Polypeptide chain initiation

Goss, D. J., Parkhurst, L. J., Mehta, H. B., Woodley, C. L., and Wahba, A.J. (1984). Studies on the role of eukaryotic nucleotide exchange factor in polypeptide chain initiation. 

Tzamarias, D., Roussou, I., and Thireos, G. (1989). Coupling of GCN4 mRNA translational activation with decreased rates of polypeptide chain initiation. Cell 57, 947-954. 

For most polypeptide chains initiation begins with one of the three initiation codons, most commonly the methionine codon AUG. When properly placed in an mRNA chain, GUG may also serve as a bacterial initiation codon. In such cases, it codes for methionine rather than for valine. Occasionally UUG, AUU, ACG, and perhaps other codons can initiate translation 288/289 This is less frequent in eukaryotes than in bacteria. The sequence of bases preceding the initiation codon must also be important for recognition of the "start" signal. 

High concentrations of hemin inhibit the transport of ALA synthase into the mitochondria, where one of the substrates, succinyl-CoA, is formed. Thus, heme synthesis is inhibited until enough globin is made to react with any heme already formed. Low concentrations, or the absence, of hemin is the signal that globin is not needed this protein (and, therefore, globin) synthesis is inhibited. In the absence of hemin, a protein kinase is activated this phosphorylates an initiation factor of (eukaryotic) protein synthesis, eIF-2, which then inhibits polypeptide chain initiation (Chap. 17) and hence inhibits globin synthesis. 

D. Scheme for steps so far detected in polypeptide chain initiation. 

In their model Euss and Koch (40) propose that after infection there is a non-specific reduction in the overall rate of polypeptide chain initiation. However, since each mEEA has its own intrinsic translational efficiency, not all mEEAs are affected to the same extent, and there is a differential reduction in the translation of individual host and viral mEEA species. In a somewhat different model, Carrasco and Smith (4I, 42) propose that upon contact of the virus with the cell membrane, a viral coat protein associates with the membrane, changing the normal monovalent-ion gradient. During the course of viral EEA translation into progeny coat protein, these proteins are also inserted into the membrane, and the membrane alterations continue. Eventually, sodiim leaks into the cell, there is an increase in the concentration of monovalent-ions inside the cells, and cellular but not viral protein chain initiation is inhibited. 

Interestingly, in reticulocyte lysates, the effect of ds RNA is similar to that resulting from heme deficiency (63), that is, polypeptide chain initiation is inhibited. (For an extensive discussion, see chapter 10 of this book).. The ds RNA appears to induce a protein kinase which phosphorylates initiation factor eIF-2 (63). Experiments in our laboratory (M. Jaye, unpublished data) have failed to detect the presence of any phosphorylated components in the crude initiation factor preparation after virus infection corresponding to those elicited in the presence of ds RNA. 

Initiation factor 3 (IF-3) is one of three protein cofactors necessary for polypeptide chain initiation. It is now known that such initiation proceeds via dissociation of 70 S ribosomes into 30 S and 50 S subunits. IF-3, by binding tightly to the 30 S subunit, strongly favors this dissociation, and also appears to have the additional function of directing the binding of 30 S ribosomal subunits to start signals in messenger RNA. Recently, a structurally detailed hypothetical mechanism account-... 

Clemens, M. J., 1976, Functional relationships between reticulocyte polypeptide chain initiation factor (IF-MP) and the translational inhibitor involved in regulation of protein synthesis by haemin, Eur. J. Biochem. 66 413. 

Clemens, M. J., Safer, B., Merrick, W. C., Anderson, W. F., and London, I. M., 1975, Inhibition of protein synthesis in rabbit reticulocyte lysates by double stranded RNA and oxidized glutathione Indirect mode of action on polypeptide chain initiation, Proc. Natl. Acad. Sci. USA 72 1286. 

Nuss, D. L., and Koch, G., 1976 , Variation in the relative synthesis of immunoglobulin G and non-immunoglobulin G proteins in cultured MFC-11 cells with changes in the overall rate of polypeptide chain initiation and elongation, J. Mol. Biol. 102 601. 

Pain, V. M., Lewis, J. A., Huvos, P., Henshaw, E. C., and Clemens, M. J., 1980, The effects of amino acid starvation on regulation of polypeptide chain initiation in Ehrlich ascites tumor cells, J. Biol. Chem. 255 1486. 

Siekierka, J., Mauser, L., and Ochoa, S., 1982, Mechanism of polypeptide chain initiation in eukaryotes and its control by phosphorylation of the a subunit of initiation factor 2, Proc. Natl. Acad. Sci. USA 79 2537. 

An essential part of the rationale presented above under (a) consists of the identification of altered products of mitochondrial protein synthesis as a result of the mutation. Although this is not a sufficient criterion for mitochondrial specification (since an altered protein might arise as a result of a mutational alteration in a component of the mitochondrial protein-synthesizing machinery, i.e., one of the mt r- or tRNAs, (as in poky Neu-rospora), it is a necessary one. We have therefore devoted considerable effort to demonstrating the capability of the mitochondria in the mutant to perform some form of protein synthesis. We did this by showing that they were capable of incorporating (1) labeled formate into formylmethionyl-puromycin as a measure of mitochondrial polypeptide-chain initiations (see also next section) (2) labeled leucine into mitochondrial membrane proteins in a reaction that is insensitive to cycloheximide (CHX), but sensitive to chloramphenicol (CAP) and (3) that continued exposure of cells to the latter led to their conversion to petite phenocopies, s5 of characteristic aspects of their phenotype, such as the presence of cytochrome b, and that this change was reversed on removal of the inhibitor (see also Table I). 

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