COOPERATION OF THE 5'-UNTRANSLATED AND 3'-UNTRANSLATED REGIONS OF ORNITHINE DECARBOXYLASE MESSENGER-RNA AND INHIBITORY ROLE OF ITS 3'-UNTRANSLATED REGION IN REGULATING THE TRANSLATIONAL EFFICIENCY OF HYBRID RNA SPECIES VIA CELLULAR FACTOR(S)

The 5' untranslated region (UTR) has an inhibitory role in the transla tability of ornithine decarboxylase (ODC) mRNA and of hybrid mRNA spec ies, whereas the ODC 3' UTR causes a partial release of this inhibitio n. We designed experiments to explore whether the co-operation between ODC 5' UTR and 3' UTR in the translational regulation is due to a dir ect interaction of those sequences or whether it is mediated by their interaction with cellular factor(s). We stably transfected Chinese ham ster ovary (CHO)-K1 cells and transiently transfected COS-I cells with expression vectors carrying different chimaeric DNAs having the lucif erase (LUC) coding sequence as reporter gene, the ODC 5' UTR or the OD C 3' UTR, or both, in the appropriate positions. We compared the resul ts obtained by assaying the LUC activities of both transfected cell li nes with each chimaeric DNA with those observed by translating the hyb rid RNAs in a translation system in vitro. When the ODC 3' UTR was pre sent, we observed a partial release of the translation inhibition owin g to the ODC 5' UTR only in vivo. The releasing effect was restored in vitro by the addition of cytoplasmic extracts from wild-type CHO-K1 o r COS-I cells, prepared 2 and 8 h after their release from serum starv ation. We also observed a partial inhibition of the translatability of the hybrid RNA owing to the presence of the ODC 3' UTR itself; the tr anslational efficiency could be rescued by cell extract from 8 h serum -stimulated cells. The co-operation between the ODC-UTRs might be medi ated by factors expressed by cells during particular phases of the cel l cycle. Excess copies of the ODC-UTRs, expressed in trans, could comp ete in binding limited amounts of such regulatory factors and remove t hem from interaction with the endogenous ODC mRNA. This phenomenon sho uld be reflected by modifications of the kinetics of ODC and/or LUC ac tivities during serum stimulation. The overexpression of the ODC 3' UT R determined an increase in both endogenous ODC activity and LUC activ ity. Moreover, in the transfectants expressing the hybrid RNA species bearing the ODC 3' UTR the basal ODC activity is higher than that obse rved in control cells. We suggest that excess copies of the ODC 3' UTR mis-regulate the endogenous ODC translatability, probably by tying up regulatory molecules expressed by cells in limited amounts and seques tering them from the ODC mRNA species they should interact with.