Co-translational trimerization of the reovirus cell attachment protein

The reovirus cell attachment protein, sigma 1, is a trimer with a 'lollipop' structure. Recent findings indicate that the N-terminal fibrous tail and the C-terminal globular head each possess a distinct trimerization domain. The region responsible for N-terminal trimerization (formation of a triple alpha-helical coiled-coil) is located at the N-terminal one-third of sigma 1. In this study, we investigated the temporality and ATP requirement of this trimerization event in the context of sigma 1 biogenesis. In vitro co-synthesis of the full-length (FL) and a C-terminally truncated (d44) sigma 1 protein revealed a preference for homotrimer over heterotrimer formation, suggesting that assembly at the N-terminus occurs co-translationally. This was corroborated by the observation that polysome-associated sigma 1 chains were trimeric as well as monomeric. Truncated proteins (d234 and d294) with C-terminal deletions exceeding half the length of sigma 1 were found to trimerize post-translationally. This trimerization did not require ATP since it proceeded normally in the presence of apyrase. In contrast, formation of stable FL sigma 1 trimers was inhibited by apyrase treatment. Collectively, our data suggest that assembly of nascent sigma 1 chains at the N-terminus is intrinsically ATP independent, and occurs co-translationally when the ribosomes have traversed past the midpoint of the mRNA.