Abstract
Crude cytoplasmic extracts prepared from HeLa cells actively incorporate amino acids but show little initiation of new peptides (as seen by labeling of N-terminal amino acids). In contrast, extracts prepared from cells subjected to prior inhibition of protein synthesis show a significant amount of polypeptide initiation indicated by formation of peptides with radioactive N-terminal methionine. The same result was obtained whether prior inhibition occurred with cycloheximide or by starvation for an essential amino acid. Cellular response to suppression of protein synthesis appears to be mediated through production of RNA, since it is inhibited by actinomycin but appears in the presence of cycloheximide. The crude extracts continue initiating new polypeptides for at least 10 min in vitro. It is postulated that enhancement of in vitro initiation described here is related to the apparent stimulation of initiation of translation seen in vivo.
Keywords: translation, regulation, cell response, Edman degradation
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Selected References
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- Berns A. J., Strous G. J., Bloemendal H. Heterologous in vitro synthesis of lens -crystallin polypeptide. Nat New Biol. 1972 Mar 1;236(61):7–9. doi: 10.1038/newbio236007a0. [DOI] [PubMed] [Google Scholar]
- Chatterjee N. K., Kerwar S. S., Weissbach H. Initiation of protein synthesis in HeLa cells. Proc Natl Acad Sci U S A. 1972 Jun;69(6):1375–1379. doi: 10.1073/pnas.69.6.1375. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Crystal R. G., Anderson W. F. Initiation of hemoglobin synthesis: comparison of model reactions that use artificial templates with those using natural messenger RNA. Proc Natl Acad Sci U S A. 1972 Mar;69(3):706–711. doi: 10.1073/pnas.69.3.706. [DOI] [PMC free article] [PubMed] [Google Scholar]
- EAGLE H. Amino acid metabolism in mammalian cell cultures. Science. 1959 Aug 21;130(3373):432–437. doi: 10.1126/science.130.3373.432. [DOI] [PubMed] [Google Scholar]
- Edman P. Sequence determination. Mol Biol Biochem Biophys. 1970;8:211–255. doi: 10.1007/978-3-662-12834-3_8. [DOI] [PubMed] [Google Scholar]
- Fan H., Penman S. Regulation of protein synthesis in mammalian cells. II. Inhibition of protein synthesis at the level of initiation during mitosis. J Mol Biol. 1970 Jun 28;50(3):655–670. doi: 10.1016/0022-2836(70)90091-4. [DOI] [PubMed] [Google Scholar]
- Greenberg J. R. High stability of messenger RNA in growing cultured cells. Nature. 1972 Nov 10;240(5376):102–104. doi: 10.1038/240102a0. [DOI] [PubMed] [Google Scholar]
- Gross K., Ruderman J., Jacobs-Lorena M., Baglioni C., Gross P. R. Cell-free synthesis of histones directed by messenger RNA from sea urchin embryos. Nat New Biol. 1973 Feb 28;241(113):272–274. doi: 10.1038/newbio241272a0. [DOI] [PubMed] [Google Scholar]
- Jackson R., Hunter T. Role of methionine in the initiation of haemoglobin synthesis. Nature. 1970 Aug 15;227(5259):672–676. doi: 10.1038/227672a0. [DOI] [PubMed] [Google Scholar]
- Jacobs-Lorena M., Baglioni C., Borun T. W. Translation of messenger RNA for histones from HeLa cells by a cell-free extract from mouse ascites tumor. Proc Natl Acad Sci U S A. 1972 Aug;69(8):2095–2099. doi: 10.1073/pnas.69.8.2095. [DOI] [PMC free article] [PubMed] [Google Scholar]
- KRUH J., SCHAPIRA G., LAREAU J., DREYFUS J. C. ACTIVATION DE LA SYNTH'ESE ACELLULAIRE DE L'H'EMOGLOBINE PAR L'ACIDE RIBONUCL'EIQUE. II. ACTION DE FRACTIONS D'ACIDE RIBONUCL'EIQUE DE R'ETICULOCYTES OBTENUES PAR CENTRIFUGATION EN GRADIENT DE SACCHAROSE. Biochim Biophys Acta. 1964 Aug 12;87:669–681. [PubMed] [Google Scholar]
- Kappen L. S., Suzuki H., Goldberg I. H. Inhibition of reticulocyte peptide-chain initiation by pactamycin: accumulation of inactive ribosomal initiation complexes. Proc Natl Acad Sci U S A. 1973 Jan;70(1):22–26. doi: 10.1073/pnas.70.1.22. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Lane C. D., Marbaix G., Gurdon J. B. Rabbit haemoglobin synthesis in frog cells: the translation of reticulocyte 9 s RNA in frog oocytes. J Mol Biol. 1971 Oct 14;61(1):73–91. doi: 10.1016/0022-2836(71)90207-5. [DOI] [PubMed] [Google Scholar]
- Legon S., Jackson R. J., Hunt T. Control of protein synthesis in reticulocyte lysates by haemin. Nat New Biol. 1973 Jan 31;241(109):150–152. doi: 10.1038/newbio241150a0. [DOI] [PubMed] [Google Scholar]
- Leibowitz R., Penman S. Regulation of protein synthesis in HeLa cells. 3. Inhibition during poliovirus infection. J Virol. 1971 Nov;8(5):661–668. doi: 10.1128/jvi.8.5.661-668.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Lockard R. E., Lingrel J. B. The synthesis of mouse hemoglobin beta-chains in a rabbit reticulocyte cell-free system programmed with mouse reticulocyte 9S RNA. Biochem Biophys Res Commun. 1969 Oct 8;37(2):204–212. doi: 10.1016/0006-291x(69)90720-7. [DOI] [PubMed] [Google Scholar]
- Lodish H. G., Housman D., Jacobsen M. Initiation of hemoglobin synthesis. Specific inhibition by antibiotics and bacteriophage ribonucleic acid. Biochemistry. 1971 Jun 8;10(12):2348–2356. doi: 10.1021/bi00788a027. [DOI] [PubMed] [Google Scholar]
- McCormick W., Penman S. Regulation of protein synthesis in HeLa cells: translation at elevated temperatures. J Mol Biol. 1969 Jan;39(2):315–333. doi: 10.1016/0022-2836(69)90320-9. [DOI] [PubMed] [Google Scholar]
- Means A. R., Comstock J. P., Rosenfeld G. C., O'Malley B. W. Ovalbumin messenger RNA of chick oviduct: partial characterization, estrogen dependence, and translation in vitro. Proc Natl Acad Sci U S A. 1972 May;69(5):1146–1150. doi: 10.1073/pnas.69.5.1146. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Rosenfeld G. C., Comstock J. P., Means A. R., O'Malley B. W. Estrogen-induced synthesis of ovalbumin messenger RNA and its translation in a cell-free system. Biochem Biophys Res Commun. 1972 Feb 25;46(4):1695–1703. doi: 10.1016/0006-291x(72)90805-4. [DOI] [PubMed] [Google Scholar]
- Singer R. H., Penman S. Stability of HeLa cell mRNA in actinomycin. Nature. 1972 Nov 10;240(5376):100–102. doi: 10.1038/240100a0. [DOI] [PubMed] [Google Scholar]
- Stewart-Blair M. L., Yanowitz I. S., Goldberg I. H. Inhibition of synthesis of new globin chains in reticulocyte lysates by pactamycin. Biochemistry. 1971 Nov;10(23):4198–4206. doi: 10.1021/bi00799a007. [DOI] [PubMed] [Google Scholar]
- Vaughan M. H., Jr, Pawlowski P. J., Forchhammer J. Regulation of protein synthesis initiation in HeLa cells deprived of single essential amino acids. Proc Natl Acad Sci U S A. 1971 Sep;68(9):2057–2061. doi: 10.1073/pnas.68.9.2057. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Weiner A. M., Platt T., Weber K. Amino-terminal sequence analysis of proteins purified on a nanomole scale by gel electrophoresis. J Biol Chem. 1972 May 25;247(10):3242–3251. [PubMed] [Google Scholar]
- van Venrooij W. J., Henshaw E. C., Hirsch C. A. Effects of deprival of glucose or individual amino acids on polyribosome distribution and rate of protein synthesis in cultured mammalian cells. Biochim Biophys Acta. 1972 Jan 18;259(1):127–137. doi: 10.1016/0005-2787(72)90480-7. [DOI] [PubMed] [Google Scholar]
