Mapping of interacting domains between the Nucleoprotein and Z matrix protein of New World arenaviruses

LEVINGSTON MACLEOD J.M.; CASABONA J.C.; GÓMEZ G.; LOUREIRO M.E.; WILDA, M.; LÓPEZ N.

Brugge

Congreso; Negative Strand Virus Meeting; 2010

Negative Strand Virus Meeting

The New World arenaviruses include Tacaribe virus (TCRV) and its closely related pathogenic Junin virus (JUNV), whose bisegmented genome encodes the nucleoprotein (N), the precursor of the envelope glycoprotein complex (GP), the polymerase (L) and a small RING finger matrix protein (Z). We have recently developed a reverse genetic system that allows the packaging of TCRV-like nucleocapsids along with Z and GP of JUNV into infectious virus-like particles (VLPs). We demostrated that Z is sufficient to recruit N within Z-containing VLPs, and that the integrity of the RING structure and residue L79 within Z are critical for Z-N interactions involved in the incorporation of both nucleocapsids and GP into budding particles (J. Virol. 83: 7029–7039, 2009).To find additional residues important for assembly, Z mutants generated by single alanine substitutions were examined for their ability to colocalize with N in vivo, to recruit N into Z-directed VLPs and to drive infectious VLP formation. Thus, we could identify further residues within Z that participate in its binding to N. The Z-binding region within TCRV N was also mapped. The interaction of N deletion mutants with Z protein from either TCRV or JUNV was assessed by coimmunoprecipitation assays and N mutant-incorporation into VLPs was tested. The C-terminal region of N, which comprises a putative Zinc finger motif, was found to be required for both intracellular N-Z interaction and the assembly of N into Z-directed VLPs.