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Indian Journal of Virology
Year : 2008, Volume : 19, Issue : 1

Print ISSN : 0970-2822.

P-06. NSs encoded by Peanut bud necrosis virus (tomato) is a multifunctional enzyme

Bhushan Lokesh, Srisathiyanarayanan D., Bhat Amruta S., Savithri H.S.

Department of Biochemistry, Indian Institute of Science, Bangalore, India.

Abstracts of the papers presented at the International Conference of Indian Virological Society on “Emerging and Re-emerging viral Diseases of the Tropics and Subtropics” at Indian Agricultural Research Institute, New Delhi, India, December 11–14, 2007.

Summary

Peanut bud necrosis virus (PBNV) belongs to the genus Tospovirus. NSs of tomato spotted wilt virus (type member of tospovirus) has been implicated as suppressor of gene silencing in vivo. With a view to elucidate the mechanism by which NSs could act as a suppressor of gene silencing and examine the multifunctional role of NSs in the life cycle of the virus, the PBNV (To) NSs gene was amplified by RT-PCR, cloned into pRSET C vector and overexpressed in E.coli C43 (DE3) pLysS cells. The recombinant NSs (rNSs) protein was purified by Ni-NTA chromatography. Purified rNSs exhibited an RNA stimulated NTPase activity with ATP as the preferred substrate. Optimum temperature and pH for this activity are 25°C and 7.0 respectively. NTPase activity of rNSs is metal ion dependent and is inhibited by EDTA as well as an ATP analog. The rNSs could also hydrolyze dNTPs. In addition to NTPase and dNTPase activities, rNSs exhibited 5’ RNA/DNA phosphatase activity. We propose that NSs removes the 5’ phosphate from dsRNA/SiRNA, as a result of which Dicer/RICS complex is not able to recognize the respective substrates. Hence, PTGS pathway is not triggered upon infection of PBNV. Interestingly, rNSs specifically binds to ssRNA and also interacts with nucleoprotein (NP) of PBNV. Our results suggest that C-terminus of NP is not important for its interaction with NSs. NP is involved in encapsidation of the viral genome and in viral replication. Thus, via the interaction with NP, NSs could also play role in viral replication/assembly.

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