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The mRNA encoding the beta-subunit of the mitochondrial H(+)-ATP synthase (beta-F1-ATPase) is localized in an approx. 150 nm structure of the hepatocyte of mammals. In the present study, we have investigated the cis- and trans-acting factors involved in the generation of the ribonucleoprotein complex containing beta-F1-ATPase mRNA. Two cis-acting elements (beta1.2 and 3'beta) have been identified. The beta1.2 element is placed in the open reading frame, downstream of the region encoding the mitochondrial pre-sequence of the protein. The 3'beta element is the 3' non-translated region of the mRNA. Complex sets of proteins from the soluble and non-soluble fractions of the liver interact with the beta1.2 and 3'beta elements. A soluble p88, present also in reticulocyte lysate, displays binding specificity for both the cis-acting elements. Sedimentation and high-resolution in situ hybridization experiments showed that the structure containing the rat liver beta-F1-ATPase mRNA is found in fractions of high sucrose concentration, where large polysomes sediment. Treatment of liver extracts with EDTA promoted the mobilization of beta-F1-ATPase mRNA to fractions of lower sucrose concentration, suggesting that the structure containing beta-F1-ATPase mRNA is a large polysome. Finally, in vitro reconstitution experiments with reticulocyte lysate, using either the full-length, mutant or chimaeric versions of beta-F1-ATPase mRNA, reveal that the assembly of the beta-F1-ATPase mRNA polysome requires the co-operation of both the cis-acting mRNA determinants. The present study illustrates the existence of an intramolecular RNA cross-talking required for the association of the mRNA with the translational machinery.

Translation in vitro of the mammalian nucleus-encoded mRNA for the beta subunit of mitochondrial H(+)-ATP synthase (beta-mRNA) of oxidative phosphorylation is promoted by a 150 nt translational enhancer sequence in the 3'-untranslated region (3' UTR). Titration of the eukaryotic initiation factor eIF4E with cap analogue revealed that translation of capped beta-mRNA was pseudo-cap independent. The 3' UTR of beta-mRNA stimulates the translation of heterologous uncapped mRNA species, both when the 3' UTR is placed at the 3' end and at the 5' end of the transcripts. The 3' UTRs of the alpha subunit of mitochondrial H(+)-ATP synthase (alpha-F1-ATPase) and subunit IV of cytochrome c oxidase (COX IV) mRNA species, other nucleus-encoded transcripts of oxidative phosphorylation, do not have the same activity in translation as the 3' UTR of beta-mRNA. On dicistronic RNA species, the 3' UTR of beta-mRNA, and to a smaller extent that of COX IV mRNA, is able to promote the translation of the second cistron to a level comparable to the activity of internal ribosome entry sites (IRESs) described in picornavirus mRNA species. These results indicate that the 3' UTRs of certain mRNA species of oxidative phosphorylation have IRES-like functional activity. Riboprobes of the active 3' UTRs on dicistronic assays formed specific RNA-protein complexes when cross-linked by UV to proteins of the lysate, suggesting that cytoplasmic translation of the mRNA species bearing an active 3' UTR is assisted by specific RNA-protein interactions.