Skip to main content
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1988 Apr;8(4):1748–1755. doi: 10.1128/mcb.8.4.1748

Haploid-specific transcription of protamine-myc and protamine-T-antigen fusion genes in transgenic mice.

T A Stewart 1, N B Hecht 1, P G Hollingshead 1, P A Johnson 1, J A Leong 1, S L Pitts 1
PMCID: PMC363335  PMID: 3380096

Abstract

The protamines are small, basic, arginine-rich proteins synthesized postmeiotically in the testes. Analysis of the regulation of synthesis of the protamine mRNA and protein is restricted by the difficulty in culturing and manipulating the cells in which transcription and translation occur. To avoid these problems, we have produced transgenic mice carrying fusion genes in which sequences 5' to the mouse protamine-2 gene have been linked to exons 2 and 3 of the mouse c-myc gene and, separately, to the simian virus 40 (SV40) early region. We show here that the prot.myc gene is correctly regulated; transcription is detected only in the round spermatids. In one family of transgenic mice carrying the 5' protamine-SV40 T-antigen fusion gene, SV40 early-region mRNA accumulated to the highest level in the testes but was also detected in the thymuses, brains, hearts, and preputial glands of the animals. Although we have demonstrated specific transcription of these fusion genes in the round spermatids, we were not able to detect the SV40 T-antigen protein.

Full text

PDF
1748

Images in this article

Selected References

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

  1. Adams J. M., Harris A. W., Pinkert C. A., Corcoran L. M., Alexander W. S., Cory S., Palmiter R. D., Brinster R. L. The c-myc oncogene driven by immunoglobulin enhancers induces lymphoid malignancy in transgenic mice. Nature. 1985 Dec 12;318(6046):533–538. doi: 10.1038/318533a0. [DOI] [PubMed] [Google Scholar]
  2. Ammer H., Henschen A., Lee C. H. Isolation and amino-acid sequence analysis of human sperm protamines P1 and P2. Occurrence of two forms of protamine P2. Biol Chem Hoppe Seyler. 1986 Jun;367(6):515–522. doi: 10.1515/bchm3.1986.367.1.515. [DOI] [PubMed] [Google Scholar]
  3. Balhorn R., Weston S., Thomas C., Wyrobek A. J. DNA packaging in mouse spermatids. Synthesis of protamine variants and four transition proteins. Exp Cell Res. 1984 Feb;150(2):298–308. doi: 10.1016/0014-4827(84)90572-x. [DOI] [PubMed] [Google Scholar]
  4. Battey J., Moulding C., Taub R., Murphy W., Stewart T., Potter H., Lenoir G., Leder P. The human c-myc oncogene: structural consequences of translocation into the IgH locus in Burkitt lymphoma. Cell. 1983 Oct;34(3):779–787. doi: 10.1016/0092-8674(83)90534-2. [DOI] [PubMed] [Google Scholar]
  5. Baty D., Barrera-Saldana H. A., Everett R. D., Vigneron M., Chambon P. Mutational dissection of the 21 bp repeat region of the SV40 early promoter reveals that it contains overlapping elements of the early-early and late-early promoters. Nucleic Acids Res. 1984 Jan 25;12(2):915–932. doi: 10.1093/nar/12.2.915. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Bellvé A. R., Anderson E., Hanley Bowdoin L. Synthesis and amino acid composition of basic proteins in mammalian sperm nuclei. Dev Biol. 1975 Dec;47(2):349–365. doi: 10.1016/0012-1606(75)90289-4. [DOI] [PubMed] [Google Scholar]
  7. Brinster R. L., Chen H. Y., Messing A., van Dyke T., Levine A. J., Palmiter R. D. Transgenic mice harboring SV40 T-antigen genes develop characteristic brain tumors. Cell. 1984 Jun;37(2):367–379. doi: 10.1016/0092-8674(84)90367-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Chirgwin J. M., Przybyla A. E., MacDonald R. J., Rutter W. J. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry. 1979 Nov 27;18(24):5294–5299. doi: 10.1021/bi00591a005. [DOI] [PubMed] [Google Scholar]
  9. Coelingh J. P., Monfoort C. H., Rozijn T. H., Leuven J. A., Schiphof R., Steyn-Parvé E. P., Braunitzer G., Schrank B., Ruhfus A. The complete amino acid sequence of the basic nuclear protein of bull spermatozoa. Biochim Biophys Acta. 1972 Nov 28;285(1):1–14. doi: 10.1016/0005-2795(72)90174-2. [DOI] [PubMed] [Google Scholar]
  10. Cohen G. L., Wright P. J., DeLucia A. L., Lewton B. A., Anderson M. E., Tegtmeyer P. Critical spatial requirement within the origin of simian virus 40 DNA replication. J Virol. 1984 Jul;51(1):91–96. doi: 10.1128/jvi.51.1.91-96.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Dani C., Blanchard J. M., Piechaczyk M., El Sabouty S., Marty L., Jeanteur P. Extreme instability of myc mRNA in normal and transformed human cells. Proc Natl Acad Sci U S A. 1984 Nov;81(22):7046–7050. doi: 10.1073/pnas.81.22.7046. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Eick D., Piechaczyk M., Henglein B., Blanchard J. M., Traub B., Kofler E., Wiest S., Lenoir G. M., Bornkamm G. W. Aberrant c-myc RNAs of Burkitt's lymphoma cells have longer half-lives. EMBO J. 1985 Dec 30;4(13B):3717–3725. doi: 10.1002/j.1460-2075.1985.tb04140.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Gluzman Y. SV40-transformed simian cells support the replication of early SV40 mutants. Cell. 1981 Jan;23(1):175–182. doi: 10.1016/0092-8674(81)90282-8. [DOI] [PubMed] [Google Scholar]
  14. Hanahan D. Heritable formation of pancreatic beta-cell tumours in transgenic mice expressing recombinant insulin/simian virus 40 oncogenes. Nature. 1985 May 9;315(6015):115–122. doi: 10.1038/315115a0. [DOI] [PubMed] [Google Scholar]
  15. Hecht N. B., Kleene K. C., Yelick P. C., Johnson P. A., Pravtcheva D. D., Ruddle F. H. Mapping of haploid expressed genes: genes for both mouse protamines are located on chromosome 16. Somat Cell Mol Genet. 1986 Mar;12(2):203–208. doi: 10.1007/BF01560667. [DOI] [PubMed] [Google Scholar]
  16. Kleene K. C., Distel R. J., Hecht N. B. Nucleotide sequence of a cDNA clone encoding mouse protamine 1. Biochemistry. 1985 Jan 29;24(3):719–722. doi: 10.1021/bi00324a027. [DOI] [PubMed] [Google Scholar]
  17. Kleene K. C., Distel R. J., Hecht N. B. Translational regulation and deadenylation of a protamine mRNA during spermiogenesis in the mouse. Dev Biol. 1984 Sep;105(1):71–79. doi: 10.1016/0012-1606(84)90262-8. [DOI] [PubMed] [Google Scholar]
  18. Kleene K. C., Distel R. J., Hecht N. B. cDNA clones encoding cytoplasmic poly(A)+ RNAs which first appear at detectable levels in haploid phases of spermatogenesis in the mouse. Dev Biol. 1983 Aug;98(2):455–464. doi: 10.1016/0012-1606(83)90375-5. [DOI] [PubMed] [Google Scholar]
  19. Kolk A. H., Samuel T. Isolation, chemical and immunological characterization of two strongly basic nuclear proteins from human spermatozoa. Biochim Biophys Acta. 1975 Jun 26;393(2):307–319. doi: 10.1016/0005-2795(75)90057-4. [DOI] [PubMed] [Google Scholar]
  20. Krawetz S. A., Connor W., Dixon G. H. Cloning of bovine P1 protamine cDNA and the evolution of vertebrate P1 protamines. DNA. 1987 Feb;6(1):47–57. doi: 10.1089/dna.1987.6.47. [DOI] [PubMed] [Google Scholar]
  21. Lee C. H., Mansouri A., Hecht W., Hecht N. B., Engel W. Nucleotide sequence of a bovine protamine cDNA. Biol Chem Hoppe Seyler. 1987 Feb;368(2):131–135. doi: 10.1515/bchm3.1987.368.1.131. [DOI] [PubMed] [Google Scholar]
  22. Mayer J. F., Jr, Chang T. S., Zirkin B. R. Spermatogenesis in the mouse 2. Amino acid incorporation into basic nucleoproteins of mouse spermatids and spermatozoa. Biol Reprod. 1981 Dec;25(5):1041–1051. doi: 10.1095/biolreprod25.5.1041. [DOI] [PubMed] [Google Scholar]
  23. Mazrimas J. A., Corzett M., Campos C., Balhorn R. A corrected primary sequence for bull protamine. Biochim Biophys Acta. 1986 Jul 25;872(1-2):11–15. doi: 10.1016/0167-4838(86)90141-x. [DOI] [PubMed] [Google Scholar]
  24. McKay D. J., Renaux B. S., Dixon G. H. Human sperm protamines. Amino-acid sequences of two forms of protamine P2. Eur J Biochem. 1986 Apr 1;156(1):5–8. doi: 10.1111/j.1432-1033.1986.tb09540.x. [DOI] [PubMed] [Google Scholar]
  25. McKay D. J., Renaux B. S., Dixon G. H. The amino acid sequence of human sperm protamine P1. Biosci Rep. 1985 May;5(5):383–391. doi: 10.1007/BF01116555. [DOI] [PubMed] [Google Scholar]
  26. Palmiter R. D., Brinster R. L. Germ-line transformation of mice. Annu Rev Genet. 1986;20:465–499. doi: 10.1146/annurev.ge.20.120186.002341. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Peschon J. J., Behringer R. R., Brinster R. L., Palmiter R. D. Spermatid-specific expression of protamine 1 in transgenic mice. Proc Natl Acad Sci U S A. 1987 Aug;84(15):5316–5319. doi: 10.1073/pnas.84.15.5316. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Piechaczyk M., Yang J. Q., Blanchard J. M., Jeanteur P., Marcu K. B. Posttranscriptional mechanisms are responsible for accumulation of truncated c-myc RNAs in murine plasma cell tumors. Cell. 1985 Sep;42(2):589–597. doi: 10.1016/0092-8674(85)90116-3. [DOI] [PubMed] [Google Scholar]
  29. Robbins P. D., Rio D. C., Botchan M. R. trans Activation of the simian virus 40 enhancer. Mol Cell Biol. 1986 Apr;6(4):1283–1295. doi: 10.1128/mcb.6.4.1283. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Sautiere P., Belaiche D., Martinage A., Loir M. Primary structure of the ram (Ovis aries) protamine. Eur J Biochem. 1984 Oct 1;144(1):121–125. doi: 10.1111/j.1432-1033.1984.tb08439.x. [DOI] [PubMed] [Google Scholar]
  31. Stewart T. A., Bellvé A. R., Leder P. Transcription and promoter usage of the myc gene in normal somatic and spermatogenic cells. Science. 1984 Nov 9;226(4675):707–710. doi: 10.1126/science.6494906. [DOI] [PubMed] [Google Scholar]
  32. Stewart T. A., Pattengale P. K., Leder P. Spontaneous mammary adenocarcinomas in transgenic mice that carry and express MTV/myc fusion genes. Cell. 1984 Oct;38(3):627–637. doi: 10.1016/0092-8674(84)90257-5. [DOI] [PubMed] [Google Scholar]
  33. Wilcox J. N., Roberts J. L., Chronwall B. M., Bishop J. F., O'Donohue T. Localization of proopiomelanocortin mRNA in functional subsets of neurons defined by their axonal projections. J Neurosci Res. 1986;16(1):89–96. doi: 10.1002/jnr.490160109. [DOI] [PubMed] [Google Scholar]
  34. Yelick P. C., Balhorn R., Johnson P. A., Corzett M., Mazrimas J. A., Kleene K. C., Hecht N. B. Mouse protamine 2 is synthesized as a precursor whereas mouse protamine 1 is not. Mol Cell Biol. 1987 Jun;7(6):2173–2179. doi: 10.1128/mcb.7.6.2173. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Molecular and Cellular Biology are provided here courtesy of Taylor & Francis

RESOURCES