Skip to main content
NCBI home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1985 Mar;82(5):1341–1345. doi: 10.1073/pnas.82.5.1341

Transfer RNA is an essential component of the ubiquitin- and ATP-dependent proteolytic system.

A Ciechanover, S L Wolin, J A Steitz, H F Lodish
PMCID: PMC397256  PMID: 2983338

Abstract

Protein degradation via the nonlysosomal ATP-dependent pathway in rabbit reticulocytes involves a number of components. In the initial event, ubiquitin, an abundant 76-residue polypeptide, becomes covalently linked to the protein substrate in an ATP-requiring reaction. Once marked in this way, the conjugated protein is proteolyzed in a reaction that also requires ATP. Ubiquitin-marking appears to be important to the progression of cells from one stage to another of the cell cycle; it may also be involved in gene activation. Here we show that tRNA is another essential component of the system. Ribonucleases strongly inhibit the ubiquitin- and ATP-dependent degradation of 125I-labeled bovine serum albumin in the reticulocyte system in vitro. RNAs extracted from fractions of the reticulocyte extract or from mouse cells restore proteolytic activity. When the RNA is fractionated by gel electrophoresis, only the tRNA fraction is active in restoring proteolysis. Furthermore, pure mouse tRNAHis, isolated by immunoprecipitation with patient autoimmune sera, restores the proteolytic activity. The possibility that the level of uncharged tRNA in mammalian cells regulates the ubiquitin- and ATP-dependent proteolytic system is discussed.

Full text

PDF
1342

Images in this article

1343Image
on p.1343
1344Image
on p.1344
1344Image
on p.1344

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Alwine J. C., Kemp D. J., Parker B. A., Reiser J., Renart J., Stark G. R., Wahl G. M. Detection of specific RNAs or specific fragments of DNA by fractionation in gels and transfer to diazobenzyloxymethyl paper. Methods Enzymol. 1979;68:220–242. doi: 10.1016/0076-6879(79)68017-5. [DOI] [PubMed] [Google Scholar]
  2. Busch H., Goldknopf I. L. Ubiquitin - protein conjugates. Mol Cell Biochem. 1981 Nov 13;40(3):173–187. doi: 10.1007/BF00224611. [DOI] [PubMed] [Google Scholar]
  3. Cashel M. Regulation of bacterial ppGpp and pppGpp. Annu Rev Microbiol. 1975;29:301–318. doi: 10.1146/annurev.mi.29.100175.001505. [DOI] [PubMed] [Google Scholar]
  4. Ciechanover A., Finley D., Varshavsky A. The ubiquitin-mediated proteolytic pathway and mechanisms of energy-dependent intracellular protein degradation. J Cell Biochem. 1984;24(1):27–53. doi: 10.1002/jcb.240240104. [DOI] [PubMed] [Google Scholar]
  5. Ciechanover A., Finley D., Varshavsky A. Ubiquitin dependence of selective protein degradation demonstrated in the mammalian cell cycle mutant ts85. Cell. 1984 May;37(1):57–66. doi: 10.1016/0092-8674(84)90300-3. [DOI] [PubMed] [Google Scholar]
  6. Ciehanover A., Hod Y., Hershko A. A heat-stable polypeptide component of an ATP-dependent proteolytic system from reticulocytes. Biochem Biophys Res Commun. 1978 Apr 28;81(4):1100–1105. doi: 10.1016/0006-291x(78)91249-4. [DOI] [PubMed] [Google Scholar]
  7. Cuatrecasas P., Fuchs S., Anfinsen C. B. Catalytic properties and specificity of the extracellular nuclease of Staphylococcus aureus. J Biol Chem. 1967 Apr 10;242(7):1541–1547. [PubMed] [Google Scholar]
  8. Finley D., Ciechanover A., Varshavsky A. Thermolability of ubiquitin-activating enzyme from the mammalian cell cycle mutant ts85. Cell. 1984 May;37(1):43–55. doi: 10.1016/0092-8674(84)90299-x. [DOI] [PubMed] [Google Scholar]
  9. Goldberg A. L., St John A. C. Intracellular protein degradation in mammalian and bacterial cells: Part 2. Annu Rev Biochem. 1976;45:747–803. doi: 10.1146/annurev.bi.45.070176.003531. [DOI] [PubMed] [Google Scholar]
  10. Han J. H., Harding J. D. Isolation and nucleotide sequence of a mouse histidine tRNA gene. Nucleic Acids Res. 1982 Aug 25;10(16):4891–4900. doi: 10.1093/nar/10.16.4891. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hardin J. A., Rahn D. R., Shen C., Lerner M. R., Wolin S. L., Rosa M. D., Steitz J. A. Antibodies from patients with connective tissue diseases bind specific subsets of cellular RNA-protein particles. J Clin Invest. 1982 Jul;70(1):141–147. doi: 10.1172/JCI110587. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Hershko A., Ciechanover A. Mechanisms of intracellular protein breakdown. Annu Rev Biochem. 1982;51:335–364. doi: 10.1146/annurev.bi.51.070182.002003. [DOI] [PubMed] [Google Scholar]
  13. Hershko A., Ciechanover A., Rose I. A. Resolution of the ATP-dependent proteolytic system from reticulocytes: a component that interacts with ATP. Proc Natl Acad Sci U S A. 1979 Jul;76(7):3107–3110. doi: 10.1073/pnas.76.7.3107. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Hershko A., Eytan E., Ciechanover A., Haas A. L. Immunochemical analysis of the turnover of ubiquitin-protein conjugates in intact cells. Relationship to the breakdown of abnormal proteins. J Biol Chem. 1982 Dec 10;257(23):13964–13970. [PubMed] [Google Scholar]
  15. Hershko A., Heller H., Elias S., Ciechanover A. Components of ubiquitin-protein ligase system. Resolution, affinity purification, and role in protein breakdown. J Biol Chem. 1983 Jul 10;258(13):8206–8214. [PubMed] [Google Scholar]
  16. Hershko A., Leshinsky E., Ganoth D., Heller H. ATP-dependent degradation of ubiquitin-protein conjugates. Proc Natl Acad Sci U S A. 1984 Mar;81(6):1619–1623. doi: 10.1073/pnas.81.6.1619. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Lerner M. R., Andrews N. C., Miller G., Steitz J. A. Two small RNAs encoded by Epstein-Barr virus and complexed with protein are precipitated by antibodies from patients with systemic lupus erythematosus. Proc Natl Acad Sci U S A. 1981 Feb;78(2):805–809. doi: 10.1073/pnas.78.2.805. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. MANDELSTAM J. The intracellular turnover of protein and nucleic acids and its role in biochemical differentiation. Bacteriol Rev. 1960 Sep;24(3):289–308. doi: 10.1128/br.24.3.289-308.1960. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Mathews M. B., Bernstein R. M. Myositis autoantibody inhibits histidyl-tRNA synthetase: a model for autoimmunity. Nature. 1983 Jul 14;304(5922):177–179. doi: 10.1038/304177a0. [DOI] [PubMed] [Google Scholar]
  20. Maxam A. M., Gilbert W. Sequencing end-labeled DNA with base-specific chemical cleavages. Methods Enzymol. 1980;65(1):499–560. doi: 10.1016/s0076-6879(80)65059-9. [DOI] [PubMed] [Google Scholar]
  21. Pösö A. R., Mortimore G. E. Requirement for alanine in the amino acid control of deprivation-induced protein degradation in liver. Proc Natl Acad Sci U S A. 1984 Jul;81(14):4270–4274. doi: 10.1073/pnas.81.14.4270. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Rosa M. D., Hendrick J. P., Jr, Lerner M. R., Steitz J. A., Reichlin M. A mammalian tRNAHis-containing antigen is recognized by the polymyositis-specific antibody anti-Jo-1. Nucleic Acids Res. 1983 Feb 11;11(3):853–870. doi: 10.1093/nar/11.3.853. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Scornik O. A. Faster protein degradation in response to decreases steady state levels of amino acylation of tRNAHis in Chinese hamster ovary cells. J Biol Chem. 1983 Jan 25;258(2):882–886. [PubMed] [Google Scholar]
  24. Scornik O. A., Ledbetter M. L., Malter J. S. Role of aminoacylation of histidyl-tRNA in the regulation of protein degradation in Chinese hamster ovary cells. J Biol Chem. 1980 Jul 10;255(13):6322–6329. [PubMed] [Google Scholar]
  25. Seglen P. O., Gordon P. B. Amino acid control of autophagic sequestration and protein degradation in isolated rat hepatocytes. J Cell Biol. 1984 Aug;99(2):435–444. doi: 10.1083/jcb.99.2.435. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Varshavsky A., Levinger L., Sundin O., Barsoum J., Ozkaynak E., Swerdlow P., Finley D. Cellular and SV40 chromatin: replication, segregation, ubiquitination, nuclease-hypersensitive sites, HMG-containing nucleosomes, and heterochromatin-specific protein. Cold Spring Harb Symp Quant Biol. 1983;47(Pt 1):511–528. doi: 10.1101/sqb.1983.047.01.061. [DOI] [PubMed] [Google Scholar]
  27. Walter P., Blobel G. Disassembly and reconstitution of signal recognition particle. Cell. 1983 Sep;34(2):525–533. doi: 10.1016/0092-8674(83)90385-9. [DOI] [PubMed] [Google Scholar]
  28. Wolin S. L., Steitz J. A. The Ro small cytoplasmic ribonucleoproteins: identification of the antigenic protein and its binding site on the Ro RNAs. Proc Natl Acad Sci U S A. 1984 Apr;81(7):1996–2000. doi: 10.1073/pnas.81.7.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES