Stimulation of polypeptide initiation in vitro after protein synthesis inhibition in vivo in HeLa cells

Crude cytoplasmic extracts prepared from HeLa cells actively incorporate amino acids but show little initiation of new peptides (as seen by labeling of N-terminal amino acids). In contrast, extracts prepared from cells subjected to prior inhibition of protein synthesis show a significant amount of polypeptide initiation indicated by formation of peptides with radioactive N-terminal methionine. The same result was obtained whether prior inhibition occurred with cycloheximide or by starvation for an essential amino acid. Cellular response to suppression of protein synthesis appears to be mediated through production of RNA, since it is inhibited by actinomycin but appears in the presence of cycloheximide. The crude extracts continue initiating new polypeptides for at least 10 min in vitro. It is postulated that enhancement of in vitro initiation described here is related to the apparent stimulation of initiation of translation seen in vivo.