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The base following stop codons in mammalian genes is strongly biased, suggesting that it might be important for the termination event. This proposal has been tested experimentally both in vivo by using the human type I iodothyronine deiodinase mRNA and the recoding event at the internal UGA codon and in vitro by measuring the ability of each of the 12 possible 4-base stop signals to direct the eukaryotic polypeptide release factor to release a model peptide, formylmethionine, from the ribosome. The internal UGA in the deiodinase mRNA is used as a codon for incorporation of selenocysteine into the protein. Changing the base following this UGA codon affected the ratio of termination to selenocysteine incorporation in vivo at this codon: 1:3 (C or U) and 3:1 (A or G). These UGAN sequences have the same order of efficiency of termination as was found with the in vitro termination assay (4th base: A G> > C U). The efficiency of in vitro termination varied in the same manner over a 70-fold range for the UAAN series and over an 8-fold range for the UGAN and UAGN series. There is a correlation between the strength of the signals and how frequently they occur at natural termination sites. Together these data suggest that the base following the stop codon influences translational termination efficiency as part of a larger termination signal in the expression of mammalian genes.

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We investigated the mechanisms by which previous "priming" activation of group I metabotropic glutamate receptors (mGluRs) facilitates the persistence of long-term potentiation (LTP) in area CA1 of rat hippocampal slices. Priming of LTP was elicited by either pharmacological or synaptic activation of mGluRs before a weak tetanic stimulus that normally produced only a rapidly decaying phase of LTP that did not involve protein synthesis or mGluRs. Pharmacological priming of LTP persistence by a selective group I mGluR agonist was blocked by an inhibitor of group I mGluRs and by inhibitors of translation, but not by a transcriptional inhibitor. The same mGluR agonist increased (35)S-methionine incorporation into slice proteins. LTP could also be facilitated using a synaptic stimulation priming protocol, and this effect was similarly blocked by group I mGluR and protein synthesis inhibitors. Furthermore, using a two-pathway protocol, the synaptic priming of LTP was found to be input-specific. To test for the contribution of group I mGluRs and protein synthesis to LTP in nonprimed slices, a longer duration control tetanization protocol was used to elicit a more slowly decaying form of LTP than did the weak tetanus used in the previous experiments. The persistence of the LTP induced by this stronger tetanus was dependent on mGluR activation and protein synthesis but not on transcription. Together, these results suggest that mGluRs couple to nearby protein synthesis machinery to homosynaptically regulate an intermediate phase of LTP dependent on new proteins made from pre-existing mRNA.

SummaryHuman mitochondria contain their own genome, encoding 13 polypeptides that are synthesized within the organelle. The molecular processes that govern and facilitate this mitochondrial translation remain unclear. Many key factors have yet to be characterized—for example, those required for translation termination. All other systems have two classes of release factors that either promote codon-specific hydrolysis of peptidyl-tRNA (class I) or lack specificity but stimulate the dissociation of class I factors from the ribosome (class II). One human mitochondrial protein has been previously identified in silico as a putative member of the class I release factors. Although we could not confirm the function of this factor, we report the identification of a different mitochondrial protein, mtRF1a, that is capable in vitro and in vivo of terminating translation at UAA/UAG codons. Further, mtRF1a depletion in HeLa cells led to compromised growth in galactose and increased production of reactive oxygen species.

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