Kishore Chiruvella - Academia.edu (original) (raw)

Papers by Kishore Chiruvella

Nature Communications

DNA double-strand breaks (DSBs) threaten genomic stability, since their persistence can lead to l... more DNA double-strand breaks (DSBs) threaten genomic stability, since their persistence can lead to loss of critical genetic information, chromosomal translocations or rearrangements, and cell death. DSBs can be repaired through the nonhomologous end-joining pathway (NHEJ), which processes and ligates DNA ends efficiently to prevent or minimize sequence loss. Polymerase λ (Polλ), one of the Family X polymerases, fills sequence gaps of DSB substrates with a strict specificity for a base-paired primer terminus. There is little information regarding Polλ’s approach to engaging such substrates. We used in vitro polymerization and cell-based NHEJ assays to explore the contributions of conserved loop regions toward DSB substrate specificity and utilization. In addition, we present multiple crystal structures of Polλ in synapsis with varying biologically relevant DSB end configurations, revealing how key structural features and hydrogen bonding networks work in concert to stabilize these tenuo...

Journal of Tropical Resources and Sustainable Science (JTRSS), May 3, 2015

Hempedu Bumi, Andrographis paniculata a popular Malay ethnomedicinal plant used to treat various ... more Hempedu Bumi, Andrographis paniculata a popular Malay ethnomedicinal plant used to treat various kinds of diseases like common cold, fever, liver disease and snake bite. Andrographolide a bitter compound present in the plant posses many medicinal properties like anti-inflammatory, antiviral, anti-HIV and promising anticancer activity. Large amount of the compound is being extracted from the plants so there is a need for commercial cultivation of this plant, but seed dormancy proved to be a major obstacle for this process. So production of non-dormant synthetic seeds is an alternative to the dormant natural seeds for the mass cultivation of this plant. Hence the present study is aimed to produce somatic embryos and synthetic seeds using plant tissue culture technique. The zygotic embryos from immature seeds of A. paniculata were inoculated on to MS media containing various auxins (NAA, IBA, 2, 4-D), and cytokinins (BAP, TDZ and Kinetin) for the induction of somatic embryos. Out of all the treatments used, only 1.5 mg/L NAA + 0.75 mg/L Kinetin produced somatic embryos after 6 weeks of culture and the number of somatic embryos per explant was 9.8 ± 0.6. NAA alone produced rhizogenic calli and NAA along with BAP produced hard friable white callus, 2, 4-D alone and 2, 4-D with cytokinins (BAP, Kinetin) produced creamy callus, while IBA alone and in combination with cytokinins (BAP, Kinetin), TDZ resulted in germination of embryos into seedlings. The somatic embryos produced were coated with sodium alginate solution and made into synthetic seeds. The synthetic seeds that have been produced can be used as planting material and for conservation of the plant for future uses.

Nucleic Acids Research, 2019

DNA double-strand breaks (DSBs) resulting from reactive oxygen species generated by exposure to U... more DNA double-strand breaks (DSBs) resulting from reactive oxygen species generated by exposure to UV and ionizing radiation are characterized by clusters of lesions near break sites. Such complex DSBs are repaired slowly, and their persistence can have severe consequences for human health. We have therefore probed DNA break repair containing a template 8-oxo-7,8-dihydro-2′-guanosine (8OG) by Family X Polymerase μ (Pol μ) in steady-state kinetics and cell-based assays. Pol μ tolerates 8OG-containing template DNA substrates, and the filled products can be subsequently ligated by DNA Ligase IV during Nonhomologous end-joining. Furthermore, Pol μ exhibits a strong preference for mutagenic bypass of 8OG by insertion of adenine. Crystal structures reveal that the template 8OG is accommodated in the Pol μ active site with none of the DNA substrate distortions observed for Family X siblings Pols β or λ. Kinetic characterization of template 8OG bypass indicates that Pol μ inserts adenosine nuc...

Scientific reports, Jan 23, 2016

Kluyveromyces lactis hAT-transposase 1 (Kat1) generates hairpin-capped DNA double strand breaks l... more Kluyveromyces lactis hAT-transposase 1 (Kat1) generates hairpin-capped DNA double strand breaks leading to MAT-switching (MATa to MATα). Using purified Kat1, we demonstrate the importance of terminal inverted repeats and subterminal repeats for its endonuclease activity. Kat1 promoted joining of the transposon end into a target DNA molecule in vitro, a biochemical feature that ties Kat1 to transposases. Gas-phase Electrophoretic Mobility Macromolecule analysis revealed that Kat1 can form hexamers when complexed with DNA. Kat1 point mutants were generated in conserved positions to explore structure-function relationships. Mutants of predicted catalytic residues abolished both DNA cleavage and strand-transfer. Interestingly, W576A predicted to be impaired for hairpin formation, was active for DNA cleavage and supported wild type levels of mating-type switching. In contrast, the conserved CXXH motif was critical for hairpin formation because Kat1 C402A/H405A completely blocked hairpinn...

Revista Brasileira de Farmacognosia, 2015

Drug resistance to human infectious diseases caused by pathogens lead to premature deaths through... more Drug resistance to human infectious diseases caused by pathogens lead to premature deaths through out the world. Plants are sources for wide variety of drugs used for treating various diseases. Systematic screening of medicinal plants for the search of new antimicrobial drug candidates that can inhibit the growth of pathogens or kill with no toxicity to host is being continued by many laboratories. Here we review the phytochemical investigations and biological activities of Meliaceae. The mahogany (Meliaceae) is family of timber trees with rich source for limonoids. So far, amongst the different members of Meliaceae, Azadirachta indica and Melia dubia have been identified as the potential plant systems possessing a vast array of biologically active compounds which are chemically diverse and structurally complex. Despite biological activities on different taxa of Meliaceae have been carried out, the information of antibacterial and antifungal activity is a meager with exception to Azadirachta indica. Together we provide new insights of Meliaceae members demonstrating as a potential source as antimicrobial agents using in vitro studies.

This review focuses on cinchona/oak bark substitute, Soymida febrifuga (Meliaceae), also known as... more This review focuses on cinchona/oak bark substitute, Soymida febrifuga (Meliaceae), also known as Indian red wood whose medical virtues are used for treating various health ailments in folk and traditional medicine. Here we review the ex situ conservation strategies for the mass multiplication of this endemic plant by micropropagation due to its difficulty in rooting of stem cuttings and high seedling mortality rates. Further we also discuss phytochemical investigations from plant parts led to the isolation of tetranortriterpenoids, flavonoids, limonoids, alkaloids, tannins and glycosides. These chemical constituents were reported to be cytotoxic and strikingly are antimalarial, anti-inflammatory, antioxidant, antidiabetic, antiallergic, antifungal, antiulcer, spasmolytic, insect antifeedant, antihelminthic, antibacterial and anticancer in action. Overall this review highlights and compiles the available information of Soymida febrifuga propagation, medicinal importance of secondary...

Physiology and Molecular Biology of Plants, 2015

Bixa orellana L. (Bixaceae) is a multipurpose tree grown for the production of commercially impor... more Bixa orellana L. (Bixaceae) is a multipurpose tree grown for the production of commercially important dyes. In the present study, an efficient, reproducible protocol was developed for direct plant regeneration from in vitro derived petiole explants of Bixa orellana L. Murashige and Skoog medium (MS) supplemented with 2-isopentenyl adenine (9.8 μM) and naphthalene acetic acid (10.7 μM) was found to be optimum for production of high frequency of shoot organogenesis. Subculturing of the shoots onto the fresh MS medium containing similar concentrations of 2-iP (9.8 μM) and NAA (10.7 μM) produced elongated shoots. Elongated shoots when placed onto MS medium supplemented with 1.7 μM indole-3-acetic acid and 14.7 μM 2-iP produced optimal rooting. Rooted plantlets were acclimatized and transplanted to the field successfully. Histological investigation revealed the origin of shoot primordia, from subepidermal cells of petiole explants. The regeneration protocol developed in this study can be useful for mass in vitro propagation and effective genetic transformation of commercially important edible dye yielding tree species.

Annual Research & Review in Biology, 2014

Asian Pacific Journal of Tropical Biomedicine, 2014

Cold Spring Harbor perspectives in biology, 2013

Nonhomologous end joining (NHEJ) refers to a set of genome maintenance pathways in which two DNA ... more Nonhomologous end joining (NHEJ) refers to a set of genome maintenance pathways in which two DNA double-strand break (DSB) ends are (re)joined by apposition, processing, and ligation without the use of extended homology to guide repair. Canonical NHEJ (c-NHEJ) is a well-defined pathway with clear roles in protecting the integrity of chromosomes when DSBs arise. Recent advances have revealed much about the identity, structure, and function of c-NHEJ proteins, but many questions exist regarding their concerted action in the context of chromatin. Alternative NHEJ (alt-NHEJ) refers to more recently described mechanism(s) that repair DSBs in less-efficient backup reactions. There is great interest in defining alt-NHEJ more precisely, including its regulation relative to c-NHEJ, in light of evidence that alt-NHEJ can execute chromosome rearrangements. Progress toward these goals is reviewed.

Proceedings of the National Academy of Sciences of the United States of America, Jan 28, 2014

Transposable elements (TEs) have had a major influence on shaping both prokaryotic and eukaryotic... more Transposable elements (TEs) have had a major influence on shaping both prokaryotic and eukaryotic genomes, largely through stochastic events following random or near-random insertions. In the mammalian immune system, the recombination activation genes1/2 (Rag1/2) recombinase has evolved from a transposase gene, demonstrating that TEs can be domesticated by the host. In this study, we uncovered a domesticated transposase, Kluyveromyces lactis hobo/Activator/Tam3 (hAT) transposase 1 (Kat1), operating at the fossil imprints of an ancient transposon, that catalyzes the differentiation of cell type. Kat1 induces mating-type switching from mating type a (MATa) to MATα in the yeast K. lactis. Kat1 activates switching by introducing two hairpin-capped DNA double-strand breaks (DSBs) in the MATa1-MATa2 intergenic region, as we demonstrate both in vivo and in vitro. The DSBs stimulate homologous recombination with the cryptic hidden MAT left alpha (HMLα) locus resulting in a switch of the cel...

DNA Repair, 2014

LIG4/Dnl4 is the DNA ligase that (re)joins DNA double-strand breaks (DSBs) via nonhomologous end ... more LIG4/Dnl4 is the DNA ligase that (re)joins DNA double-strand breaks (DSBs) via nonhomologous end joining (NHEJ), an activity supported by binding of its tandem BRCT domains to the ligase accessory protein XRCC4/Lif1. We screened a panel of 88 distinct ligase mutants to explore the structure-function relationships of the yeast Dnl4 BRCT domains and inter-BRCT linker in NHEJ. Screen results suggested two distinct classes of BRCT mutations with differential effects on Lif1 interaction as compared to NHEJ completion. Validated constructs confirmed that D800K and GG(868:869)AA mutations, which target the Lif1 binding interface, showed a severely defective Dnl4-Lif1 interaction but a less consistent and often small decrease in NHEJ activity in some assays, as well as nearly normal levels of Dnl4 accumulation at DSBs. In contrast, mutants K742A and KTT(742:744)ATA, which target the β3-α2 region of the first BRCT domain, substantially decreased NHEJ function commensurate with a large defect in Dnl4 recruitment to DSBs, despite a comparatively greater preservation of the Lif1 interaction. Together, these separation-of-function mutants indicate that Dnl4 BRCT1 supports DSB recruitment and NHEJ in a manner distinct from Lif1 binding and reveal a complexity of Dnl4 BRCT domain functions in support of stable DSB association.

Notulae Scientia Biologicae, 2014

Scientific Reports, 2014

Intake of fruits rich in antioxidants in daily diet is suggested to be cancer preventive. Sapota ... more Intake of fruits rich in antioxidants in daily diet is suggested to be cancer preventive. Sapota is a tropical fruit grown and consumed extensively in several countries including India and Mexico. Here we show that methanolic extracts of Sapota fruit (MESF) induces cytotoxicity in a dose-dependent manner in cancer cell lines. Cell cycle analysis suggested activation of apoptosis, without arresting cell cycle progression. Annexin V-propidium iodide double-staining demonstrated that Sapota fruit extracts potentiate apoptosis rather than necrosis in cancer cells. Loss of mitochondrial membrane potential, upregulation of proapoptotic proteins, activation of MCL-1, PARP-1, and Caspase 9 suggest that MESF treatment leads to activation of mitochondrial pathway of apoptosis. More importantly, we show that MESF treatment leads to significant inhibition of tumor growth and a 3-fold increase in the life span of tumor bearing animals compared to untreated tumor mice.

Journal of King Saud University - Science, 2015

Fruiting bodies of three wild polypores namely Ganoderma applanatum, Ganoderma resinaceum and Gan... more Fruiting bodies of three wild polypores namely Ganoderma applanatum, Ganoderma resinaceum and Ganoderma curtisi, collected from Ondo State Southwest Nigeria, were tested for their antimicrobial activity using the agar well diffusion method. Ethanol, methanol, methanol: ethanol (50:50), water, ethyl acetate and n-hexane were used as the extraction solvent.The partially purified extract of the mushrooms was tested for antimicrobial action against twelve microorganisms in the genera Pseudomonas aeruginosa NCIB 950, Serratia marcescens NCIB 1377, Micrococcus luteus NCIB 196, Bacillus cereus NCIB 6345, Bacillus substilis NCIB 3610, Klebsiella ozaenae, Escherichia coli, Staphylococcus aureus, Streptococcus pneumonia, Candida albicans Aspergillus fumigatus and Saccharomyces cerevisiae. The percentage yield for the various solvent was found to range between 8.10 and 0.90 with the mixture of methanol ethanol as solvent having the highest value while the least was ethyl acetate The methanol, ethanol and ethyl acetate extract of the Ganoderma has antimicrobial activity against the test organism with the growth inhibition as the index parameter. The susceptibility of the organism to the antimicrobial activity of the extract vary among the different organism tested. The highest zone of inhibition was recorded in Bacillus subtilis (28.3) while the least was observed in Aspergillus fumigatus. The zone of inhibition was found to be highest in bacteria than fungi. Comparison of the antimicrobial activity of the extract with some selected commercial antibiotics showed that the latter was found to be more effective in inhibiting the growth of the tested pathogen than the former. Phytochemical screeninig of the extract reveal the presence of saponins while alkaloids were absent. The presence of these bioactive molecules justifies the use of the Ganoderma species in ethnomedicine.

PLoS Genetics, 2013

DNA ligase IV (Dnl4 in budding yeast) is a specialized ligase used in non-homologous end joining ... more DNA ligase IV (Dnl4 in budding yeast) is a specialized ligase used in non-homologous end joining (NHEJ) of DNA doublestrand breaks (DSBs). Although point and truncation mutations arise in the human ligase IV syndrome, the roles of Dnl4 in DSB repair have mainly been examined using gene deletions. Here, Dnl4 catalytic point mutants were generated that were severely defective in auto-adenylation in vitro and NHEJ activity in vivo, despite being hyper-recruited to DSBs and supporting wild-type levels of Lif1 interaction and assembly of a Ku-and Lif1-containing complex at DSBs. Interestingly, residual levels of especially imprecise NHEJ were markedly higher in a deletion-based assay with Dnl4 catalytic mutants than with a gene deletion strain, suggesting a role of DSB-bound Dnl4 in supporting a mode of NHEJ catalyzed by a different ligase. Similarly, next generation sequencing of repair joints in a distinct single-DSB assay showed that dnl4-K466A mutation conferred a significantly different imprecise joining profile than wild-type Dnl4 and that such repair was rarely observed in the absence of Dnl4. Enrichment of DNA ligase I (Cdc9 in yeast) at DSBs was observed in wild-type as well as dnl4 point mutant strains, with both Dnl4 and Cdc9 disappearing from DSBs upon 59 resection that was unimpeded by the presence of catalytically inactive Dnl4. These findings indicate that Dnl4 can promote mutagenic end joining independently of its catalytic activity, likely by a mechanism that involves Cdc9.

Plant Tissue Culture and Biotechnology, 2012

Soymida febrifuga (Roxb.) A. Juss., (Meliaceae) an indigenous lofty deciduous tree, endemic to In... more Soymida febrifuga (Roxb.) A. Juss., (Meliaceae) an indigenous lofty deciduous tree, endemic to India, has extensive pharmacological activity. Continuous destruction of plants due to environmental and geopolitical instability has posed a major threat to endemic tree species. In vitro propagation techniques are also problematic due to high incidence of shoot tip necrosis (STN). The effect of various culture conditions on regeneration and STN of Soymida febrifuga were investigated. Nodal segments from field grown plants were used as explants and cultured on MS supplemented with different concentrations of BA, KN, NAA and IAA. The synergistic effect of BA (2 mg/l) and NAA (0.2 mg/l) induced a mean of 6.3 ± 0.09 shoots from the nodal explants with a frequency of 80.4%. Frequent subculturing of nodal explants from in vitro derived shoots increases the number of multiple shoots, but the regenerated shoots exhibited the symptoms of STN. Various factors such as strength of the media, differe...