Two classes of translational control RNA: their role in the regulation of protein synthesis.

Abstract

Two classes of translational control RNA (tcRNA) have been isolated from embryonic chick muscle. One of these classes, the tcRNA isolated from messenger ribonucleoprotein particles (mRNP-tcRNA), is effective in inhibiting the translation of mRNP-mRNA while having little if any effect on polysomal mRNA. The other class, polysome-tcRNA, has no effect on mRNP-mRNA while it stimulates the translation of polysomal mRNA. Translational control RNA (tcRNA) has been isolated from initiation factor preparations and has been demonstrated to specifically inhibit the translation of heterologous mRNAs (3). The inhibition by muscle tcRNA results from the inability of the globin mRNA to bind to ribosomes during the formation of the initiation complex when tcRNA is added to the cell-free system (6). Due to the specific nature of this inhibition, it is likely that the tcRNA interacts directly with mRNA. However, the involvement of initiation factors in this tcRNA-mediated inhibition cannot be excluded.In order to determine if tcRNA is effective in the regulation of protein synthesis within the cell from which it is derived, we have isolated mRNA from both polysomes and messenger ribonucleoprotein particles (mRNPs) of embryonic chick muscles and tested the ability of muscle tcRNA to inhibit the translation of these mRNAs. We report here that tcRNA may be obtained from either the polysomal or mRNP sources. Both tcRNA preparations inhibit globin synthesis, while only the tcRNA obtained from mRNPs is effective in inhibiting the Abbreviations: tcRNA, translational control RNA; mINP, messenger ribonucleoprotein particles. * Permanent address: Tygerberg Hospital, Capetown, South Africa. 1523translation of muscle mRNAs; and, in this case, only the translation of mRNP-mRNA is inhibited. It therefore is likely that at least two classes of tcRNA are present in muscle. It was of particular interest that the tcRNA effective in inhibiting muscle mRNP-mRNA translation contains a poly(U) tract that is absent in the polysomal class of tcRNA. From these results, a model is proposed to explain the manner by which tcRNA specifically regulates the utilization of cytoplasmic mRNAs. MATERIALS AND METHODSThe preparation of rabbit reticulocyte lysates and the conditions for cell-free protein synthesis were as described (3) except that 50 AM hemin and 15 jAg of muscle initiation factor 3 (IF3) were present in the reaction mixture, which had a final volume of 0.1 ml. Incubations were for 30 min at 300. Twentyfive one hundredth nmol each of 20 amino acids containing 8 jACi of 3H-labeled amino-acid mixture (New England Nuclear Corp.) was added to each incubation mixture. Where indicated, 2 ,g of tcRNA and 15 ug of poly(A)-containing mRNA from muscle were added to the cell-free, amino-acid incorporating systems.The mRNA and tcRNA preparations were prepared from 14-day embryonic chick leg muscle in the following manner. Approximately 45-50 g of muscle was gently homogenized in 0.25 M KC1, 5 MM M\gCI2, 20 mM Tris-HCl (pH 7.4) as described (7). The 10,00...

Regeneration of renal proximal tubules after mercuric chloride injury is accompanied by increased binding of aminoacyl-transfer ribonucleic acid

“…For regenerating liver and liver during microsomalenzyme induction, evidence has been obtained for a decrease in the activity of an inhibitor of aminoacyltRNA binding (Goodchild & Nicholls, 1976;Tominaga et al, 1975), as well as an increase in the activity of elongation factor 1 (Cappon & Nicholls, 1974a (Petryshyn & Nicholls, 1976). Further, the absence of a differential effect on initiation suggests that changes in an inhibitor affecting initiation, such as reported for tissues other than kidney, cannot account for the results reported here (Gross & Rabinowitz, 1972;Lodish & Desalu, 1973;Levin et al, 1975;Bester et al, 1975). The increased incorporation with the supernatant fraction of regenerating kidneys could not be attributed to the amount of mRNA in the preparation, since the increase was present when excess of poly(U) directed the peptide formation as well as when endogenous mRNA in the control-liver ribosome preparation directed incorporation.…”

Non-polyadenylated 22 S ribonucleoprotein particle is insensitive to translational inhibitor RNA of cryptobiotic gastrulae of Artemia salina

“…These mRNPs as well as mRNAs isolated thereof are inactive as templates in a cell-free translational system, owing to their bound translational inhibitor RNA, which can be dissociated in vitro from poly(A) mRNP or mRNA by poly(U)-Sepharose affinity chromatography in the presence of 10 mM EDTA (2). Similar translational inhibitor RNAs associated with mRNPs have also been detected in the embryonic chicken muscle (3,4), duck erythroblasts (5), rat connective tissue (6) as well as in Artemia salina embryos (7).…”

Novel mechanism for translational control in regulation of ferritin synthesis by iron.

“…Despite the considerable variation in the template activity of the post-ribosomal mRNA preparations (normal rat livers: 189,000-695,000 cpm; iron-treated rat liver: 261,000-554,000 cpm), very little variation was observed in the ratio: incorporation of radioactivity into ferritin/incorporation of radioactivity into total protein (Table 2, last column) within the two types of post-ribosomal mRNA preparations (normal rat liver: 0.39-0.56; iron-treated rat liver: 0.11-0.13). Bester et al (43) have demonstrated the presence of translational control RNA (tcRNA) in the post-ribosomal supernatant from embryonic chick muscle and have shown that it inhibits the translation of the corresponding messenger ribonucleoprotein (mRNP). We have preliminary evidence that some of our post-ribosomal mRNA preparations contained variable amounts of translational control RNA which would explain the observed differences in their activities in the wheat germ system.…”