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The identification and characterization of a testis-specific cDNA during spermatogenesis

Abstract

Using bioinformatics and experimental validation, we obtained a cDNA (named srsf) which was exclusively expressed in the mouse testes. RT-PCR analysis showed that srsf mRNA was not expressed in the gonad during the sex determination period or during embryogenesis. In developing mouse tests, srsf expression was first detected on post-natal day 10, reached its highest level on day 23, and then reduced to and remained at a moderate level throughout adulthood. In situ hybridization analysis demonstrated that srsf mRNA was expressed in pachytene spermatocytes and round spermatids in the testes. The predicted protein contains one RNA-binding domain (RBD) and a serine-arginine rich domain (RS), which are characterized by some splicing factors of SR family members. These findings indicate that srsf may play a role during spermatogenesis.

Abbreviations

A:

type Aspermatogonia

In:

intermediate spermatogonia

B:

type B spermatogonia

PL:

preleptotene spermatocytes

L:

leptotene spermatocytes

Zs:

zygotene spermatocytes

EPs:

early pachytene spermatocytes

LPs:

late pachytene spermatocytes

Ds:

diplotene spermatocytes

MI:

first meiosis

MII:

second meiosis

Rs:

round spermatids

Es:

elongated spermatids

S16:

step 16 spermatids

E:

early

L:

late

SG:

spermatogonia

SC:

spermatocytes

ST:

round spermatid.

References

  1. 1.

    Kleen, K.C. Patterns of transcriptional regulation in the mammalian testes. Mol. Report Dev. 423 (1996) 268–281.

    Article  Google Scholar 

  2. 2.

    Venables, J.P. Alternative splicing in the tests. Curr. Opin. Genet. Dev. 12 (2002) 615–619.

    PubMed  CAS  Article  Google Scholar 

  3. 3.

    Fu, X.D. Specific commitment of different pre-mRNA to splicing by single SR proteins. Nature 365 (1993) 82–85.

    PubMed  CAS  Article  Google Scholar 

  4. 4.

    Portal, D. and Joaqu’ın, M. An early ancestor in the evolution of splicing: a Trypanosoma cruzi serine-arginin-rich protein (TcSR) is functional in cis-splicing. Mol. Biochem. Parasitol. 127 (2003) 37–46.

    PubMed  CAS  Article  Google Scholar 

  5. 5.

    Roland, T. and James, L. Determinants of SR protein specificity. Curr. Opin. Cell Biol. 11 (1999) 358–362.

    Article  Google Scholar 

  6. 6.

    Krämer, A. The structure and function of proteins involved in mammalian pre-mRNA splicing. Annu. Rev. Biochem. 65 (1996) 367–409.

    PubMed  Article  Google Scholar 

  7. 7.

    Fu, X.D. The superfamily of arginine/serine-rich splicing factors. RNA 1 (1995) 663–680.

    PubMed  CAS  Google Scholar 

  8. 8.

    Manley, J.L. and Tacke, R. SR proteins and splicing control. Genes Dev. 9 (1996) 284–293.

    Google Scholar 

  9. 9.

    Valcárcel, J. and Green, M.R. The SR protein family: pleiotropic functions in pre-mRNA splicing. Trends Biochem. Sci. 21 (1996) 296–301.

    PubMed  Article  Google Scholar 

  10. 10.

    Hastings, M.L. and Krainer, A.R. Splicing in the new millennium. Curr. Opin. Cell Biol. 13 (2001) 302–308.

    PubMed  CAS  Article  Google Scholar 

  11. 11.

    Meissner, M., Lopato, S., Gotzmann, J., Sauermann, G. and Barta, A. Proto-oncoprotein TLS/FUS is associated to the nuclear mitrix and complexed with splicing factors PTB, SRm160, and SR proteins. Exp. Cell Res. 283 (2003) 184–195.

    PubMed  CAS  Article  Google Scholar 

  12. 12.

    Zhu, J. and Krainer, A.R. Pre-mRNA splicing in the absence of a SR protein SR domain. Genes 14 (2000) 3166–3178.

    CAS  Article  Google Scholar 

  13. 13.

    Wang, J., Takegaki, Y. and Manliy, J.L. Targeted disruption of an essential vertebrate gene ASF/SF2 is required for cell viability. Genes Dev. 10 (1996) 2588–2599.

    PubMed  CAS  Google Scholar 

  14. 14.

    Stefan, W., Simion, C., Ivshina, M. and Nickerson, J.A. In vitro FRAP reveals the ATP-dependent nuclear mobilization of the exon junction complex protein SRm160. J. Cell Biol. 164 (2004) 843–850.

    Article  Google Scholar 

  15. 15.

    Stefan, W., Simion, C. and Nickerson, J.A. The spatial targeting and nuclear matrix binding domains of SR m160. Proc. Natl. Acad. Sci. U.S.A. 100 (2003) 3269–3274.

    Article  Google Scholar 

  16. 16.

    Dijkman, H.B.P.M., Mentzel, S, de Jong, A.S., Assmann, K.J.M. RNA in situ hybridization using digoxigenin-labeled cRNA probes. Biochemica 2(1995) 21–25

    Google Scholar 

  17. 17.

    Thompson, J.D., Higgins, D.G., Gibson, and T.J. CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res. 11 (1994) 4673–4680.

    Google Scholar 

  18. 18.

    Birney, E., Kumar, S. and Krainer, A.R. Analysis of the RNA-recognition motif and RS and RGG domains: conservation in metazoan pre-mRNA splicing factors. Nucleic Acids Res. 21 (1993) 5803–5816.

    PubMed  CAS  Google Scholar 

  19. 19.

    Zhang, D., Penttila, T.L., Morris, P.L. and Roeder, R.G. Cell-and stage-specific high-level expression of TBP-related factor 2 (TRF2) during mouse spermatogenesis. Mech. Dev. 106 (2001) 203–205.

    PubMed  CAS  Article  Google Scholar 

  20. 20.

    Ina, S., Tsunekawa, N., Nakamura, A. and Noce, T. Expression of the mouse Aven gene during spermatogenesis, analyzed by subtraction screening using Mvh-knockout mice. Gene Expr. Patterns 5 (2003) 635–638.

    Article  Google Scholar 

  21. 21.

    Bellve, A.R., Cavicchia, J.C., Millette, C.F., O’Brien, D.A., Bhatnagar, Y.N. and Dym, M. Spermatogenic cells of the prepuberal mouse: isolation and morphological characterization. J. Cell Biol. 74 (1977) 68–85.

    PubMed  CAS  Article  Google Scholar 

  22. 22.

    Taizo, K. and Masaki, F.T. Unique and redundant functions of SR proteins, a conserved family of splicing factors, in Caenorhabditis elegants development. Mech. Dev. 95 (2000) 67–76.

    Article  Google Scholar 

  23. 23.

    Mattox, W., McGuffin, M.E. and Baker, B.S. A Negative feedback mechanism revealed by functional analysis of the alternative isoforms of the Drosophila splicing regulator transformer-2. Genetics 143 (1996) 303–314.

    PubMed  CAS  Google Scholar 

  24. 24.

    Rugh, R. The mouse, its reproduction and development. Burgess Publishing Company, Minneapolis. (1968).

    Google Scholar 

  25. 25.

    Sassone-Corsi, P. Unique chromatin remodeling and transcriptional regulation in spermatogenesis. Science 296 (2002) 2176–2178

    PubMed  CAS  Article  Google Scholar 

  26. 26.

    Kimmins, S., Kotaja, N., Davidson, I. and Sassone-Corsi, P. Testis-specific transcription mechanisms promoting male germ-cell differentiation. Repro. 128 (2004) 5–12

    CAS  Article  Google Scholar 

  27. 27.

    Baoan, L., Mahalakshmi, N. and Mackay, D.R. Ovoll regulates meiotic pachytene progression during spermatogenesis by repressing Id2 expression. Development 132 (2005) 1463–1473

    Article  Google Scholar 

  28. 28.

    Tung, K-S. and Bilanchone, V. The pachytene checkpoint prevents accumulation and phosphorylation of the meiosis-specific transcription factor Ndt80. Proc. Natl. Acad. Sci. U.S.A. 97 (2000) 12187–12192.

    PubMed  CAS  Article  Google Scholar 

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Correspondence to Ping Song.

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Chen, Y., Hu, J., Song, P. et al. The identification and characterization of a testis-specific cDNA during spermatogenesis. Cell. Mol. Biol. Lett. 11, 80–89 (2006). https://doi.org/10.2478/s11658-006-0008-4

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Key Words

  • Expression pattern
  • Spermatogenesis
  • Splicing factor