A mouse protein that localizes to acrosome and sperm tail is regulated by Y-chromosome

BMC Cell Biology

Rachel A Jesudasan²  Vishnu M Dhople¹  Manju S Devi²  Rupa Bhattacharya³ 

[1] Department of Functional Genomics, Interfaculty Institute for Genetics and Functional Genomics, Universitätsmedizin Greifswald, Friedrich-Ludwig-Jahn-Straße 15 a, Greifswald 17487, Germany;Centre for Cellular and Molecular Biology, Hyderabad, Andhra Pradesh, India;Present Address: Lawrenceville, New Jersey 08648, USA

关键词: Yq-del mice;    Co-IP;    Sperm tail;    Acrosome;    MAST;   

Others  :  855069
DOI  :  10.1186/1471-2121-14-50

 received in 2013-07-09, accepted in 2013-11-14,  发布年份 2013

【 摘 要 】

Background

Acrosomal proteins play crucial roles in the physiology of fertilization. Identification of proteins localizing to the acrosome is fundamental to the understanding of its contribution to fertilization. Novel proteins are still being reported from acrosome. In order to capture yet unreported proteins localizing to acrosome in particular and sperm in general, 2D-PAGE and mass spectrometry analysis of mouse sperm proteins was done.

Results

One of the protein spots identified in the above study was reported in the NCBI database as a hypothetical protein from Riken cDNA 1700026L06 that localizes to chromosome number 2. Immunofluorescence studies using the antibody raised in rabbit against the recombinant protein showed that it localized to mouse acrosome and sperm tail. Based on the localization of this protein, it has been named mouse acrosome and sperm tail protein (MAST, [Q7TPM5 (http://www.ncbi.nlm.nih.gov/protein/Q7TPM5 webcite)]). This protein shows 96% identity to the rat spermatid specific protein RSB66. Western blotting showed that MAST is expressed testis-specifically. Co-immunoprecipitation studies using the MAST antibody identified two calcium-binding proteins, caldendrin and calreticulin as interacting partners of MAST. Caldendrin and calreticulin genes localize to mouse chromosomes 5 and 8 respectively. In a Yq-deletion mutant mouse, that is subfertile and has a deletion of 2/3rd of the long arm of the Y chromosome, MAST failed to localize to the acrosome. Western blot analysis however, revealed equal expression of MAST in the testes of wild type and mutant mice. The acrosomal calcium-binding proteins present in the MAST IP-complex were upregulated in sperms of Yq-del mice.

Conclusions

We have identified a mouse acrosomal protein, MAST, that is expressed testis specifically. MAST does not contain any known motifs for protein interactions; yet it complexes with calcium-binding proteins localizing to the acrosome. The misexpression of all the proteins identified in a complex in the Yq-del mice invokes the hypothesis of a putative pathway regulated by the Y chromosome. The role of Y chromosome in the regulation of this complex is however not clear from the current study.

【 授权许可】

   
2013 Bhattacharya et al.; licensee BioMed Central Ltd.

【 图 表 】

【 参考文献 】

• [1]Baccetti B, Afzelius BA: The biology of the sperm cell. Monogr Dev Biol 1976, 10:1-254.
• [2]Guo R, Yu Z, Guan J, Ge Y, Ma J, Li S, Wang S, Xue S, Han D: Stage-specific and tissue-specific expression characteristics of differentially expressed genes during mouse spermatogenesis. Mol Reprod Dev 2004, 67(3):264-272.
• [3]Huang XY, Guo XJ, Shen J, Wang YF, Chen L, Xie J, Wang NL, Wang FQ, Zhao C, Huo R, et al.: Construction of a proteome profile and functional analysis of the proteins involved in the initiation of mouse spermatogenesis. J Proteome Res 2008, 7(8):3435-3446.
• [4]Eddy EM: Male germ cell gene expression. Recent Prog Horm Res 2002, 57:103-128.
• [5]Buffone MG, Foster JA, Gerton GL: The role of the acrosomal matrix in fertilization. Int J Dev Biol 2008, 52(5–6):511-522.
• [6]Korfanty J, Toma A, Wojtas A, Rusin A, Vydra N, Widlak W: Identification of a new mouse sperm acrosome-associated protein. Reproduction 2012, 143(6):749-757.
• [7]Schultz N, Hamra FK, Garbers DL: A multitude of genes expressed solely in meiotic or postmeiotic spermatogenic cells offers a myriad of contraceptive targets. Proc Natl Acad Sci U S A 2003, 100(21):12201-12206.
• [8]Guo X, Zhang P, Huo R, Zhou Z, Sha J: Analysis of the human testis proteome by mass spectrometry and bioinformatics. Proteomics Clin Appl 2008, 2(12):1651-1657.
• [9]Naaby-Hansen S: Electrophoretic map of acidic and neutral human spermatozoal proteins. J Reprod Immunol 1990, 17(3):167-185.
• [10]Li L-W, Fan L-Q, Zhu W-B, Nie H-C, Sun B-L, Luo K-L, Liao T-T, Tang L, Lu G-X: Establishment of a high-resolution 2-D reference map of human spermatozoal proteins from 12 fertile sperm-bank donors. Asian J Androl 2007, 9(3):312-320.
• [11]Sigrist CJ, Cerutti L, De Castro E, Langendijk-Genevaux PS, Bulliard V, Bairoch A, Hulo N: PROSITE, a protein domain database for functional characterization and annotation. Nucleic Acids Res 2010, 38(Database issue):D161-D166.
• [12]Toure A, Szot M, Mahadevaiah SK, Rattigan A, Ojarikre OA, Burgoyne PS: A new deletion of the mouse Y chromosome long arm associated with the loss of Ssty expression, abnormal sperm development and sterility. Genetics 2004, 166(2):901-912.
• [13]Oliva R, De Mateo S, Estanyol JM: Sperm cell proteomics. Proteomics 2009, 9(4):1004-1017.
• [14]Chen X, Hu T, Liang G, Yang M, Zong S, Miao S, Koide SS, Wang L: A novel testis protein, RSB-66, interacting with INCA1 (inhibitor of Cdk interacting with cyclin A1). Biochem Cell Biol 2008, 86(4):345-351.
• [15]Nakamura M, Michikawa Y, Baba T, Okinaga S, Arai K: Calreticulin is present in the acrosome of spermatids of rat testis. Biochem Biophys Res Commun 1992, 186(2):668-673.
• [16]Spungin B, Breitbart H: Calcium mobilization and influx during sperm exocytosis. J Cell Sci 1996, 109(Pt 7):1947-1955.
• [17]Naaby-Hansen S, Wolkowicz MJ, Klotz K, Bush LA, Westbrook VA, Shibahara H, Shetty J, Coonrod SA, Reddi PP, Shannon J, et al.: Co-localization of the inositol 1,4,5-trisphosphate receptor and calreticulin in the equatorial segment and in membrane bounded vesicles in the cytoplasmic droplet of human spermatozoa. Mol Hum Reprod 2001, 7(10):923-933.
• [18]Jorgensen CS, Heegaard NH, Holm A, Hojrup P, Houen G: Polypeptide binding properties of the chaperone calreticulin. Eur J Biochem 2000, 267(10):2945-2954.
• [19]Park BJ, Lee DG, Yu JR, Jung SK, Choi K, Lee J, Kim YS, Lee JI, Kwon JY, Singson A, et al.: Calreticulin, a calcium-binding molecular chaperone, is required for stress response and fertility in Caenorhabditis elegans. Mol Biol Cell 2001, 12(9):2835-2845.
• [20]Seidenbecher CI, Langnaese K, Sanmarti-Vila L, Boeckers TM, Smalla KH, Sabel BA, Garner CC, Gundelfinger ED, Kreutz MR: Caldendrin, a novel neuronal calcium-binding protein confined to the somato-dendritic compartment. J Biol Chem 1998, 273(33):21324-21331.
• [21]Laube G, Seidenbecher CI, Richter K, Dieterich DC, Hoffmann B, Landwehr M, Smalla KH, Winter C, Bockers TM, Wolf G, et al.: The neuron-specific Ca2 + -binding protein caldendrin: gene structure, splice isoforms, and expression in the rat central nervous system. Mol Cell Neurosci 2002, 19(3):459-475.
• [22]Redecker P, Kreutz MR, Bockmann J, Gundelfinger ED, Boeckers TM: Brain synaptic junctional proteins at the acrosome of rat testicular germ cells. J Histochem Cytochem 2003, 51(6):809-819.
• [23]Guo J, Zhu P, Wu C, Yu L, Zhao S, Gu X: In silico analysis indicates a similar gene expression pattern between human brain and testis. Cytogenet Genome Res 2003, 103(1–2):58-62.
• [24]De Blas G, Michaut M, Trevino CL, Tomes CN, Yunes R, Darszon A, Mayorga LS: The intraacrosomal calcium pool plays a direct role in acrosomal exocytosis. J Biol Chem 2002, 277(51):49326-49331.
• [25]Jehan Z, Vallinayagam S, Tiwari S, Pradhan S, Singh L, Suresh A, Reddy HM, Ahuja YR, Jesudasan RA: Novel noncoding RNA from human Y distal heterochromatic block (Yq12) generates testis-specific chimeric CDC2L2. Genome Res 2007, 17(4):433-440.
• [26]Towbin H, Staehelin T, Gordon J: Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. 1979. Biotechnology 1992, 24:145-149.