Skip to main content
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1995 Jan;15(1):561–571. doi: 10.1128/mcb.15.1.561

Calspermin gene transcription is regulated by two cyclic AMP response elements contained in an alternative promoter in the calmodulin kinase IV gene.

Z Sun 1, P Sassone-Corsi 1, A R Means 1
PMCID: PMC232013  PMID: 7799965

Abstract

The transcript for the high-affinity Ca2+/calmodulin-binding protein calspermin is generated from the gene encoding Ca2+/calmodulin-dependent protein kinase IV only in postmeiotic germ cells during spermatogenesis. We demonstrate that this testis-specific calspermin transcript can be produced in heterologous cells by utilization of a promoter located in an intron of the calmodulin (CaM) kinase IV gene. Critical motifs within this promoter are two cyclic AMP response element (CRE)-like sequences located about -70 and -50 bp upstream of the transcriptional initiation site. Both CRE motifs are footprinted by the authentic testis-specific transcriptional activator CREM tau or by CREM tau present in adult testis nuclear extract. Whereas a 2.1-kb DNA fragment containing the calspermin promoter is inactive when transfected into NIH 3T3 cells, activity can be restored by cotransfection of CREM tau and protein kinase A or CaM kinase IV but not CaM kinase II alpha. Restoration of activity is greatly reduced by mutation of the two CRE motifs. Since CRE-like motifs have been identified in many genes uniquely expressed in postmeiotic germ cells, which contain abundant CREM tau protein, we suggest that CREM tau may function as one transcription factor responsible for the expression of postmeiotic germ cell-specific genes.

Full Text

The Full Text of this article is available as a PDF (514.8 KB).

Selected References

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

  1. Brokaw C. J., Nagayama S. M. Modulation of the asymmetry of sea urchin sperm flagellar bending by calmodulin. J Cell Biol. 1985 Jun;100(6):1875–1883. doi: 10.1083/jcb.100.6.1875. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bunick D., Johnson P. A., Johnson T. R., Hecht N. B. Transcription of the testis-specific mouse protamine 2 gene in a homologous in vitro transcription system. Proc Natl Acad Sci U S A. 1990 Feb;87(3):891–895. doi: 10.1073/pnas.87.3.891. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Cruzalegui F. H., Means A. R. Biochemical characterization of the multifunctional Ca2+/calmodulin-dependent protein kinase type IV expressed in insect cells. J Biol Chem. 1993 Dec 15;268(35):26171–26178. [PubMed] [Google Scholar]
  4. Dash P. K., Karl K. A., Colicos M. A., Prywes R., Kandel E. R. cAMP response element-binding protein is activated by Ca2+/calmodulin- as well as cAMP-dependent protein kinase. Proc Natl Acad Sci U S A. 1991 Jun 1;88(11):5061–5065. doi: 10.1073/pnas.88.11.5061. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Delmas V., Laoide B. M., Masquilier D., de Groot R. P., Foulkes N. S., Sassone-Corsi P. Alternative usage of initiation codons in mRNA encoding the cAMP-responsive-element modulator generates regulators with opposite functions. Proc Natl Acad Sci U S A. 1992 May 15;89(10):4226–4230. doi: 10.1073/pnas.89.10.4226. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Delmas V., van der Hoorn F., Mellström B., Jégou B., Sassone-Corsi P. Induction of CREM activator proteins in spermatids: down-stream targets and implications for haploid germ cell differentiation. Mol Endocrinol. 1993 Nov;7(11):1502–1514. doi: 10.1210/mend.7.11.8114765. [DOI] [PubMed] [Google Scholar]
  7. Epstein P. N., Overbeek P. A., Means A. R. Calmodulin-induced early-onset diabetes in transgenic mice. Cell. 1989 Sep 22;58(6):1067–1073. doi: 10.1016/0092-8674(89)90505-9. [DOI] [PubMed] [Google Scholar]
  8. Foulkes N. S., Borrelli E., Sassone-Corsi P. CREM gene: use of alternative DNA-binding domains generates multiple antagonists of cAMP-induced transcription. Cell. 1991 Feb 22;64(4):739–749. doi: 10.1016/0092-8674(91)90503-q. [DOI] [PubMed] [Google Scholar]
  9. Foulkes N. S., Mellström B., Benusiglio E., Sassone-Corsi P. Developmental switch of CREM function during spermatogenesis: from antagonist to activator. Nature. 1992 Jan 2;355(6355):80–84. doi: 10.1038/355080a0. [DOI] [PubMed] [Google Scholar]
  10. Foulkes N. S., Schlotter F., Pévet P., Sassone-Corsi P. Pituitary hormone FSH directs the CREM functional switch during spermatogenesis. Nature. 1993 Mar 18;362(6417):264–267. doi: 10.1038/362264a0. [DOI] [PubMed] [Google Scholar]
  11. Gorman C. M., Moffat L. F., Howard B. H. Recombinant genomes which express chloramphenicol acetyltransferase in mammalian cells. Mol Cell Biol. 1982 Sep;2(9):1044–1051. doi: 10.1128/mcb.2.9.1044. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. 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]
  13. Howard T., Balogh R., Overbeek P., Bernstein K. E. Sperm-specific expression of angiotensin-converting enzyme (ACE) is mediated by a 91-base-pair promoter containing a CRE-like element. Mol Cell Biol. 1993 Jan;13(1):18–27. doi: 10.1128/mcb.13.1.18. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Johnson P. A., Bunick D., Hecht N. B. Protein binding regions in the mouse and rat protamine-2 genes. Biol Reprod. 1991 Jan;44(1):127–134. doi: 10.1095/biolreprod44.1.127. [DOI] [PubMed] [Google Scholar]
  15. Jones D. A., Glod J., Wilson-Shaw D., Hahn W. E., Sikela J. M. cDNA sequence and differential expression of the mouse Ca2+/calmodulin-dependent protein kinase IV gene. FEBS Lett. 1991 Sep 2;289(1):105–109. doi: 10.1016/0014-5793(91)80919-t. [DOI] [PubMed] [Google Scholar]
  16. Koide Y., Ono T., Yamashita K. Calspermin: a 32K-dalton calmodulin-binding protein in central nervous system and male reproductive system; property and distribution. Cell Calcium. 1986 Dec;7(5-6):329–338. doi: 10.1016/0143-4160(86)90036-9. [DOI] [PubMed] [Google Scholar]
  17. Langford K. G., Shai S. Y., Howard T. E., Kovac M. J., Overbeek P. A., Bernstein K. E. Transgenic mice demonstrate a testis-specific promoter for angiotensin-converting enzyme. J Biol Chem. 1991 Aug 25;266(24):15559–15562. [PubMed] [Google Scholar]
  18. Laoide B. M., Foulkes N. S., Schlotter F., Sassone-Corsi P. The functional versatility of CREM is determined by its modular structure. EMBO J. 1993 Mar;12(3):1179–1191. doi: 10.1002/j.1460-2075.1993.tb05759.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Lefevre C., Imagawa M., Dana S., Grindlay J., Bodner M., Karin M. Tissue-specific expression of the human growth hormone gene is conferred in part by the binding of a specific trans-acting factor. EMBO J. 1987 Apr;6(4):971–981. doi: 10.1002/j.1460-2075.1987.tb04847.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Means A. R., Cruzalegui F. Differential gene expression from a single transcription unit during spermatogenesis. Recent Prog Horm Res. 1993;48:79–97. doi: 10.1016/b978-0-12-571148-7.50007-5. [DOI] [PubMed] [Google Scholar]
  21. Means A. R., Cruzalegui F., LeMagueresse B., Needleman D. S., Slaughter G. R., Ono T. A novel Ca2+/calmodulin-dependent protein kinase and a male germ cell-specific calmodulin-binding protein are derived from the same gene. Mol Cell Biol. 1991 Aug;11(8):3960–3971. doi: 10.1128/mcb.11.8.3960. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Mellström B., Naranjo J. R., Foulkes N. S., Lafarga M., Sassone-Corsi P. Transcriptional response to cAMP in brain: specific distribution and induction of CREM antagonists. Neuron. 1993 Apr;10(4):655–665. doi: 10.1016/0896-6273(93)90167-p. [DOI] [PubMed] [Google Scholar]
  23. Montminy M. R., Bilezikjian L. M. Binding of a nuclear protein to the cyclic-AMP response element of the somatostatin gene. Nature. 1987 Jul 9;328(6126):175–178. doi: 10.1038/328175a0. [DOI] [PubMed] [Google Scholar]
  24. Montminy M. R., Sevarino K. A., Wagner J. A., Mandel G., Goodman R. H. Identification of a cyclic-AMP-responsive element within the rat somatostatin gene. Proc Natl Acad Sci U S A. 1986 Sep;83(18):6682–6686. doi: 10.1073/pnas.83.18.6682. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Ohmstede C. A., Bland M. M., Merrill B. M., Sahyoun N. Relationship of genes encoding Ca2+/calmodulin-dependent protein kinase Gr and calspermin: a gene within a gene. Proc Natl Acad Sci U S A. 1991 Jul 1;88(13):5784–5788. doi: 10.1073/pnas.88.13.5784. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Oliva R., Dixon G. H. Vertebrate protamine genes and the histone-to-protamine replacement reaction. Prog Nucleic Acid Res Mol Biol. 1991;40:25–94. doi: 10.1016/s0079-6603(08)60839-9. [DOI] [PubMed] [Google Scholar]
  27. Ono T., Slaughter G. R., Cook R. G., Means A. R. Molecular cloning sequence and distribution of rat calspermin, a high affinity calmodulin-binding protein. J Biol Chem. 1989 Feb 5;264(4):2081–2087. [PubMed] [Google Scholar]
  28. 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]
  29. Quinn P. G. Distinct activation domains within cAMP response element-binding protein (CREB) mediate basal and cAMP-stimulated transcription. J Biol Chem. 1993 Aug 15;268(23):16999–17009. [PubMed] [Google Scholar]
  30. Robinson M. O., McCarrey J. R., Simon M. I. Transcriptional regulatory regions of testis-specific PGK2 defined in transgenic mice. Proc Natl Acad Sci U S A. 1989 Nov;86(21):8437–8441. doi: 10.1073/pnas.86.21.8437. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Shapiro D. J., Sharp P. A., Wahli W. W., Keller M. J. A high-efficiency HeLa cell nuclear transcription extract. DNA. 1988 Jan-Feb;7(1):47–55. doi: 10.1089/dna.1988.7.47. [DOI] [PubMed] [Google Scholar]
  32. Sheng M., Thompson M. A., Greenberg M. E. CREB: a Ca(2+)-regulated transcription factor phosphorylated by calmodulin-dependent kinases. Science. 1991 Jun 7;252(5011):1427–1430. doi: 10.1126/science.1646483. [DOI] [PubMed] [Google Scholar]
  33. Slaughter G. R., Means A. R. Analysis of expression of multiple genes encoding calmodulin during spermatogenesis. Mol Endocrinol. 1989 Oct;3(10):1569–1578. doi: 10.1210/mend-3-10-1569. [DOI] [PubMed] [Google Scholar]
  34. Smale S. T., Baltimore D. The "initiator" as a transcription control element. Cell. 1989 Apr 7;57(1):103–113. doi: 10.1016/0092-8674(89)90176-1. [DOI] [PubMed] [Google Scholar]
  35. Smale S. T., Schmidt M. C., Berk A. J., Baltimore D. Transcriptional activation by Sp1 as directed through TATA or initiator: specific requirement for mammalian transcription factor IID. Proc Natl Acad Sci U S A. 1990 Jun;87(12):4509–4513. doi: 10.1073/pnas.87.12.4509. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Sun P., Enslen H., Myung P. S., Maurer R. A. Differential activation of CREB by Ca2+/calmodulin-dependent protein kinases type II and type IV involves phosphorylation of a site that negatively regulates activity. Genes Dev. 1994 Nov 1;8(21):2527–2539. doi: 10.1101/gad.8.21.2527. [DOI] [PubMed] [Google Scholar]
  37. Tamura T., Makino Y., Mikoshiba K., Muramatsu M. Demonstration of a testis-specific trans-acting factor Tet-1 in vitro that binds to the promoter of the mouse protamine 1 gene. J Biol Chem. 1992 Mar 5;267(7):4327–4332. [PubMed] [Google Scholar]
  38. Tash J. S., Krinks M., Patel J., Means R. L., Klee C. B., Means A. R. Identification, characterization, and functional correlation of calmodulin-dependent protein phosphatase in sperm. J Cell Biol. 1988 May;106(5):1625–1633. doi: 10.1083/jcb.106.5.1625. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Waeber G., Habener J. F. Novel testis germ cell-specific transcript of the CREB gene contains an alternatively spliced exon with multiple in-frame stop codons. Endocrinology. 1992 Oct;131(4):2010–2015. doi: 10.1210/endo.131.4.1396344. [DOI] [PubMed] [Google Scholar]
  40. Waeber G., Meyer T. E., LeSieur M., Hermann H. L., Gérard N., Habener J. F. Developmental stage-specific expression of cyclic adenosine 3',5'-monophosphate response element-binding protein CREB during spermatogenesis involves alternative exon splicing. Mol Endocrinol. 1991 Oct;5(10):1418–1430. doi: 10.1210/mend-5-10-1418. [DOI] [PubMed] [Google Scholar]
  41. Wolgemuth D. J., Watrin F. List of cloned mouse genes with unique expression patterns during spermatogenesis. Mamm Genome. 1991;1(4):283–288. doi: 10.1007/BF00352340. [DOI] [PubMed] [Google Scholar]
  42. Yamamoto K. K., Gonzalez G. A., Biggs W. H., 3rd, Montminy M. R. Phosphorylation-induced binding and transcriptional efficacy of nuclear factor CREB. Nature. 1988 Aug 11;334(6182):494–498. doi: 10.1038/334494a0. [DOI] [PubMed] [Google Scholar]
  43. Zambrowicz B. P., Harendza C. J., Zimmermann J. W., Brinster R. L., Palmiter R. D. Analysis of the mouse protamine 1 promoter in transgenic mice. Proc Natl Acad Sci U S A. 1993 Jun 1;90(11):5071–5075. doi: 10.1073/pnas.90.11.5071. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Zambrowicz B. P., Palmiter R. D. Testis-specific and ubiquitous proteins bind to functionally important regions of the mouse protamine-1 promoter. Biol Reprod. 1994 Jan;50(1):65–72. doi: 10.1095/biolreprod50.1.65. [DOI] [PubMed] [Google Scholar]
  45. de Groot R. P., den Hertog J., Vandenheede J. R., Goris J., Sassone-Corsi P. Multiple and cooperative phosphorylation events regulate the CREM activator function. EMBO J. 1993 Oct;12(10):3903–3911. doi: 10.1002/j.1460-2075.1993.tb06068.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. van der Hoorn F. A., Tarnasky H. A. Factors involved in regulation of the RT7 promoter in a male germ cell-derived in vitro transcription system. Proc Natl Acad Sci U S A. 1992 Jan 15;89(2):703–707. doi: 10.1073/pnas.89.2.703. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. von der Ahe D., Janich S., Scheidereit C., Renkawitz R., Schütz G., Beato M. Glucocorticoid and progesterone receptors bind to the same sites in two hormonally regulated promoters. Nature. 1985 Feb 21;313(6004):706–709. doi: 10.1038/313706a0. [DOI] [PubMed] [Google Scholar]

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

RESOURCES