Oligomerization, secretion, and biological function of an anchor-free parainfluenza virus type 2 (PI2) fusion protein

A number of studies indicate that the transmembrane domain, the cytoplasmic domain, or both regions of viral surface glycoproteins are involved in qua ternary structure formation. In this report, the transmembrane domain and c ytoplasmic tail coding sequence of the fusion (F) glycoprotein gene from pa rainfluenza type 2 virus was truncated by PCR and the resulting gene (PI2F' ) was expressed in HeLa-T4 cells by using the vaccinia virus-T7 transient e xpression system. Pulse-chase experiments indicated that the anchor-free P1 2F' was expressed and processed into F-1 and F-2 subunits. Both the process ed and the unprocessed anchor-free P12F' proteins were found to be efficien tly secreted into the culture medium. Examination of the oligomeric form of the anchor-free P12F' by chemical cross-linking demonstrated that it assem bles posttranslationally into dimers and trimers with a pattern similar to that of the wild-type PI2F protein. In an effort to better understand the b iological properties of the truncated form of P12F', we anchored PI2F' by a glycosyl-phosphatidylinositol (GPI) linkage. The GPI-anchored P12F' protei n, when coexpressed with PI2HN, did not induce cell fusion seen as syncytiu m formation, but was found to initiate lipid mixing (hemifusion) as observe d by transfer of R-18 rhodamine from red blood cells to the GPI-PI2F'/PI2HN cotransfected cells. The results therefore indicate that the extracellular domain of the PI2 fusion protein contains not only the structural informat ion sufficient to direct assembly into higher oligomers, but also is compet ent to initiate membrane fusion, suggesting that the anchor-free P12F' may be useful for further structural studies. (C) 2000 Academic Press.