Browsing by Author "Indra Singh"

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Alterations in antioxidative machinery and growth parameters upon application of nitric oxide donor that reduces detrimental effects of cadmium in rice seedlings with increasing days of growth
(Elsevier B.V., 2020) Prerna Singh; Indra Singh; Kavita Shah
Finding its way into the agricultural lands through various sources Cadmium (Cd) has been proved to be deleterious in plant systems. Cd-induced oxidative stress tampers with both plant's physiology and biochemistry. Nitric oxide (NO) is a key signaling molecule in plant stress response and plays a significant role in many regulatory pathways of plant development. This study investigates the effect on rice seedlings grown under 50 µM Cd/SNP alone or in combination in the growth medium, on roots and shoots of rice cv. HUR 3022 at 1, 4, and 7 days of growth. Plants treated with Cd alone showed diminished growth, decreased length and weight, lower cell viability and less chlorophyll. Application of equimolar quantity of SNP to cadmium-stressed rice within the growth medium repressed Cd-uptake and reversed the Cd-induced detrimental effects. Rice plants treated with only Cd showed high oxidative stress. Specific activity of antioxidant enzymes such as guaiacol peroxidase, ascorbate peroxidase and glutathione reductase were increased. Antioxidants like glutathione and ascorbate had lower levels in cadmium treated plants. A high amount of reactive oxygen species were generated in cadmium treated plants which were supported by the histochemical studies for - hydrogen peroxide and superoxide anion. Application of SNP in rice plants under cadmium-stress in the growth medium mitigated the Cd-induced oxidative stress by elevating the levels of antioxidants and reducing the amount of reactive oxygen species produced. These results were supported by reduced specific activity of oxidative enzymes. The results indicate that NO when supplemented exogenously, diminishes the deleterious effects of Cd in rice plants. Our study presents report from the early days of plant development i.e., 1,4,7 days of growth because physiological and biochemical changes occurring during these early developmental days may provide significant insights into rice plant's adaptation strategy towards any perceived stress. © 2020 SAAB
Designing Self-Inhibitory fusion peptide analogous to viral spike protein against novel severe acute respiratory syndrome (SARS-CoV-2)
(Taylor and Francis Ltd., 2022) Indra Singh; Shalini Singh; Krishna Kumar Ojha; Neetu Singh Yadav
COVID-19 is a highly contagious viral infection caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which is declared pandemic by the World Health Organization (WHO). The spike protein of SARS-CoV-2 is a key component playing a pivotal role in facilitating viral fusion as well as release of genome into the host cell. Till date there is no clinically approved vaccine or drug available against Covid-19. We designed four hydrophobic inhibitory peptides (ITPs) based on WWIHS (Wimley and White interfacial hydrophobicity scale) score, targeting the HR1 domain of spike protein. Two inhibitory peptides out of four have a strong affinity to the hydrophobic surface of HR1 domain in pre-fusion spike protein. The MD simulation result showed the strong accommodation of ITPs with HR1 domain surface. These self-inhibitory peptides mimic the function of HR2 by binding to HR1 domain, thus inhibiting the formation of HR1-HR2 post-fusion complex, which is a key structure for virus-host tropism. Communicated by Ramaswamy H. Sarma. © 2021 Informa UK Limited, trading as Taylor & Francis Group.
Development of B cell epitopes-based enzyme linked immune sorbent assay for detection of bovine anti-Mullerian hormone
(Springer Science and Business Media Deutschland GmbH, 2023) Prasanna Pal; Anjali Aggarwal; Y.S. Rajput; Rajib Deb; Vinay G. Joshi; Arvind Kumar Verma; Avijit Haldar; Indra Singh; Sonika Grewal; Sachinandan De
The present study aimed to generate antibodies against predicted B cell epitopic peptides encoding bAMH for developing different ELISA models. Sandwich ELISA was determined to be an excellent technique for assessing bAMH in bovine plasma based on sensitivity tests. The assay's specificity, sensitivity, inter- and intra-assay CV, recovery %, Lower limit of quantification (LLOQ), and Upper limit of quantification (ULOQ) were determined. The test was selective since it did not bind to AMH-related growth and differentiation factors (LH and FSH) or non-related components (BSA, progesterone). The intra-assay CV was 5.67%, 3.12%, 4.94%, 3.61% and 4.27% for 72.44, 183.11, 368.24, 522.24 and 732.25 pg/ml AMH levels, respectively. At the same time, the inter-assay CV was 8.77%, 7.87%, 4.53%, 5.76% and 6.70% for 79.30, 161.27, 356.30, 569.33 and 798.19 pg/ml AMH levels, respectively. The average (Mean ± SEM) recovery percentages were 88–100%. LLOQ was 5 pg/ml and ULOQ at 50 µg/ml (CV < 20%). In conclusion, we developed a new highly sensitive ELISA against bAMH using epitope specific antibodies. © 2023, King Abdulaziz City for Science and Technology.
Development of CD163 receptor-based enzyme-linked immunosorbent assay for diagnosis of porcine reproductive and respiratory syndrome virus
(Springer Science and Business Media Deutschland GmbH, 2022) Rajib Deb; Ajay Kumar Yadav; Gyanendra Singh Sengar; Joyshikh Sonowal; D. Lalita; Seema Rani Pegu; Indra Singh; Ningthoukhongjam Linda; Pranab Jyoti Das; Satish Kumar; Prasanna Pal; Souvik Paul; Swaraj Rajkhowa; Vivek Kumar Gupta
Porcine reproductive and respiratory syndrome (PRRS) is an important economical disease in the global swine industry. The accurate detection of the PRRS virus (PRRSV) antigen is essential for the disease control and prevention programme. In this study, an indirect enzyme-linked immunosorbent test (PRRSVCD163-iELISA) was developed for the detection of the PRRSV antigen in samples of post-mortem swine tissue using the recombinant pig CD163 receptor protein as the capture ligand. The test was found to be specific for PRRSV, with no cross-reactions with other prevalent pig viral pathogens. The assay was validated by testing 217 post-mortem porcine tissue samples and the results were found to be satisfactory with a relative accuracy of 88.88%. Our assay is also quite precise, with intra- and inter-assay CVs of 6% and 10%, respectively. These findings imply that the PRRSVCD163-iELISA developed is capable of detecting the PRRSV antigen in swine post-mortem tissue samples. This research showed that porcine CD163, the PRRSV cellular receptor, can be exploited to build a diagnostic technique for the detection of PRRSV antigen. © 2022, King Abdulaziz City for Science and Technology.
Evidences for structural basis of altered ascorbate peroxidase activity in cadmium-stressed rice plants exposed to jasmonate
(Kluwer Academic Publishers, 2014) Indra Singh; Kavita Shah
Binding interactions of cadmium (Cd) with rice ascorbate peroxidase (OsAPX) in presence or absence of jasmonate was examined in-silico. OsAPX is a 250 amino acid long protein with 90 % sequence similarity to soybean-APX. The 3D model of OsAPX obtained by homology modeling using soybean APX (PDBID:1OAF) as template was associated with -15975.85 kJ/mol energy, 100 % residues in favoured region, verify score of 0.85, ERRAT score 89.625 and a negative ProSA graph, suggesting OsAPX model to be of good quality, robust and reliable which was submitted with Protein Model Database with PMDBID: PM0078091. The rice ascorbate peroxidase ascorbate [OsAPX-Asc] complex had a substrate binding cavity involving residues at position 30KSCAPL35, 167RCH169 and 172R wherein ascorbate accommodated via three H-bonds involving 30Lys at the γ-edge of heme. 169His served as a bridge between heme-porphyrin of OsAPX and ascorbate creating a charge relay system. Cd bound in [OsAPX-Asc-Cd] complex at 29EKSCAPL35, a site similar to ascorbate binding site. The binding of Cd caused breaking of 169His bridge shifting the protein conformation. Cadmium exhibited four electrostatic interactions via 29Glu of OsAPX backbone. Docking of [OsAPX-Asc] with jasmonic acid (JA) resulted in [OsAPX-Asc-JA] complex where 4-H-bonds held JA to OsAPX in a cavity at γ-edge on the distal side of heme. The binding of [OsAPX-Asc-JA] to Cd show the metal to bind at a position other than that involved in binding of OsAPX with Cd alone. Results indicate that Cd does not replace iron or ascorbate or JA but binds to OsAPX on the surface at a separate site electrostatically. In presence of JA the interactions involved in formation of [OsAPXAsc] are restored which is otherwise altered by the presence of Cd. The formation and reformation of H-bond take place between the [OsAPX-Asc] and Cd/JA. It is the interaction between heme and ascorbate which is modulated differently in presence of Cd/JA. In absence of JA, Cd-binds to the [OsAPX-Asc] complex at the proximal end of APX near Asc-binding site, whereas in presence of JA, Cd-binds on the opposite site of the Asc-binding site involving 30Lys and 29Glu residues. In-silico binding studies well correlate with the wet-lab results where exogenous application of JA increased the activity of OsAPX in rice grown under Cd-stress. Therefore it is concluded that the activity of OsAPX in rice roots and shoots are compromised under Cd-stress alone. © 2014 Springer Science+Business Media.
Evidences for suppression of cadmium induced oxidative stress in presence of sulphosalicylic acid in rice seedlings
(Kluwer Academic Publishers, 2015) Indra Singh; Kavita Shah
Abstract: Role of salicylic acid in plants grown under metal contaminated soil is not yet understood. To investigate if signaling by Cd-induced H2O2 or SA, collide or integrate, rice cv. HUR3022, grown for 3, 7 and 10 day under 50 µM Cd/SA alone or in combination were studied. Dynamic distribution of Cd2+ and SA, cell viability, time-dependent changes in H2O2 levels, free proline, chlorophyll, ascorbate and glutathione pools and altered antioxidant enzymes in roots/shoots of rice were observed. Exogenous SA decelerated cell-death in Cd-stressed rice playing a positive role in protection against injury. Cd led to elevated activities of superoxide dismutase, ascorbate peroxidase, guaiacol peroxidase and phenylalanine ammonia lyase and lowered catalase and glutathione reductase activities in roots and shoots of rice seedlings. By contrast, SA + Cd2+ reversed the trend of enzyme activities, levels of non-enzymatic antioxidants and GSH in roots/shoots suggesting alleviation of Cd-induced oxidative damage in rice. Suppression of Cd-uptake in presence of SA suggest an epistatic effect of SA-signal over Cd-induced ROS-signal in rice roots. No additive effects of SA or H2O2 signals were noted in Cd-stressed rice. Results indicate integration of different signals and SA-enhanced Cd2+ tolerance due to SA-regulated Cd2+ uptake as well as the SA-elevated enzymatic and non-enzymatic antioxidant pool in rice.; Graphical abstract: Role of salicylic acid in rice plants grown under Cd-contaminated soil was studied. Cd-induced H2O2 or SA, signals both collide and/or integrate. Dynamic distribution of Cd2+ and SA, cell viability, time-dependent changes in H2O2 levels, free proline, chlorophyll, ascorbate and glutathione pools appear. Suppression of Cd-uptake in presence of SA were noted. Integration of different signals, SA-induced Cd2+ tolerance and SA-elevated enzymatic and non-enzymatic antioxidant pool in rice was observed.[Figure not available: see fulltext.] © 2015, Springer Science+Business Media Dordrecht.
Exogenous application of methyl jasmonate lowers the effect of cadmium-induced oxidative injury in rice seedlings
(Elsevier Ltd, 2014) Indra Singh; Kavita Shah
Rice seedlings grown under 50 μM cadmium alone or in combination with 5 μM methyl jasmonate were investigated for Cd-induced oxidative injury at 3, 7 and 10 days of treatment. MeJA treatments alone did not have any significant change in antioxidant enzyme activities or levels of H2O2 and O2- in roots/shoots, as compared to controls during 3-10 days. The Cd-stressed plants When supplemented with exogenous MeJA revealed significant and consistent changes in activities of antioxidant enzymes CAT, SOD, POD and GR paralleled with an increased GSH-pools than that in plants subjected to Cd-stress alone. Synthesis of GSH driven by increasing demand for GSH in response to Cd-induced oxidative stress in rice was evident. Increased activity of LOX under Cd-stress was noted. Results suggest enhanced Cd-tolerance, lowered Cd2+ uptake, an improved membrane integrity and 'switching on' of the JA-biosynthesis by LOX in the Cd-stressed rice roots/shoots exposed to MeJA. Exposure to MeJA improved antioxidant response and accumulation of antioxidants which perhaps lowered the Cd-induced oxidative stress in rice. It is this switching on/off of the JA-biosynthesis and ROS mediated signal transduction pathway involving glutathione homeostasis via GR which helps MeJA to mitigate Cd-induced oxidative injury in rice. © 2014 Elsevier Ltd. All rights reserved.
Expression of ZAT12 transcripts in transgenic tomato under various abiotic stresses and modeling of ZAT12 protein in silico
(Springer Science and Business Media B.V., 2014) Avinash Chandra Rai; Indra Singh; Major Singh; Kavita Shah
ZAT12 a C2H2-zinc-finger protein is an abiotic stress-responsive transcription factor in plants having less information about their structure. Transcription analysis proved that ZAT12 transcripts over expressed during drought, heat and salt stress conditions which led to an interest in 3-D structural studies of ZAT12 in Brassica carinata. Over-expression of BcZAT12 in transformed tomato plants under abiotic stresses, suggest role of ZAT12 in conferring stress-tolerance in tomato. Sequence analysis of ZAT12 protein (Accession No. ABB55254.1) from B. carinata revealed it as a 161 amino acid long protein with short conserved motif 140LDLXL144 in C-terminal, a leucine rich L-Box with—14EXXAXCLXXL23 motif in N-terminal region and presence of two conserved Zinc-Finger motifs “CXXCXXXXXXXQALGGHXXXH” between positions 42–62 and 85–105. The two zinc finger motifs have presence of two conserved glutamic acid (Glu) and phenylalanine (Phe) residues. Two methionine (Met) residues at position 94 and 102 present in ZF-motif-2 were absent in ZF-motif-1. The 94Met and 97Ala in ZFmotif-2 were found to be replaced by serine (Ser) in ZFmotif-1. Homology and ab initio structural modeling of ZAT12 encodedBcZAT12 protein of B. carinata resulted in robust 3-D models and were evaluated for structural motifs, associated GO terms and protein-DNA interactions. The BcZAT12 protein model, was of good quality, reliable, stable and is deposited in PMDB database (PMDB ID: PM0078213). BcZAT12 is annotated as an intracellular protein having molecular function in Zn-binding which in turn regulates signal transduction/translation processes in response to abiotic stresses in plants. Results suggest BcZAT12 protein to interact directly with one strand of dsDNA via electrostatic and H-bonds. © Springer Science+Business Media New York 2014.
In silico study of interaction between rice proteins enhanced disease susceptibility 1 and phytoalexin deficient 4, the regulators of salicylic acid signalling pathway
(2012) Indra Singh; Kavita Shah
Enhanced disease susceptibility 1 (EDS1), a plant-specific protein has homology with the eukaryotic lipase in their N-terminal halves and a unique domain at its C-termini. EDS1 is known to be an important regulator of biotic stress and an essential component of basal immunity. EDS1 interacts with its positive co-regulator phytoalexin deficient 4 (PAD4), resulting in mobilization of the salicylic acid defence pathway. Limited information regarding this interaction in rice is available. To study this interaction, a model of EDS1 and PAD4 proteins from rice was generated and validated with Accelrys DS software version 3.1 using bioinformatics interface. The in silico docking between the two proteins showed a significant protein-protein interaction between rice EDS1 and PAD4, suggesting that they form a dimeric protein complex, which, similar to that in Arabidopsis, is perhaps also important for triggering the salicylic acid signalling pathway in plants. © Indian Academy of Sciences.
Investigating the RelA Gene's role in African Swine fever tolerance: a study of Indian pigs
(Academic Press, 2025) Arutkumaran S; Rajib Deb; Shanmathi S; Soumendu Chakravarti; Pranab Jyoti Das; Gyanendra Singh Sengar; Seema Rani Pegu; Indra Singh; Satish Kumar; Meera K; Swaraj Rajkhowa; Pushpendra Pavan Kumar; Vivek Kumar Gupta
African Swine Fever (ASF) is a highly contagious transboundary viral disease affecting domestic pigs worldwide, often resulting in nearly 100 % mortality due to the lack of effective vaccines. However, wild species such as warthogs (Phacochoerus sp.) and bush pigs (Potamochoerus sp.) do not exhibit clinical symptoms of the disease, previous studies showed that amino acid substitutions in a proto-oncogene, RelA, a subunit of NF-κB found in warthogs might be responsible for their resistance. Expanding this study over an Indian breed named Doom which was considered tolerant due to lack of much information regarding their ASFV-positive cases and identifying the genetic basis of tolerance might help in creating a tolerant breed, thereby controlling the spread of the disease. So, this study was initiated to investigate the polymorphic signature in RelA gene of Doom breed similar to warthogs. Initially molecular docking studies identified a potential interaction between the N-terminal sub-domain of Rel homology domain of porcine RELA and the African Swine Fever Viral (ASFV) protein A238L, a hypothetical viral protein of unknown function. This amino acid interaction corresponds to a nucleotide sequence of exon 10 of porcine RelA gene. Further, ARMS-PCR (Amplification Refractory Mutation System-PCR) and PCR-RFLP (Restriction Fragment Length Polymorphism) were optimized to target a single nucleotide polymorphism (SNP, CCT/GCT: Pro→Ala) identified in the exon 10 region through Ensemble genome browser. PCR results showed the presence of homozygous CC genotype, coding for proline, in all pig breeds tested, including Doom. Sanger sequencing confirmed the CC genotype across all breeds, indicating that amino acid substitutions in this RELA domain were not present in Doom and therefore may not be responsible for ASF tolerance. Since disease tolerance is a polygenic trait relying on one gene might not reveal the exact cause for their tolerance against ASFV. Future research can be targeted on other set of genes playing important role in innate immune pathways or Genome Wide Association Study can be preferred to identify and associate SNPs with Doom's tolerance against ASFV. © 2025 Elsevier Ltd
Porcine Circovirus type 2 infected myocardial tissue transcriptome signature
(Elsevier B.V., 2022) Rajib Deb; Joyshikh Sonowal; Gyanendra Singh Sengar; Seema Rani Pegu; Manas Ranjan Praharaj; Waseem Akram Malla; Indra Singh; Ajay Kumar Yadav; Swaraj Rajkhowa; Pranab Jyoti Das; Jaya Bharati; Souvik Paul; Vivek Kumar Gupta
The goal of this study was to compare the global gene expression profile in cardiac tissues of pig infected with porcine circovirus 2 (PCV2) to that of healthy cells. Since PCV2 infection causes severe cardiovascular lesions, the myocardial tissue model was chosen for this study. In High-throughput transcriptome analysis, DESeq2 and CLC genomics workbench analyses revealed a total of 196 significantly differentially expressed genes (DEGs) (p-value < 0.05). Furthermore, 194 transcripts were upregulated, while only two were downregulated (HSPA6 and DNAJA1), with fold changes ranging from 16.293 to −10.002. Among the KEGG canonical pathways targeted by the DEGs in the functional analysis, adrenergic signalling in cardiomyocytes, Cardiac Muscle Contraction, Hypertrophic Cardiomyopathy (HCM), and Dilated Cardiomyopathy (DCM) tends to be enriched. The differentially expressed highly connected (DEHC) biomarker genes in pathogenicity of PCV2 infection, such as LDB3, MYOZ2, CASQ2, TNNT2, MLC2V, MYBPC3, ACTC1, TCAP, TNNI3, TRDN, CSRP3, MYL3, RYR2, LMOD2, MYH7, etc., were identified using protein–protein interaction (PPI) network analysis. The study might provide detailed information on the dysregulated genes and biological pathways in infected myocardial tissues that may be essential for PCV2-related heart pathology. © 2022 Elsevier B.V.
Reduced activity of nitrate reductase under heavy metal cadmium stress in rice: an in silico answer
(Frontiers Media S.A., 2019) Prerna Singh; Indra Singh; Kavita Shah
Cadmium is a well known toxic heavy metal, which has various detrimental effects on plant system. In plants an important enzyme involved in the production of nitric oxide, nitrate reductase, is also affected by cadmium toxicity. According to many studies cadmium has an inhibitory effect on nitrate reductase activity. Similar effect of cadmium was found in our study where an inhibitory effect of cadmium on nitrate reductase activity was noted. However, the mechanism behind this inhibition has not been explored. With the help of homology, 3-D structure of rice-nitrate reductase is modeled in this study. Its binding with nitrate, nitrite and cadmium metal in silico has been explored. The bonds formed between the enzyme-substrate complex, enzyme-cadmium and differences in interactions in presence of cadmium has been studied in detail. The present study should help in understanding the modeled structure of rice-nitrate reductase in 3-D which may in turn guide enzyme related studies in silico. The present study also provides an insight as to how cadmium interacts with nitrate reductase to alter the enzyme activity. © 2019 Singh, Singh and Shah.
Structural and functional analysis of protein
(Elsevier, 2021) Neetu Singh Yadav; Pawan Kumar; Indra Singh
This chapter deals with the topic of bioinformatics approaches applied to biology, essentially for the analysis and characterization of protein. Protein is a large and diverse class of biomolecules, involved in various functional and structural parts in all forms of life. Advances in the experimental techniques and computational methodology have facilitated a relevant understanding of the 3D arrangement of the protein structures and the molecular level insight of the protein function. The collective exponential growth of the solved 3D structures has been utilized extensively to develop an advanced computational program to classify the structure information in different hierarchical levels, to develop more accurate machine learning-based protocols for 3D-structure prediction and scoring function, and to understand the mysterious protein folding problem. This chapter discusses the state-of-the-art computational techniques oriented toward the protein structure and function prediction from the sequence. These methods, if used appropriately, can provide valuable indicators of protein structure and prediction. © 2022 Elsevier Inc. All rights reserved.
Transcriptome signatures of host tissue infected with African swine fever virus reveal differential expression of associated oncogenes
(Springer, 2024) Rajib Deb; Gyanendra Singh Sengar; Joyshikh Sonowal; Seema Rani Pegu; Pranab Jyoti Das; Indra Singh; Soumendu Chakravarti; Arutkumaran Selvaradjou; Nitin Attupurum; Swaraj Rajkhowa; Vivek Kumar Gupta
African swine fever (ASF) has emerged as a threat to swine production worldwide. Evasion of host immunity by ASF virus (ASFV) is well understood. However, the role of ASFV in triggering oncogenesis is still unclear. In the present study, ASFV-infected kidney tissue samples were subjected to Illumina-based transcriptome analysis. A total of 2463 upregulated and 825 downregulated genes were differentially expressed (p < 0.05). A literature review revealed that the majority of the differentially expressed host genes were key molecules in signaling pathways involved in oncogenesis. Bioinformatic analysis indicated the activation of certain oncogenic KEGG pathways, including basal cell carcinoma, breast cancer, transcriptional deregulation in cancer, and hepatocellular carcinoma. Analysis of host-virus interactions revealed that the upregulated oncogenic RELA (p65 transcription factor) protein of Sus scrofa can interact with the A238L (hypothetical protein of unknown function) of ASFV. Differential expression of oncogenes was confirmed by qRT-PCR, using the H3 histone family 3A gene (H3F3A) as an internal control to confirm the RNA-Seq data. The levels of gene expression indicated by qRT-PCR matched closely to those determined through RNA-Seq. These findings open up new possibilities for investigation of the mechanisms underlying ASFV infection and offer insights into the dynamic interaction between viral infection and oncogenic processes. However, as these investigations were conducted on pigs that died from natural ASFV infection, the role of ASFV in oncogenesis still needs to be investigated in controlled experimental studies. © The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature 2024.