VACCINIA VIRUS A17L GENE-PRODUCT IS ESSENTIAL FOR AN EARLY STEP IN VIRION MORPHOGENESIS

Vaccinia virus (VV) A17L gene encodes a 23-kDa protein that is proteol ytically cleaved to generate a 21-kDa product that is incorporated int o the viral particles. We have previously shown that the 21-kDa protei n forms a stable complex with the W 14-kDa envelope protein and sugges ted that the 21-kDa protein may serve to anchor the 14-kDa protein to the envelope of the virion (D. Rodriguez, J. R. Rodriguez, and M. Este ban, J. Virol. 67:3435-3440, 1993). To study the role of the 21-kDa pr otein in virion assembly, in this investigation we generated a W recom binant, WindA17L, that contains an inducible A17L gene regulated by th e E. call repressor/operator system. In the absence of the inducer, sh utoff of the A17L gene was complete, and this shutoff correlated with a reduction in virus yields of about 3 log units. Although early and l ate viral polypeptides are normally synthesized in the absence of the A17L gene product, proteolytic processing of the major p4a and p4b cor e proteins was clearly impaired under these conditions. Electron micro scopy examination of cells infected in the absence of isopropylthiogal actopyranoside (IPTG) revealed that virion morphogenesis was completel y arrested at a very early stage, even prior to the formation of cresc ent-shaped membranes, which are the first distinguishable viral struct ures. Only electron-dense structures similar to rifampin bodies, but d evoid of membranes, could be observed in the cytoplasm of cells infect ed with WindA17L under nonpermissive conditions. Considering the most recent assembly model presented by Sodeik et al. (B. Sodeik, R. W. Dom s, M. Ericsson, G. Hiller, C. E. Machamer, W. van't Hof, G. van Meer, B. Moss, and G. Griffiths, J. Cell Biol. 121:521-541, 1993), we propos e that this protein is targeted to the intermediate compartment and is involved in the recruitment of these membranes to the viral factories , where it forms the characteristic crescent structures that subsequen tly result in the formation of virions.