Identification of retroviral late domains as determinants of particle size

Retroviral Gag proteins, in the absence of any other viral products, induce budding and release of spherical, virus-like particles from the plasma mem brane. Gag-produced particles, like those of authentic retrovirions, are no t uniform in diameter but nevertheless fall within a fairly narrow distribu tion of sizes. For the human immunodeficiency virus type 1 (HIV-1) Gag prot ein, we recently reported that elements important for controlling particle size are contained within the C-terminal region of Gag, especially within t he p6 sequence (L. Garnier, L. Ratner, B. Rovinski, S.-X. Cao, and J. W. Wi lls, J. Virol. 72:4667-4677, 1998). Deletions and substitutions throughout this sequence result in the release of very large particles. Because the si ze determinant could not be mapped to any one of the previously defined fun ctions within p6, it seemed likely that its activity requires the overall p roper folding of this region of Gag. This left open the possibility of the size determinant residing in a subdomain of p6, and in this study, we exami ned whether the late domain (the region of Gag that is critical for the vir us-cell separation step) is involved in controlling particle size. We found that particles of normal size are produced when p6 is replaced with the to tally unrelated late domain sequences from Rous sarcoma virus (contained in its p2b sequence) or equine infectious anemia virus (contained in p9). In addition, we found that the large particles released in the absence of p6 r equire the entire CA and adjacent spacer peptide sequences, whereas these i nternal sequences of HIV-1 Gag are not needed for budding (or proper size) when a late domain is present. Thus, it appears the requirements for buddin g are very different in the presence and absence of p6.