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. 1979 Aug;76(8):3809–3813. doi: 10.1073/pnas.76.8.3809

Identification of separate mRNAs coding for the alpha and beta subunits of thyrotropin.

N C Vamvakopoulos, I A Kourides
PMCID: PMC383924  PMID: 291041

Abstract

A 9S mRNA, purified from mouse thyrotropic pituitary tumors by sucrose density gradient centrifugation of poly(A)-enriched mRNA, directed the synthesis of only alpha and beta subunits of thyrotropin in the reticulocyte lysate translation system. Analysis of radioiodinated 9S mRNA, repurified by oligo(dT)-cellulose chromatography and sucrose gradient centrifugation, yielded two species of RNA on urea/polyacrylamide gel electrophoresis. The major RNA species contained 620 nucleotides, and the minor RNA species contained 560 nucleotides. Unlabeled 9S mRNA was further purified by urea/polyacrylamide gel electrophoresis; the mRNAs were separately eluted from slices of the gel containing material migrating with an apparent length of 620 and 560 nucleotides. Translation of these mRNAs in the reticulocyte lysate showed that the longer mRNA coded for the alpha subunit and the shorter mRNA coded for the beta subunit of mouse thyrotropin. Because more alpha than beta subunit of thyrotropin was consistently synthesized, unbalanced amounts of thyrotropin subunits appear to be synthesized by translation of unbalanced amounts of individual mRNAs. We have demonstrated that the synthesis of thyrotropin is directed by two separate mRNA molecules, each coding for a different subunit of the hormone.

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Selected References

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

  1. Aviv H., Leder P. Purification of biologically active globin messenger RNA by chromatography on oligothymidylic acid-cellulose. Proc Natl Acad Sci U S A. 1972 Jun;69(6):1408–1412. doi: 10.1073/pnas.69.6.1408. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Birken S., Fetherston J., Desmond J., Canfield R., Boime I. Partial amino acid sequence of the preprotein form of the alpha subunit of human choriogonadotropin and identification of the site of subsequent proteolytic cleavage. Biochem Biophys Res Commun. 1978 Dec 29;85(4):1247–1253. doi: 10.1016/0006-291x(78)91137-3. [DOI] [PubMed] [Google Scholar]
  3. Blackman M. R., Gershengorn M. C., Weintraub B. D. Excess production of free alpha subunits by mouse pituitary thyrotropic tumor cells in vitro. Endocrinology. 1978 Feb;102(2):499–508. doi: 10.1210/endo-102-2-499. [DOI] [PubMed] [Google Scholar]
  4. Blobel G., Dobberstein B. Transfer of proteins across membranes. I. Presence of proteolytically processed and unprocessed nascent immunoglobulin light chains on membrane-bound ribosomes of murine myeloma. J Cell Biol. 1975 Dec;67(3):835–851. doi: 10.1083/jcb.67.3.835. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Chin W. W., Habener J. F., Kieffer J. D., Maloof F. Cell-free translation of the messenger RNA coding for the alpha subunit of thyroid-stimulating hormone. J Biol Chem. 1978 Nov 25;253(22):7985–7988. [PubMed] [Google Scholar]
  6. Commerford S. L. Iodination of nucleic acids in vitro. Biochemistry. 1971 May 25;10(11):1993–2000. doi: 10.1021/bi00787a005. [DOI] [PubMed] [Google Scholar]
  7. Crystal R. G., Elson N. A., Anderson W. F. Initiation of globin synthesis: assays. Methods Enzymol. 1974;30:101–127. doi: 10.1016/0076-6879(74)30014-6. [DOI] [PubMed] [Google Scholar]
  8. Daniels-McQueen S., McWillians D., Birken S., Canfield R., Landefeld T., Boime I. Identification of mRNAs encoding the alpha and beta subunits of human choriogonadotropin. J Biol Chem. 1978 Oct 10;253(19):7109–7114. [PubMed] [Google Scholar]
  9. Favre A., Morel C., Scherrer K. The secondary structure and poly(A) content of globin messenger RNA as a pure RNA and in polyribosome-derived ribonucleoprotein complexes. Eur J Biochem. 1975 Sep 1;57(1):147–157. doi: 10.1111/j.1432-1033.1975.tb02285.x. [DOI] [PubMed] [Google Scholar]
  10. Franklin R. M. Purification and properties of the replicative intermediate of the RNA bacteriophage R17. Proc Natl Acad Sci U S A. 1966 Jun;55(6):1504–1511. doi: 10.1073/pnas.55.6.1504. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Furth J., Moy P., Hershman J. M., Ueda G. Thyrotropic tumor syndrome. A multiglandular disease induced by sustained deficiency of thyroid hormones. Arch Pathol. 1973 Oct;96(4):217–226. [PubMed] [Google Scholar]
  12. Gould H. J., Hamlyn P. H. The molecular weight of rabbit globin messenger RNA's. FEBS Lett. 1973 Mar 15;30(3):301–304. doi: 10.1016/0014-5793(73)80674-x. [DOI] [PubMed] [Google Scholar]
  13. Hagen C., McNeilly A. S. Identification of human luteinizing hormone, follicle-stimulating hormone, luteinizing hormone beta-subunit and gonadotrophin alpha-subunit in foetal and adult pituitary glands. J Endocrinol. 1975 Oct;67(1):49–57. doi: 10.1677/joe.0.0670049. [DOI] [PubMed] [Google Scholar]
  14. Jackson R. C., Blobel G. Post-translational cleavage of presecretory proteins with an extract of rough microsomes from dog pancreas containing signal peptidase activity. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5598–5602. doi: 10.1073/pnas.74.12.5598. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Kaplan S. L., Grumbach M. M., Aubert M. L. alpha and beta glycoprotein hormone subunits (hLH, hFSH, hCG) in the serum and pituitary of the human fetus. J Clin Endocrinol Metab. 1976 May;42(5):995–998. doi: 10.1210/jcem-42-5-995. [DOI] [PubMed] [Google Scholar]
  16. Kessler S. W. Rapid isolation of antigens from cells with a staphylococcal protein A-antibody adsorbent: parameters of the interaction of antibody-antigen complexes with protein A. J Immunol. 1975 Dec;115(6):1617–1624. [PubMed] [Google Scholar]
  17. Kourides I. A., Ridgway E. C., Weintraub B. D., Bigos S. T., Gershengorn M. C., Maloof F. Thyrotropin-induced hyperthyroidism: use of alpha and beta subunit levels to identify patients with pituitary tumors. J Clin Endocrinol Metab. 1977 Sep;45(3):534–543. doi: 10.1210/jcem-45-3-534. [DOI] [PubMed] [Google Scholar]
  18. Kourides I. A., Weintraub B. D., Ridgway E. C., Maloof F. Pituitary secretion of free alpha and beta subunit of human thyrotropin in patients with thyroid disorders. J Clin Endocrinol Metab. 1975 May;40(5):872–885. doi: 10.1210/jcem-40-5-872. [DOI] [PubMed] [Google Scholar]
  19. Kourides I. A., Weintraub B. D., Rosen S. W., Ridgway E. C., Kliman B., Maloof F. Secretion of alpha subunit of glycoprotein hormones by pituitary adenomas. J Clin Endocrinol Metab. 1976 Jul;43(1):97–106. doi: 10.1210/jcem-43-1-97. [DOI] [PubMed] [Google Scholar]
  20. Kourides I. A., Weintraub B. D. mRNA-directed biosynthesis of alpha subunit of thyrotropin: translation in cell-free and whole-cell systems. Proc Natl Acad Sci U S A. 1979 Jan;76(1):298–302. doi: 10.1073/pnas.76.1.298. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  22. Laskey R. A., Mills A. D. Quantitative film detection of 3H and 14C in polyacrylamide gels by fluorography. Eur J Biochem. 1975 Aug 15;56(2):335–341. doi: 10.1111/j.1432-1033.1975.tb02238.x. [DOI] [PubMed] [Google Scholar]
  23. Lockhard R. E., Rajbhandary U. L. Nucleotide sequences at the 5'termini of rabbit alpha and beta globin mRNA. Cell. 1976 Dec;9(4 Pt 2):747–760. doi: 10.1016/0092-8674(76)90138-0. [DOI] [PubMed] [Google Scholar]
  24. Maniatis T., Jeffrey A., van deSande H. Chain length determination of small double- and single-stranded DNA molecules by polyacrylamide gel electrophoresis. Biochemistry. 1975 Aug 26;14(17):3787–3794. doi: 10.1021/bi00688a010. [DOI] [PubMed] [Google Scholar]
  25. Maxam A. M., Gilbert W. A new method for sequencing DNA. Proc Natl Acad Sci U S A. 1977 Feb;74(2):560–564. doi: 10.1073/pnas.74.2.560. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Palacios R., Sullivan D., Summers N. M., Kiely M. L., Schimke R. T. Purification of ovalbumin messenger ribonucleic acid by specific immunoadsorption of ovalbumin-synthesizing polysomes and millipore partition of ribonucleic acid. J Biol Chem. 1973 Jan 25;248(2):540–548. [PubMed] [Google Scholar]
  27. Pelham H. R., Jackson R. J. An efficient mRNA-dependent translation system from reticulocyte lysates. Eur J Biochem. 1976 Aug 1;67(1):247–256. doi: 10.1111/j.1432-1033.1976.tb10656.x. [DOI] [PubMed] [Google Scholar]
  28. Pierce J. G. Eli Lilly lecture. The subunits of pituitary thyrotropin--their relationship to other glycoprotein hormones. Endocrinology. 1971 Dec;89(6):1331–1344. doi: 10.1210/endo-89-6-1331. [DOI] [PubMed] [Google Scholar]
  29. Pierce J. G., Liao T., Howard S. M., Shome B., Cornell J. S. Studies on the structure of thyrotropin: its relationship to luteinizing hormone. Recent Prog Horm Res. 1971;27:165–212. doi: 10.1016/b978-0-12-571127-2.50029-8. [DOI] [PubMed] [Google Scholar]
  30. Prentice L. G., Ryan R. J. LH and its subunits in human pituitary, serum and urine. J Clin Endocrinol Metab. 1975 Feb;40(2):303–312. doi: 10.1210/jcem-40-2-303. [DOI] [PubMed] [Google Scholar]
  31. Roberts B. E., Paterson B. M. Efficient translation of tobacco mosaic virus RNA and rabbit globin 9S RNA in a cell-free system from commercial wheat germ. Proc Natl Acad Sci U S A. 1973 Aug;70(8):2330–2334. doi: 10.1073/pnas.70.8.2330. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Rosen S. W., Weintraub B. D. Ectopic production of the isolated alpha subunit of the glycoprotein hormones. A quantitative marker in certain cases of cancer. N Engl J Med. 1974 Jun 27;290(26):1441–1447. doi: 10.1056/NEJM197406272902601. [DOI] [PubMed] [Google Scholar]
  33. Spielman L. L., Bancroft F. C. Pregrowth hormone: evidence for conversion to growth hormone during synthesis on membrane-bound polysomes. Endocrinology. 1977 Sep;101(3):651–658. doi: 10.1210/endo-101-3-651. [DOI] [PubMed] [Google Scholar]
  34. Tashjian A. H., Jr, Weintraub B. D., Barowsky N. J., Rabson A. S., Rosen S. W. Subunits of human chorionic gonadotropin: unbalanced synthesis and secretion by clonal cell strains derived from a bronchogenic carcinoma. Proc Natl Acad Sci U S A. 1973 May;70(5):1419–1422. doi: 10.1073/pnas.70.5.1419. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Vaitukaitis J. L., Ross G. T., Braunstein G. D., Rayford P. L. Gonadotropins and their subunits: basic and clinical studies. Recent Prog Horm Res. 1976;32:289–331. doi: 10.1016/b978-0-12-571132-6.50019-1. [DOI] [PubMed] [Google Scholar]
  36. Weintraub B. D., Rosen S. W. Ectopic production of the isolated beta subunit of human chorionic gonadotropin. J Clin Invest. 1973 Dec;52(12):3135–3142. doi: 10.1172/JCI107513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Weintraub B. D., Stannard B. S. Precursor-product relationships in the biosynthesis and secretion of thyrotropin and its subunits by mouse thyrotropic tumor cells. FEBS Lett. 1978 Aug 15;92(2):303–307. doi: 10.1016/0014-5793(78)80775-3. [DOI] [PubMed] [Google Scholar]
  38. Zasloff M., Ginder G. D., Felsenfeld G. A new method for the purification and identification of covalently closed circular DNA molcules. Nucleic Acids Res. 1978 Apr;5(4):1139–1152. doi: 10.1093/nar/5.4.1139. [DOI] [PMC free article] [PubMed] [Google Scholar]

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