Differential stimulation of capped mRNA translation in vitro by cap binding protein

Abstract

The cap structure, m⁷GpppN, which is present at the 5′ end of most eukaryotic mRNAs, facilitates the initiation of protein synthesis¹. Rabbit reticulocytes and mouse L and ascites cells have been shown to contain a protein (molecular weight (MW) 24,000) that specifically recognizes the cap of reovirus mRNA and several other capped mRNAs². That protein partially co-purifies with eIF-3 and eIF-4B (refs 2, 3), two eukaryotic initiation factors involved in mRNA binding to ribosomes⁴. We have isolated essentially homogeneous 24,000-MW polypeptide which stimulated translation of capped mRNAs in vitro⁵. Consistent with those findings, we now report that among the many protein synthesis factors tested, preparations of eIF-3 and eIF-4B that also contained the 24,000-MW cap binding protein (CBP) markedly increased translation in HeLa cell extracts of capped mRNAs but not of naturally uncapped viral messengers. Moreover, removal of the 24,000-MW protein diminished the ability of eIF-3 to stimulate translation of capped mRNA. Purified CBP, which had no effect on uncapped satellite tobacco necrosis virus (STNV) RNA, stimulated translation of this RNA after addition of a cap. The protein also relieved translational competition between capped and naturally uncapped mRNAs in favour of the capped species. These findings support the hypothesis that the CBP participates in the regulation of eukaryotic mRNA translation in normal and virus-infected cells⁶.