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Functional characterization of rNSs protein of PBNV - a suppressor of gene silencing

Bhushan, Lokesh and Srisathiyanarayanan, * and Savithri, HS (2008) Functional characterization of rNSs protein of PBNV - a suppressor of gene silencing. In: Indian Journal Of Virology, 19 (1). p. 53.

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Abstract

RNA interference (RNAi) or post transcriptional gene silencing is a novel mechanism by which the organism protects itself against invasion by viruses. Viruses encode proteins that suppress this defense mechanism and establish themselves in susceptible hosts. There is no sequence similarity among different viral encoded supressors of gene silencing and their mechanism of suppression is also different. Peanut bud necrosis virus (PBNV) belonging to the tospovirus genus has emerged as major pathogen causing severe crop loss in many species of Leguminosae, Solanaceae and Cucurbitaceae crops all over India. A Tospovirus, infecting tomato plants in Karnataka, was isolated and purified and was shown to be a strain of PBNV. Among the proteins encoded by this virus, the NSs protein from the small RNA genome is implicated to be a suppressor of gene silencing in plants. The NSs gene of PBNV was cloned in pRSET C vector and overexpressed in E. coli C43 cells. The recombinant NSs (rNSs) protein was purified by Ni-NTA chromatography. The purified protein exhibited CD and fluorescence spectra typical of globular proteins. In order to understand the role of NSs in suppression of gene silencing, the functional characterization of the protein was undertaken. Interestingly, the rNSs protein exhibited RNA stimulated ATPase activity, which was optimal at pH 7.0 and at room temperature. The activity was inhibited completely by the addition of 5 mM EDTA or ATP analog. The rNSs could also hydrolyze other NTPs and dNTPs. In addition to the NTPase and dNTPase activities, the rNSs exhibited 5’phospatase activity. One of the requirements for recognition of dsRNA/siRNA by the DICER/RICS complex is that they should have a 5’ phosphate. It is possible that the removal of 5’phosphate by the NSs renders the dsRNA/siRNA incapable of binding to DICER/RISC complex leading to suppression of gene silencing.

Item Type: Journal Article
Additional Information: Copyright of this article belongs to Indian Virological Society.
Department/Centre: Division of Biological Sciences > Biochemistry
Date Deposited: 22 Jul 2009 10:20
Last Modified: 22 Jul 2009 10:20
URI: http://eprints.iisc.ernet.in/id/eprint/17829

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