[Studies on bovine leukemia virus envelope glycoprotein gp30 YXXL sequences in virus infection and fusogenic activity].

The bovine leukemia virus (BLV) envelope transmembrane protein (gp30) contains three YXXL sequences at its cytoplasmic tail. It is known that N-terminal two of these sequences participate in the induction of B cell activation when chimeric proteins in which cytoplasmic domain of CD8-alpha has been replaced with that of BLV gp30 are stimulated by anti-CD8-alpha antibody. In addition to such signal transduction activity, the two tyrosines in the YXXL sequences also appear to involve infection with high viral loads and their maintenance in the sheep experimentally infected with BLV. To analyze detailed biological relevance of these sequences in vitro, we constructed a full-length BLV-infectious molecular clone with two copies of long terminal repeats (LTRs), designated pBLV-IF, and then changed residues Y487, L490, Y498, L501 or Y487 plus Y498 in gp30 cytoplasmic tail to Ala by site-directed mutagenesis. Introduction of molecular clones of wild-type and mutants into COS-1 cells revealed that all mutated molecular clones synthesized matured envelope proteins and released virus particles into growth medium. Serial passages of transient transfectants with the molecular clones and cell-free inoculation resulted in the reduction of infectivity by mutation of Y498 and Y487 plus Y498. In viral penetration, but not specific virus-cell binding, mutations of Y498 and Y487 plus Y498 substantially reduced the potential. Mutations of Y487, Y498 and Y487 plus Y498 increased syncytium-forming potential with increasing expression of envelope proteins on cell surface. In contrast, mutations of L490 and L501 affected neither infectivity nor syncytium-formation. Altogether, our data indicate that the YXXL sequences play a critical role during virus penetration and fusogenesity in the life cycle of BLV and regulate surface expression of envelope proteins.