therefore have become a threat to economically important crops across the world. Here, we identified a novel molecular function of the betasatellite-encoded pathogenicity determinant βC1. The tomato leaf curl Patna betasatellite (ToLCPaB)-encoded βC1 protein was found to exhibit novel ATPase activity in the presence of the divalent metal ion cofactor MgCl(2). Moreover, ATPase activity was confirmed to be ubiquitously displayed by βC1 proteins encoded by diverse betasatellites. Mutational and sequence analysis showed that conserved lysine/arginine residues at positions 49/50 and 91 of βC1 proteins are essential for their ATPase activity. Biochemical studies revealed that the DNA-binding activity of the βC1 protein was interfered with by the binding of ATP to the protein. Mutating arginine 91 of βC1 to alanine reduced its DNA-binding activity. The results of docking studies provided evidence for an overlap of the ATP-binding and DNA-binding regions of βC1 and for the importance of arginine 91 for both ATP-binding and DNA-binding activities. A mutant betasatellite with a specifically βC1-ATPase dominant negative mutation was found to induce symptoms on Nicotiana benthamiana plants similar to those induced by wild-type betasatellite infection. The ATPase function of βC1 was found to be negatively associated with geminivirus-betasatellite DNA accumulation, despite the positive influence of this ATPase function on the accumulation of replication-associated protein (Rep) and βC1 transcripts. IMPORTANCE Most satellites influence the pathogenesis of their helper viruses. Here, we characterized the novel molecular function of βC1, a nonstructural pathogenicity determinant protein encoded by a betasatellite. We demonstrated the display of ATPase activity by this βC1 protein. Additionally, we confirmed the ubiquitous display of ATPase activity by βC1 proteins encoded by diverse betasatellites. The lysine/arginine residues conserved at positions 49 and 91 of βC1 were found to be crucial for its ATPase function. DNA-binding activity of βC1 was found to be reduced in the presence of ATP. Inhibition of ATPase activity of βC1 in the presence of an excess concentration of cold ATP, GTP, CTP, or UTP suggested that the purified βC1 can also hydrolyze other cellular nucleoside triphosphates (NTPs) besides ATP in vitro. These results established the importance of the ATPase and DNA-binding activities of the βC1 protein in regulating geminivirus-betasatellite DNA accumulation in the infected plant cell.

Summary Geminivirus disease complexes potentially interfere with plants physiology and cause disastrous effects on a wide range of economically important crops throughout the world. Diverse geminivirus betasatellite associations exacerbate the epidemic threat for global food security. Our previous study showed that βC1, the pathogenicity determinant of geminivirus betasatellites induce symptom development by disrupting the ultrastructure and function of chloroplasts. Here we explored the betasatellite‐virus‐chloroplast interaction in the scope of viral pathogenesis as well as plant defence responses, using Nicotiana benthamiana—Radish leaf curl betasatellite (RaLCB) as the model system. We have shown an interaction between RaLCB‐encoded βC1 and one of the extrinsic subunit proteins of oxygen‐evolving complex of photosystem II both in vitro and in vivo. Further, we demonstrate a novel function of the Nicotiana benthamiana oxygen‐evolving enhancer protein 2 (PsbP), in that it binds DNA, including geminivirus DNA. Transient silencing of PsbP in N. benthamiana plants enhances pathogenicity and viral DNA accumulation. Overexpression of PsbP impedes disease development during the early phase of infection, suggesting that PsbP is involved in generation of defence response during geminivirus infection. In addition, βC1‐PsbP interaction hampers non‐specific binding of PsbP to the geminivirus DNA. Our findings suggest that betasatellite‐encoded βC1 protein accomplishes counter‐defence by physical interaction with PsbP reducing the ability of PsbP to bind geminivirus DNA to establish infection.

Tomato spotted wilt virus (TSWV, species Tomato spotted wilt orthotospovirus) is an economically important plant virus that infects multiple horticultural crops on a global scale. TSWV encodes a non-structural protein NSs that acts as a suppressor of host RNA silencing machinery during infection. Despite extensive structural and functional analyses having been carried out on TSWV NSs, its protein-interacting targets in host plants are still largely unknown. Here, we systemically investigated NSs-interacting proteins in Nicotiana benthamiana via affinity purification and mass spectrometry (AP-MS) analysis. Forty-three TSWV NSs-interacting candidates were identified in N. benthamiana. Gene Ontology (GO) and protein&ndash, protein interaction (PPI) network analyses were carried out on their closest homologs in tobacco (Nicotiana tabacum), tomatoes (Solanum lycopersicum) and Arabidopsis (Arabidopsis thaliana). The results showed that NSs preferentially interacts with plant defense-related proteins such as calmodulin (CaM), importin, carbonic anhydrase and two heat shock proteins (HSPs): HSP70 and HSP90. As two major nodes in the PPI network, CaM and importin subunit &alpha, were selected for the further verification of their interactions with NSs via yeast two-hybrid (Y2H) screening. Our work suggests that the downstream signaling, transportation and/or metabolic pathways of host-NSs-interacting proteins may play critical roles in NSs-facilitated TSWV infection.

Extraviral components that can influence the accumulation and pathogenesis of their associated helper viruses are known as 'satellites'. The maintenance of satellites requires their ability to associate with their helper viruses. Satellites can be categorized as either satellite viruses or satellite nucleic acids based on their ability to encode capsid proteins. Understanding the biology of satellites is important since they are pathogenic to a wide range of plant, animal, and yeast organisms. Most satellites influence the pathogenesis of their helper viruses by altering the interaction between the host and helper virus. However, the molecular mechanism that governs the trilateral interaction between host, satellites, and helper virus remains largely unexplored. This review comprehensively describes details of the association and interaction of helper viruses with satellite viruses, satellite RNAs, and satellite DNAs, and their implications for pathogenesis.

Under constant climatic change and the existing environment, plants are exposed to several adverse factors such as extreme temperatures, high salt, heavy metals (HMs), drought, and pathogens. Prolonged exposure to such adverse conditions results in phytotoxicity and overall retardation of growth and development. Excess of HMs is one of the significant factors that affects plant growth significantly. Plants develop several defense mechanisms at the cellular and physiological levels to survive HMs stress. However, these internal defense mechanisms are not enough to overcome excessive HMs stress. Plants generate various secondary messengers to activate cell signaling, which results in the activation of several transcriptional responses associated with plant defense. Different transcription factors (TFs) gene families such as WRKY, my elob lastosis (MYB), basic leucine zipper domain (bZIP), basic helix-loop-helix (bHLH), APETALA2/ethylene-responsive element binding protein (AP2/EREBP), Cys2-His2 (C2H2), receptor genes/proteins, and kinases, like mitogen-activated protein kinase (MAPK), play an essential role in the regulation of stress using hormone-mediated pathways under HMs stress. Additionally, phytohormones and their signaling network also contribute to regulating cellular functions at the molecular level to combat HMs stress. Several mechanisms have been explored to understand the defense mechanism of plants under HMs stress. However, many more advanced molecular approaches are required to explore these defense mechanisms. Herein, we have described OMICS’ approaches to improve the plants' internal defense system to counteract HMs stress.

Summary Begomoviruses have emerged as a group of plant pathogens that cause devastating diseases in a wide range of crops in tropical and subtropical regions of the world. Betasatellites, the circular single‐stranded DNA molecules with the size of almost half of that of the associated helper begomoviruses, are often essential for the production of typical disease symptoms in several virus‐host systems. Association of betasatellites with begomoviruses results in more severe symptoms in the plants and affects the yield of numerous crops leading to huge agroeconomic losses. βC1, the only protein encoded by betasatellites, plays a multifaceted role in the successful establishment of infection. This protein counteracts the innate defence mechanisms of the host, like RNA silencing, ubiquitin‐proteasome system and defence responsive hormones. In the last two decades, the molecular aspect of betasatellite pathogenesis has attracted much attention from the researchers worldwide, and reports have shown that βC1 protein aggravates the helper begomovirus disease complex by modulating specific host factors. This review discusses the molecular aspects of the pathogenesis of betasatellites, including various βC1‐host factor interactions and their effects on the suppression of defence responses of the plants.

Plants face a continuous threat of abiotic stresses under the changing environment. Because of climate change, water scarcity has been shown to be a significant environmental constraint on plant productivity. Droughts in particular have been shown to affect plant growth and development, lead to alteration in quality and quantity of crop production, and result in global food insecurity. Phytohormones are known to play critical roles in regulating diverse processes of plant adaption to a drought environment. They regulate cellular functions at molecular levels via various cell signaling. Among various phytohormones, abscisic acid (ABA) is known for its role in drought-stress tolerance in plants. Other phytohormones such as auxins, brassinosteroids (BRs), cytokinins (CK), ethylene (ET), gibberellins (GA), jasmonic acid (JA), and salicylic acid (SA) are also crucial in plant drought tolerance. Several plant growth-promoting microbes have been reported to enhance the phytohormone levels in plants to mitigate the negative effect of drought. However, the transgenic approach appears to be a boon to engineering the genes responsible for regulating phytohormones to develop a drought-tolerant trait. Expression analyses have revealed that genes encoding transcription factors such as bZIP11, DREB2, MYB14, MYB48, WRKY2, WRKY56, WRKY108715, and RD22 play a very crucial role in phytohormone mediated drought response. Furthermore, exogenous applications of phytohormones are shown to enhance endogenous phytohormones. This review highlights the most recent advancements in phytohormone-mediated drought tolerance in major crop plants.

The compound microsatellites consist of two or more individual microsatellites, originate from mutation or imperfection in simple repeat sequences. The reports on systematic analysis of the occurrence, size and density of compound microsatellite (cSSR) types are very rare. Our study indicates that cSSRs are clustered at specific regions in the begomovirus genomes. cSSRs were overrepresented in majority of begomovirus genomes indicating that they might have some functional significance. Further, non-random distribution pattern of cSSR in begomovirus genomes was significantly correlated with the recombination breakpoint positions in the genome. The analysis of cSSR regions in the viral genome indicates the presence of stem loop (hairpin) secondary structure. The significance of these findings in biology of geminiviruses is discussed based on our present understanding of recombination and repetitive DNA. To our knowledge, this is the first analysis suggesting the possible association between recombination and microsatellites in any viral genome.

Microsatellites are known to exhibit ubiquitous presence across all kingdoms of life including viruses. Members of the Caulimoviridae family severely affect growth of vegetable and fruit plants and reduce economic yield in diverse cropping systems worldwide. Here, we analyzed the nature and distribution of both simple and complex microsatellites present in complete genome of 44 species of Caulimoviridae. Our results showed, in all analyzed genomes, genome size and GC content had a weak influence on number, relative abundance and relative density of microsatellites, respectively. For each genome, mono- and dinucleotide repeats were found to be highly predominant and are overrepresented in genome of majority of caulimoviruses. AT/TA and GAA/AAG/AGA was the most abundant di- and trinucleotide repeat motif, respectively. Repeats larger than trinucleotide were rarely found in these genomes. Comparative study of occurrence, abundance and density of microsatellite among available RNA and DNA viral genomes indicated that simple repeats were least abundant in genomes of caulimoviruses. Polymorphic repeats even though rare were observed in the large intergenic region of the genome, indicating strand slippage and/or unequal recombination processes do occur in caulimoviruses. To our knowledge, this is the first analysis of microsatellites occurring in any dsDNA viral genome. Characterization of such variations in repeat sequences would be important in deciphering the origin, mutational processes, and role of repeat sequences in viral genomes.

Highlight A satellite DNA-encoded protein (βC1) is localized in the chloroplast. The intercellular events associated with βC1-induced photosynthetic inhibition and vein clearing symptom formation are discussed., Geminivirus infection often causes severe vein clearing symptoms in hosts. Recently a betasatellite has emerged as a key regulator of symptom induction. To understand the host–betasatellite interactions in the process of symptom development, a systematic study was carried out involving symptoms induced by a betasatellite associated with radish leaf curl disease (RaLCB) in Nicotiana benthamiana. It has been found that βC1 protein localized to chloroplasts of host cells, and RaLCB lacking βC1, which failed to produce symptoms, had no effect on chloroplast ultrastructure. Vein flecking induced by transiently expressed βC1 was associated with chloroplast ultrastructure. In addition, the betasatellite down-regulates expression of genes involved in chlorophyll biosynthesis as well as genes involved in chloroplast development and plastid translocation. Interestingly, the expression of key host genes involved in chlorophyll degradation remains unaffected. Betasatellite infection drastically reduced the numbers of active reaction centres and the plastoquinol pool size in leaves exhibiting vein clearing symptoms. Betasatellite-mediated impediments at different stages of chloroplast functionality affect the photosynthetic efficiency of N. benthamiana. To the best of the authors’ knowledge, this is the first evidence of a chloroplast-targeting protein encoded by a DNA virus which induces vein clearing and structurally and functionally damages chloroplasts in plants.