CAP-DEPENDENT AND CAP-INDEPENDENT TRANSLATION BY INTERNAL INITIATION OF MESSENGER-RNAS IN CELL-EXTRACTS PREPARED FROM SACCHAROMYCES-CEREVISIAE

Translation extracts were prepared from various strains of Saccharomyc es cerevisiae. The translation of mRNA molecules in these extracts was cooperatively enhanced by the presence of 5'-terminal cap structures and 3'-terminal poly(A) sequences. These cooperative effects could not be observed in other translation systems such as those prepared from rabbit reticulocytes, wheat germ, and human HeLa cells. Because the ye ast translation system mimicked the effects of the cap structure and p oly(A) tail on translational efficiency seen in vivo, this system was used to study cap-dependent and cap-independent translation of viral a nd cellular mRNA molecules. Both the 5' noncoding regions of hepatitis C virus and those of coxsackievirus B1 conferred cap-independent tran slation to a reporter coding region during translation in the yeast ex tracts; thus, the yeast translational apparatus is capable of initiati ng cap-independent translation. Although the translation of most yeast mRNAs was cap dependent, the unusually long 5' noncoding regions of m RNAs encoding cellular transcription factors TFIID and HAP4 were shown to mediate cap-independent translation in these extracts. Furthermore , both TFIID and HAP4 5' noncoding regions mediated translation of a s econd cistron when placed into the intercistronic spacer region of a d icistronic mRNA, indicating that these leader sequences can initiate t ranslation by an internal ribosome binding mechanism in this in vitro translation system. This finding raises the possibility that an intern al translation initiation mechanism exists in yeast cells for regulate d translation of endogenous mRNAs.