Pseudotype formation with La Crosse virus glycoproteins.

Abstract

Pseudotype formation is a powerful tool for analysing mechanisms of virus neutralization and entry, since it allows for analysis of glycoprotein properties without the necessity for preparing recombinant genomes. Using recombinant vaccinia viruses, we prepared pseudotypes of La Crosse virus with recombinant glycoproteins cloned from the monoclonal antibody (MAb)-resistant variant V31. The resulting pseudotypes became partially resistant to MAb 807-31. Furthermore, when the V31 glycoproteins were incorporated into a second MAb-resistant variant (V33), the pseudotyped virus became sensitive to neutralization by the MAb (807-33) originally used in its selection. These results suggest a simple technique for the incorporation of glycoprotein mutations into bunyaviruses, allowing analysis of mechanisms of neutralization and other virus entry functions.La Crosse virus (LAC) is a member of the California serogroup of the genus Bunyavirus of the family Bunyaviridae, and like all members of the family has a tri-segmented genome with segments designated L (large), M (medium) and S (small). The L RNA segment encodes the L or polymerase protein. The M segment encodes a single open reading frame that is processed into the two viral glycoproteins G1 and G2, as well as NSm -a non-structural protein of unknown function. The S segment encodes the nucleocapsid protein and another nonstructural protein (NSs) in overlapping reading frames (Elliott, 1990).The G1 and G2 glycoproteins of LAC have been expressed using vaccinia virus (VV) vectors. Several of the glycoprotein functions, such as targeting to the Golgi apparatus (a characteristic of bunyaviruses) and pH-dependent fusogenicity, are faithfully reproduced by the recombinant proteins (Bupp et

Oropouche Virus Infection and Pathogenesis Are Restricted by MAVS, IRF-3, IRF-7, and Type I Interferon Signaling Pathways in Nonmyeloid Cells

“…This stock was clarified and stored at Ϫ80°C. The titer of LACV was calculated as 2.7 ϫ 10 7 FFU/ml using the monoclonal antibodies (MAb; 807-31 and 807-33) that also were provided by Andrew Pekosz (42,43). Experiments with LACV were conducted in BSL2 and A-BSL3 facilities.…”

Analysis of the medium (M) segment sequence of Guaroa virus and its comparison to other orthobunyaviruses

“…It is conceivable that these positions contribute to the serological reaction of GROV. The mutation at (Bupp & Gonzá lez-Scarano, 1998). Amino acid positions for the GROV sequence are indicated at the end of each line.…”