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Dhrubajyoti Chattopadhyay - One of the best experts on this subject based on the ideXlab platform.
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Investigating the role of Ebp1 in Chandipura Virus infection
Journal of Biosciences, 2019Co-Authors: Dhritiman Dey, Ayae Honda, Dhrubajyoti ChattopadhyayAbstract:ErbB-3 binding protein 1 (Ebp1) is a host protein which binds ErbB-3 receptor to induce signalling events for cell growth regulation. In addition, Ebp1 also interacts with ribonucleoprotein complexes. In recent times, Ebp1 was found to play an antagonistic role in viral infections caused by Influenza and Rinderpest Viruses. In our present work we have tried to understand the role of Ebp1 in Chandipura Virus (CHPV) infection. We have observed an induction in Ebp1 expression upon CHPV infection similar to other Viruses. However, unlike other Viruses an overexpressed Ebp1 only reduces viral protein expression, but does not affect its progeny formation. Additionally, this effect is being carried out in an indirect manner, as there is no interaction between Ebp1 and viral proteins. This is despite Ebp1’s presence in viral inclusion bodies.
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Role of tryptophan 135 of Chandipura Virus phosphoprotein P in dimerization and complex formation with leader RNA: structural aspect using time resolved anisotropy and simulation
RSC Advances, 2015Co-Authors: Manini Mukherjee, Dhrubajyoti Chattopadhyay, Aditya Kumar Sarkar, Arunava Roy, Pinki Saha Sardar, Ansuman Lahiri, Sanjib GhoshAbstract:The aggregation of phosphoprotein P of Chandipura Virus (CHPV) and its interaction with viral leader (le) RNA in aqueous buffer and 40% ethylene glycol (EG)-buffer have been characterised using two single tryptophan (Trp/W) mutants W105F and W135F. The longer rotational correlation time [(θC)T] originating from overall motion observed at 300 nM concentration conforms to a globular structure of the monomer of WT and both the mutants. The (θC)T values also indicate that W135F does not form a dimer at 1500 nM concentration; while a dimer with disordered structure is predicted for both WT and W105F. The complexes of WT and W105F with le RNA at monomeric and dimeric conditions are indicated to have tight core packing. Dimerization and complex formation at both the concentrations enhance the correlation time arising from the localised motion of Trp side chain [(θC)S] for WT and W105F predicting hindered localized rotation of Trp 135. Comparative protein modelling and molecular dynamics (MD) simulations using the amino acids domain ranging from 105–168 of the full length CHPVP based on the vesicular stomatitis Virus phosphoprotein (VSVP) also indicate that formation of dimers are more feasible for WT and W105F compared to W135F.
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A peptide targeted against phosphoprotein and leader RNA interaction inhibits growth of Chandipura Virus -- an emerging rhabdoVirus.
Antiviral research, 2013Co-Authors: Arunava Roy, Dhrubajyoti Chattopadhyay, Smarajit Polley, Prasenjit Chakraborty, Sabita RoyAbstract:Abstract The fatal illness caused by Chandipura Virus (CHPV), an emerging pathogen, presently lacks any therapeutic option. Previous research suggested that interaction between the virally encoded phosphoprotein (P) and the positive sense leader RNA (le-RNA) may play an important role in the viral lifecycle. In this report, we have identified a β -sheet/loop motif in the C-terminal domain of the CHPV P protein as essential for this interaction. A synthetic peptide encompassing this motif and spanning a continuous stretch of 36 amino acids (Pep 208–243 ) was found to bind the le-RNA in vitro and inhibit CHPV growth in infected cells. Furthermore, a stretch of three amino acid residues at position 217–219 was identified as essential for this interaction, both in vitro and in infected cells. siRNA knockdown-rescue experiments demonstrated that these three amino acid residues are crucial for the leader RNA binding function of P protein in the CHPV life cycle. Mutations of these three amino acid residues render the peptide completely ineffective against CHPV. Effect of inhibition of phosphoprotein–leader RNA interaction on viral replication was assayed. Peptide Pep 208–243 tagged with a cell penetrating peptide was found to inhibit CHPV replication as ascertained by real time RT-PCR. The specific inhibition of viral growth observed using this peptide suggests a new possibility for designing of anti-viral agents against Mononegavirale group of human Viruses.
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Interaction of Chandipura Virus N and P Proteins: Identification of Two Mutually Exclusive Domains of N Involved in Interaction with P
2013Co-Authors: Arindam Mondal ¤a, Arunava Roy, Ipto Sarkar, Jishnu Mukherjee ¤b, Tridib Ganguly ¤c, Dhrubajyoti ChattopadhyayAbstract:The nucleocapsid protein (N) and the phosphoprotein (P) of nonsegmented negative-strand (NNS) RNA Viruses interact with each other to accomplish two crucial events necessary for the viral replication cycle. First, the P protein binds to the aggregation prone nascent N molecules maintaining them in a soluble monomeric (N 0) form (N 0-P complex). It is this form that is competent for specific encapsidation of the viral genome. Second, the P protein binds to oligomeric N in the nucleoprotein complex (N-RNA-P complex), and thereby facilitates the recruitment of the viral polymerase (L) onto its template. All previous attempts to study these complexes relied on co-expression of the two proteins in diverse systems. In this study, we have characterised these different modes of N-P interaction in detail and for the first time have been able to reconstitute these complexes individually in vitro in the Chandipura Virus (CHPV), a human pathogenic NNS RNA Virus. Using a battery of truncated mutants of the N protein, we have been able to identify two mutually exclusive domains of N involved in differential interaction with the P protein. An unique N-terminal binding site, comprising of amino acids (aa) 1– 180 form the N 0-P interacting region, whereas, C-terminal residues spanning aa 320–390 is instrumental in N-RNA-P interactions. Significantly, the ex-vivo data also supports these observations. Based on these results, we suggest that the P protein acts as N-specific chaperone and thereby partially masking the N-N self-association region, which leads to th
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characterization of the Chandipura Virus leader rna phosphoprotein interaction using single tryptophan mutants and its detection in viral infected cells
Biochimie, 2013Co-Authors: Arunava Roy, Manini Mukherjee, Sanjib Ghosh, Subhradip Mukhopadhyay, Shyam Sundar Maity, Dhrubajyoti ChattopadhyayAbstract:Abstract The phosphoprotein (P protein) of the Chandipura Virus (CHPV), a negative strand RNA Virus, is involved in both transcription and replication of the viral life cycle. Interaction between the P protein and the viral leader (le) RNA under in vitro conditions has been previously reported for CHPV and other negative strand RNA Viruses such as the rinderpest Virus (RPV). However, till date, the region of the P protein involved in le RNA binding remains undefined. Moreover, the in vivo occurrence of this interaction has not been studied before. Here, we have characterised the P protein–le RNA interaction, using single tryptophan mutants of the P protein. The CHPV P protein contains two tryptophan residues located at amino acid position 105 and 135 respectively. Our previous study showed that Trp 135 is located in a buried region within a less polar environment whereas Trp 105 is more solvent-exposed. In this study we have used steady state and time resolved fluorescence spectroscopy at 298 K to show that the buried tryptophan (Trp 135) is involved in the interaction with the le RNA and the more solvent exposed Trp 105 is only slightly perturbed during this interaction. We also show that Trp 135 is responsible for the dimerization of the CHPV P protein. In addition, we have been able to demonstrate for the first time that the P protein–le RNA interaction is detectable in CHPV-infected Vero-76 cells and this interaction is augmented during the replication phase of the viral cycle.
Akhilesh C. Mishra - One of the best experts on this subject based on the ideXlab platform.
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Isolation of Chandipura Virus (VesiculoVirus: Rhabdoviridae) from Sergentomyia species of sandflies from Nagpur, Maharashtra, India.
The Indian journal of medical research, 2014Co-Authors: A. B. Sudeep, Vijay P. Bondre, Gajanan N. Sapkal, M. S. Mavale, Yogesh K. Gurav, Mangesh D. Gokhale, Renjith George, Akhilesh C. MishraAbstract:Background & objectives: An outbreak of acute encephalitis syndrome was reported from Vidarbha region of Maharashtra s0 tate, India, during July 2012. Anti-IgM antibodies against Chandipura Virus (CHPV) were detected in clinical samples. Sandfly collections were done to determine their role in CHPV transmission. Methods: Twenty nine pools of Sergentomyia spp. comprising 625 specimens were processed for Virus isolation in Vero E6 cell line. Diagnostic RT-PCR targeting N-gene was carried out with the sample that showed cytopathic effects (CPE). The PCR product was sequenced, analysed and the sequences were deposited in Genbank database. Results: CPE in Vero E6 cell line infected with three pools was detected at 48 h post infection. However, Virus could be isolated only from one pool. RT-PCR studies demonstrated 527 nucleotide product that confirmed the agent as CHPV. Sequence analysis of the new isolate showed difference in 10-12 nucleotides in comparison to earlier isolates. Interpretation & conclusions: This is perhaps the first isolation of CHPV from Sergentomyia spp. in India and Virus isolation during transmission season suggests their probable role in CHPV transmission. Further studies need to be done to confirm the precise role of Sargentomyia spp. in CHPV transmission.
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Chandipura Virus encephalitis outbreak among children in nagpur division maharashtra 2007
Indian Journal of Medical Research, 2010Co-Authors: Yogesh K. Gurav, Ramesh S. Jadi, Vidya A. Arankalle, Rashmi Gunjikar, Babasaheb V. Tandale, Sanjaykumar S Tikute, Aniruddha V Jamgaonkar, Ravindra K Khadse, S V Jalgaonkar, Akhilesh C. MishraAbstract:Results: A total of 78 AES cases were recorded in children <15 yr of age. Case fatality ratio was 43.6 per cent. Male to female ratio was 1:1.2. Chandipura (CHP) was confirmed in 39 cases. CHPV RNA was detected in both CSF and serum specimens of 2 cases and in serum of 22 cases. Phylogenetic analysis showed 99.98 – 100 per cent nucleotide identity in the sequences studied. Anti-CHPV IgM antibodies were detected in CSF of 2 cases and in serum of 8 cases. Seroconversion to anti-CHPV IgM antibodies was observed in 5 cases. Clinical manifestations of CHP cases (n=38) were fever (100%), convulsion (76.3%), altered sensorium (34.2%), headache (23.7%), vomiting (44.7%) and diarrhoea (23.7%). CHPV RNA was detected in one of two pools of sandflies from affected locality. Interpretation & conclusions: Chandipura Virus was confirmed as the aetiological agent of this acute encephalitis outbreak with high case-fatality among children.
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Chandipura Virus growth kinetics in vertebrate cell lines insect cell lines embryonated eggs
Indian Journal of Medical Research, 2010Co-Authors: Ramesh S. Jadi, A. B. Sudeep, Santosh Kumar, Vidya A. Arankalle, Akhilesh C. MishraAbstract:Background & objectives: Since not much information on Chandipura Virus is available, an attempt was made to study the growth kinetics of the Virus in certain vertebrate, invertebrate cell lines and embryonated chicken eggs. Methods: Comparative study of Chandipura Virus (CHPV) growth kinetics in three vertebrate cell lines [Vero E6, Rhabdo myosarcoma (RD), Porcine stable kidney (PS) cell lines], two insect cell lines [Aedes aegypti (AA) and Phlebotomus papatasi (PP-9) cell lines] and embryonated pathogen free chicken eggs was conducted, by tissue culture infective dose 50 per cent (TCID50) and indirect immunofluorescence assay (IFA). Results: All the cell lines and embryonated egg supported the growth of CHPV and yielded high Virus titre. The vertebrate cell lines showed distinct cytopathic effect (CPE) within 4-6 h post infection (PI), while no CPE was observed in insect cell lines. PP-9 cell line was the most sensitive system to CHPV as viral antigen could be detected at 1 h PI by IFA. Interpretation & conclusions: Our results demonstrated that all the systems were susceptible to CHPV and achieved high yield of Virus. However, the PP-9 cell line had an edge over the others due to its high sensitivity to the Virus which might be useful for detection and isolation of the Virus during epidemics.
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Chandipura Virus growth kinetics in vertebrate cell lines, insect cell lines & embryonated eggs.
The Indian journal of medical research, 2010Co-Authors: Ramesh S. Jadi, A. B. Sudeep, Santosh Kumar, Vidya A. Arankalle, Akhilesh C. MishraAbstract:Background & objectives: Since not much information on Chandipura Virus is available, an attempt was made to study the growth kinetics of the Virus in certain vertebrate, invertebrate cell lines and embryonated chicken eggs. Methods: Comparative study of Chandipura Virus (CHPV) growth kinetics in three vertebrate cell lines [Vero E6, Rhabdo myosarcoma (RD), Porcine stable kidney (PS) cell lines], two insect cell lines [Aedes aegypti (AA) and Phlebotomus papatasi (PP-9) cell lines] and embryonated pathogen free chicken eggs was conducted, by tissue culture infective dose 50 per cent (TCID50) and indirect immunofluorescence assay (IFA). Results: All the cell lines and embryonated egg supported the growth of CHPV and yielded high Virus titre. The vertebrate cell lines showed distinct cytopathic effect (CPE) within 4-6 h post infection (PI), while no CPE was observed in insect cell lines. PP-9 cell line was the most sensitive system to CHPV as viral antigen could be detected at 1 h PI by IFA. Interpretation & conclusions: Our results demonstrated that all the systems were susceptible to CHPV and achieved high yield of Virus. However, the PP-9 cell line had an edge over the others due to its high sensitivity to the Virus which might be useful for detection and isolation of the Virus during epidemics.
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Development and evaluation of a real-time one step reverse-transcriptase PCR for quantitation of Chandipura Virus.
BMC infectious diseases, 2008Co-Authors: Santosh Kumar, Ramesh S. Jadi, Akhilesh C. Mishra, Sudeep B Anakkathil, Babasaheb V. Tandale, Vidya A. ArankalleAbstract:Background Chandipura Virus (CHPV), a member of family Rhabdoviridae was attributed to an explosive outbreak of acute encephalitis in children in Andhra Pradesh, India in 2003 and a small outbreak among tribal children from Gujarat, Western India in 2004. The case-fatality rate ranged from 55–75%. Considering the rapid progression of the disease and high mortality, a highly sensitive method for quantifying CHPV RNA by real-time one step reverse transcriptase PCR (real-time one step RT-PCR) using TaqMan technology was developed for rapid diagnosis.
Vidya A. Arankalle - One of the best experts on this subject based on the ideXlab platform.
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Whole Genomes of Chandipura Virus Isolates and Comparative Analysis with Other RhabdoViruses
PloS one, 2012Co-Authors: Sarah S. Cherian, Santosh Kumar, Rashmi Gunjikar, Arpita Banerjee, Vidya A. ArankalleAbstract:The Chandipura Virus (CHPV) belonging to the VesiculoVirus genus and Rhabdoviridae family, has recently been associated with a number of encephalitis epidemics, with high mortality in children, in different parts of India. No full length genome sequences of CHPV isolates were available in GenBank and little is known about the molecular markers for pathogenesis. In the present study, we provide the complete genomic sequences of four isolates from epidemics during 2003–2007. These sequences along with the deduced sequence of the prototype isolate of 1965 were analysed using phylogeny, motif search, homology modeling and epitope prediction methods. Comparison with other rhaboViruses was also done for functional extrapolations. All CHPV isolates clustered with the Isfahan Virus and maintained several functional motifs of other rhabdoViruses. A notable difference with the prototype vesiculoVirus, Vesicular Stomatitis Virus was in the L-domain flanking sequences of the M protein that are known to be crucial for interaction with host proteins. With respect to the prototype isolate, significant additional mutations were acquired in the 2003–2007 isolates. Several mutations in G mapped onto probable antigenic sites. A mutation in N mapped onto regions crucial for N-N interaction and a putative T-cell epitope. A mutation in the Casein kinase II phosphorylation site in P may attribute to increased rates of phosphorylation. Gene junction comparison revealed changes in the M-G junction of all the epidemic isolates that may have implications on read-through and gene transcription levels. The study can form the basis for further experimental verification and provide additional insights into the virulence determinants of the CHPV.
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Chandipura Virus encephalitis outbreak among children in nagpur division maharashtra 2007
Indian Journal of Medical Research, 2010Co-Authors: Yogesh K. Gurav, Ramesh S. Jadi, Vidya A. Arankalle, Rashmi Gunjikar, Babasaheb V. Tandale, Sanjaykumar S Tikute, Aniruddha V Jamgaonkar, Ravindra K Khadse, S V Jalgaonkar, Akhilesh C. MishraAbstract:Results: A total of 78 AES cases were recorded in children <15 yr of age. Case fatality ratio was 43.6 per cent. Male to female ratio was 1:1.2. Chandipura (CHP) was confirmed in 39 cases. CHPV RNA was detected in both CSF and serum specimens of 2 cases and in serum of 22 cases. Phylogenetic analysis showed 99.98 – 100 per cent nucleotide identity in the sequences studied. Anti-CHPV IgM antibodies were detected in CSF of 2 cases and in serum of 8 cases. Seroconversion to anti-CHPV IgM antibodies was observed in 5 cases. Clinical manifestations of CHP cases (n=38) were fever (100%), convulsion (76.3%), altered sensorium (34.2%), headache (23.7%), vomiting (44.7%) and diarrhoea (23.7%). CHPV RNA was detected in one of two pools of sandflies from affected locality. Interpretation & conclusions: Chandipura Virus was confirmed as the aetiological agent of this acute encephalitis outbreak with high case-fatality among children.
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Chandipura Virus growth kinetics in vertebrate cell lines insect cell lines embryonated eggs
Indian Journal of Medical Research, 2010Co-Authors: Ramesh S. Jadi, A. B. Sudeep, Santosh Kumar, Vidya A. Arankalle, Akhilesh C. MishraAbstract:Background & objectives: Since not much information on Chandipura Virus is available, an attempt was made to study the growth kinetics of the Virus in certain vertebrate, invertebrate cell lines and embryonated chicken eggs. Methods: Comparative study of Chandipura Virus (CHPV) growth kinetics in three vertebrate cell lines [Vero E6, Rhabdo myosarcoma (RD), Porcine stable kidney (PS) cell lines], two insect cell lines [Aedes aegypti (AA) and Phlebotomus papatasi (PP-9) cell lines] and embryonated pathogen free chicken eggs was conducted, by tissue culture infective dose 50 per cent (TCID50) and indirect immunofluorescence assay (IFA). Results: All the cell lines and embryonated egg supported the growth of CHPV and yielded high Virus titre. The vertebrate cell lines showed distinct cytopathic effect (CPE) within 4-6 h post infection (PI), while no CPE was observed in insect cell lines. PP-9 cell line was the most sensitive system to CHPV as viral antigen could be detected at 1 h PI by IFA. Interpretation & conclusions: Our results demonstrated that all the systems were susceptible to CHPV and achieved high yield of Virus. However, the PP-9 cell line had an edge over the others due to its high sensitivity to the Virus which might be useful for detection and isolation of the Virus during epidemics.
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Intracranial administration of P gene siRNA protects mice from lethal Chandipura Virus encephalitis.
PloS one, 2010Co-Authors: Santosh Kumar, Vidya A. ArankalleAbstract:Background In parts of India, Chandipura Virus (CHPV) has emerged as an encephalitis causing pathogen in both epidemic and sporadic forms. This pediatric disease follows rapid course leading to 55–75% mortality. In the absence of specific treatment, effectiveness of RNA interference (RNAi) was evaluated.
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Chandipura Virus growth kinetics in vertebrate cell lines, insect cell lines & embryonated eggs.
The Indian journal of medical research, 2010Co-Authors: Ramesh S. Jadi, A. B. Sudeep, Santosh Kumar, Vidya A. Arankalle, Akhilesh C. MishraAbstract:Background & objectives: Since not much information on Chandipura Virus is available, an attempt was made to study the growth kinetics of the Virus in certain vertebrate, invertebrate cell lines and embryonated chicken eggs. Methods: Comparative study of Chandipura Virus (CHPV) growth kinetics in three vertebrate cell lines [Vero E6, Rhabdo myosarcoma (RD), Porcine stable kidney (PS) cell lines], two insect cell lines [Aedes aegypti (AA) and Phlebotomus papatasi (PP-9) cell lines] and embryonated pathogen free chicken eggs was conducted, by tissue culture infective dose 50 per cent (TCID50) and indirect immunofluorescence assay (IFA). Results: All the cell lines and embryonated egg supported the growth of CHPV and yielded high Virus titre. The vertebrate cell lines showed distinct cytopathic effect (CPE) within 4-6 h post infection (PI), while no CPE was observed in insect cell lines. PP-9 cell line was the most sensitive system to CHPV as viral antigen could be detected at 1 h PI by IFA. Interpretation & conclusions: Our results demonstrated that all the systems were susceptible to CHPV and achieved high yield of Virus. However, the PP-9 cell line had an edge over the others due to its high sensitivity to the Virus which might be useful for detection and isolation of the Virus during epidemics.
Sanjay Gupta - One of the best experts on this subject based on the ideXlab platform.
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Host interactions of Chandipura Virus matrix protein
Acta tropica, 2015Co-Authors: Sreejith Rajasekharan, Jyoti Rana, Kapila Kumar, Amita Gupta, Vijay K. Chaudhary, Sanjay GuptaAbstract:The rhabdoVirus matrix (M) protein is a multifunctional virion protein that plays major role in Virus assembly and budding, Virus-induced inhibition of host gene expression and cytopathic effects observed in infected cells. The myriad roles played by this protein in the Virus biology make it a critical player in viral pathogenesis. Therefore, discerning the interactions of this protein with host can greatly facilitate our understanding of Virus infections, ultimately leading to both improved therapeutics and insight into cellular processes. Chandipura Virus (CHPV; Family Rhabdoviridae, Genus VesiculoVirus) is an emerging rhabdoVirus responsible for several outbreaks of fatal encephalitis among children in India. The present study aims to screen the human fetal brain cDNA library for interactors of CHPV M protein using yeast two-hybrid system. Ten host protein interactors were identified, three of which were further validated by affinity pull down and protein interaction ELISA. The study identified novel human host interactors for CHPV which concurred with previously described associations in other human Viruses.
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Neuroinvasion by Chandipura Virus.
Acta tropica, 2014Co-Authors: Sreejith Rajasekharan, Sahil Gulati, Jyoti Rana, Vandana Gupta, Sanjay GuptaAbstract:Abstract Chandipura Virus (CHPV) is an arthropod borne rhabdoVirus associated with acute encephalitis in children below the age of 15 years in the tropical states of India. Although the entry of the Virus into the nervous system is among the crucial events in the pathogenesis of CHPV, the exact mechanism allowing CHPV to invade the central nervous system (CNS) is currently poorly understood. In the present review, based on the knowledge of host interactors previously predicted for CHPV, along with the support from experimental data available for other encephalitic Viruses, the authors have speculated the various plausible modes by which CHPV could surpass the blood–brain barrier and invade the CNS to cause encephalitis whilst evading the host immune surveillance. Collectively, this review provides a conservative set of potential interactions that can be employed for future experimental validation with a view to better understand the neuropathogenesis of CHPV.
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Elucidating the interacting domains of Chandipura Virus nucleocapsid protein.
Advances in virology, 2013Co-Authors: Kapila Kumar, Sahil Gulati, Sreejith Rajasekharan, Jyoti Rana, Reema Gabrani, Amita Gupta, Vijay K. Chaudhary, Chakresh Kumar Jain, Sanjay GuptaAbstract:The nucleocapsid (N) protein of Chandipura Virus (CHPV) plays a crucial role in viral life cycle, besides being an important structural component of the virion through proper organization of its interactions with other viral proteins. In a recent study, the authors had mapped the associations among CHPV proteins and shown that N protein interacts with four of the viral proteins: N, phosphoprotein (P), matrix protein (M), and glycoprotein (G). The present study aimed to distinguish the regions of CHPV N protein responsible for its interactions with other viral proteins. In this direction, we have generated the structure of CHPV N protein by homology modeling using SWISS-MODEL workspace and Accelrys Discovery Studio client 2.55 and mapped the domains of N protein using PiSQRD. The interactions of N protein fragments with other proteins were determined by ZDOCK rigid-body docking method and validated by yeast two-hybrid and ELISA. The study revealed a unique binding site, comprising of amino acids 1–30 at the N terminus of the nucleocapsid protein (N1) that is instrumental in its interactions with N, P, M, and G proteins. It was also observed that N2 associates with N and G proteins while N3 interacts with N, P, and M proteins.
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Predicting the host protein interactors of Chandipura Virus using a structural similarity–based approach
Pathogens and disease, 2013Co-Authors: Sreejith Rajasekharan, Sahil Gulati, Jyoti Rana, Vandana Gupta, Sanjeev K. Sharma, Sanjay GuptaAbstract:Chandipura Virus (CHPV), alike other pathogens, exploits the cellular infrastructure of their hosts through complex network of interactions for successful infection. CHPV being a recently emerged pediatric encephalitic Virus, the mechanisms involved in the establishment of viral persistence are still ill defined. Because the protein interface between CHPV and its host provides one means by which the Virus invades and seize control of their human host machinery, the authors in this study have employed computational methods to create a network of putative protein–protein interactions between CHPV and its human host to shed light on the hitherto less-known CHPV biology. On the basis of the 2105 potential interactions predicted among 1650 human proteins and the five proteins of CHPV, the authors decipher the probable mode by which the Virus manipulates the biological pathways of its host toward its own end and replicates while evading the immune system. Identification of such conserved set of putative interactions that allow the Virus to take control of the host has the potential to deepen our understanding of the Virus-specific remodeling processes of the host cell and illuminate new arenas of disease intervention.
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predicting the host protein interactors of Chandipura Virus using a structural similarity based approach
Pathogens and Disease, 2013Co-Authors: Sreejith Rajasekharan, Sahil Gulati, Jyoti Rana, Vandana Gupta, Sanjeev K. Sharma, Sanjay GuptaAbstract:Chandipura Virus (CHPV), alike other pathogens, exploits the cellular infrastructure of their hosts through complex network of interactions for successful infection. CHPV being a recently emerged pediatric encephalitic Virus, the mechanisms involved in the establishment of viral persistence are still ill defined. Because the protein interface between CHPV and its host provides one means by which the Virus invades and seize control of their human host machinery, the authors in this study have employed computational methods to create a network of putative protein–protein interactions between CHPV and its human host to shed light on the hitherto less-known CHPV biology. On the basis of the 2105 potential interactions predicted among 1650 human proteins and the five proteins of CHPV, the authors decipher the probable mode by which the Virus manipulates the biological pathways of its host toward its own end and replicates while evading the immune system. Identification of such conserved set of putative interactions that allow the Virus to take control of the host has the potential to deepen our understanding of the Virus-specific remodeling processes of the host cell and illuminate new arenas of disease intervention.
Arunava Roy - One of the best experts on this subject based on the ideXlab platform.
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Role of tryptophan 135 of Chandipura Virus phosphoprotein P in dimerization and complex formation with leader RNA: structural aspect using time resolved anisotropy and simulation
RSC Advances, 2015Co-Authors: Manini Mukherjee, Dhrubajyoti Chattopadhyay, Aditya Kumar Sarkar, Arunava Roy, Pinki Saha Sardar, Ansuman Lahiri, Sanjib GhoshAbstract:The aggregation of phosphoprotein P of Chandipura Virus (CHPV) and its interaction with viral leader (le) RNA in aqueous buffer and 40% ethylene glycol (EG)-buffer have been characterised using two single tryptophan (Trp/W) mutants W105F and W135F. The longer rotational correlation time [(θC)T] originating from overall motion observed at 300 nM concentration conforms to a globular structure of the monomer of WT and both the mutants. The (θC)T values also indicate that W135F does not form a dimer at 1500 nM concentration; while a dimer with disordered structure is predicted for both WT and W105F. The complexes of WT and W105F with le RNA at monomeric and dimeric conditions are indicated to have tight core packing. Dimerization and complex formation at both the concentrations enhance the correlation time arising from the localised motion of Trp side chain [(θC)S] for WT and W105F predicting hindered localized rotation of Trp 135. Comparative protein modelling and molecular dynamics (MD) simulations using the amino acids domain ranging from 105–168 of the full length CHPVP based on the vesicular stomatitis Virus phosphoprotein (VSVP) also indicate that formation of dimers are more feasible for WT and W105F compared to W135F.
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A peptide targeted against phosphoprotein and leader RNA interaction inhibits growth of Chandipura Virus -- an emerging rhabdoVirus.
Antiviral research, 2013Co-Authors: Arunava Roy, Dhrubajyoti Chattopadhyay, Smarajit Polley, Prasenjit Chakraborty, Sabita RoyAbstract:Abstract The fatal illness caused by Chandipura Virus (CHPV), an emerging pathogen, presently lacks any therapeutic option. Previous research suggested that interaction between the virally encoded phosphoprotein (P) and the positive sense leader RNA (le-RNA) may play an important role in the viral lifecycle. In this report, we have identified a β -sheet/loop motif in the C-terminal domain of the CHPV P protein as essential for this interaction. A synthetic peptide encompassing this motif and spanning a continuous stretch of 36 amino acids (Pep 208–243 ) was found to bind the le-RNA in vitro and inhibit CHPV growth in infected cells. Furthermore, a stretch of three amino acid residues at position 217–219 was identified as essential for this interaction, both in vitro and in infected cells. siRNA knockdown-rescue experiments demonstrated that these three amino acid residues are crucial for the leader RNA binding function of P protein in the CHPV life cycle. Mutations of these three amino acid residues render the peptide completely ineffective against CHPV. Effect of inhibition of phosphoprotein–leader RNA interaction on viral replication was assayed. Peptide Pep 208–243 tagged with a cell penetrating peptide was found to inhibit CHPV replication as ascertained by real time RT-PCR. The specific inhibition of viral growth observed using this peptide suggests a new possibility for designing of anti-viral agents against Mononegavirale group of human Viruses.
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Interaction of Chandipura Virus N and P Proteins: Identification of Two Mutually Exclusive Domains of N Involved in Interaction with P
2013Co-Authors: Arindam Mondal ¤a, Arunava Roy, Ipto Sarkar, Jishnu Mukherjee ¤b, Tridib Ganguly ¤c, Dhrubajyoti ChattopadhyayAbstract:The nucleocapsid protein (N) and the phosphoprotein (P) of nonsegmented negative-strand (NNS) RNA Viruses interact with each other to accomplish two crucial events necessary for the viral replication cycle. First, the P protein binds to the aggregation prone nascent N molecules maintaining them in a soluble monomeric (N 0) form (N 0-P complex). It is this form that is competent for specific encapsidation of the viral genome. Second, the P protein binds to oligomeric N in the nucleoprotein complex (N-RNA-P complex), and thereby facilitates the recruitment of the viral polymerase (L) onto its template. All previous attempts to study these complexes relied on co-expression of the two proteins in diverse systems. In this study, we have characterised these different modes of N-P interaction in detail and for the first time have been able to reconstitute these complexes individually in vitro in the Chandipura Virus (CHPV), a human pathogenic NNS RNA Virus. Using a battery of truncated mutants of the N protein, we have been able to identify two mutually exclusive domains of N involved in differential interaction with the P protein. An unique N-terminal binding site, comprising of amino acids (aa) 1– 180 form the N 0-P interacting region, whereas, C-terminal residues spanning aa 320–390 is instrumental in N-RNA-P interactions. Significantly, the ex-vivo data also supports these observations. Based on these results, we suggest that the P protein acts as N-specific chaperone and thereby partially masking the N-N self-association region, which leads to th
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characterization of the Chandipura Virus leader rna phosphoprotein interaction using single tryptophan mutants and its detection in viral infected cells
Biochimie, 2013Co-Authors: Arunava Roy, Manini Mukherjee, Sanjib Ghosh, Subhradip Mukhopadhyay, Shyam Sundar Maity, Dhrubajyoti ChattopadhyayAbstract:Abstract The phosphoprotein (P protein) of the Chandipura Virus (CHPV), a negative strand RNA Virus, is involved in both transcription and replication of the viral life cycle. Interaction between the P protein and the viral leader (le) RNA under in vitro conditions has been previously reported for CHPV and other negative strand RNA Viruses such as the rinderpest Virus (RPV). However, till date, the region of the P protein involved in le RNA binding remains undefined. Moreover, the in vivo occurrence of this interaction has not been studied before. Here, we have characterised the P protein–le RNA interaction, using single tryptophan mutants of the P protein. The CHPV P protein contains two tryptophan residues located at amino acid position 105 and 135 respectively. Our previous study showed that Trp 135 is located in a buried region within a less polar environment whereas Trp 105 is more solvent-exposed. In this study we have used steady state and time resolved fluorescence spectroscopy at 298 K to show that the buried tryptophan (Trp 135) is involved in the interaction with the le RNA and the more solvent exposed Trp 105 is only slightly perturbed during this interaction. We also show that Trp 135 is responsible for the dimerization of the CHPV P protein. In addition, we have been able to demonstrate for the first time that the P protein–le RNA interaction is detectable in CHPV-infected Vero-76 cells and this interaction is augmented during the replication phase of the viral cycle.
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Characterization of the Chandipura Virus leader RNA–phosphoprotein interaction using single tryptophan mutants and its detection in viral infected cells
Biochimie, 2012Co-Authors: Arunava Roy, Manini Mukherjee, Sanjib Ghosh, Subhradip Mukhopadhyay, Shyam Sundar Maity, Dhrubajyoti ChattopadhyayAbstract:Abstract The phosphoprotein (P protein) of the Chandipura Virus (CHPV), a negative strand RNA Virus, is involved in both transcription and replication of the viral life cycle. Interaction between the P protein and the viral leader (le) RNA under in vitro conditions has been previously reported for CHPV and other negative strand RNA Viruses such as the rinderpest Virus (RPV). However, till date, the region of the P protein involved in le RNA binding remains undefined. Moreover, the in vivo occurrence of this interaction has not been studied before. Here, we have characterised the P protein–le RNA interaction, using single tryptophan mutants of the P protein. The CHPV P protein contains two tryptophan residues located at amino acid position 105 and 135 respectively. Our previous study showed that Trp 135 is located in a buried region within a less polar environment whereas Trp 105 is more solvent-exposed. In this study we have used steady state and time resolved fluorescence spectroscopy at 298 K to show that the buried tryptophan (Trp 135) is involved in the interaction with the le RNA and the more solvent exposed Trp 105 is only slightly perturbed during this interaction. We also show that Trp 135 is responsible for the dimerization of the CHPV P protein. In addition, we have been able to demonstrate for the first time that the P protein–le RNA interaction is detectable in CHPV-infected Vero-76 cells and this interaction is augmented during the replication phase of the viral cycle.