Role of the Y-located putative gonadoblastoma gene in human spermatogenesis

Figures & data

Figure 1.  Diagrammatic illustration of TSPY and TSPX domains and effects on cyclin B1-CDK1 kinase activities. (A) TSPY and TSPX proteins share significant similarity at their SET/NAP (NAP) domain, but diverge at flanking sequences, particularly at their carboxyl termini. TSPX harbors an acidic tail, which is absent in TSPY. (B) Cyclin B-CDK1 complex phosphorylates target proteins in an energy dependent manner involving ATP. (C) TSPY interacts with cyclin B and enhances while (D) TSPX also interacts with cyclin B but inhibits the cyclin B-CDK1 kinase activities. (E) Conjugating the acidic domain of TSPX to the carboxyl terminus of TSPY renders the recombinant protein to be inhibitory in the same cyclin B-CDK1 kinase assay, suggesting that the TSPX acidic domain is responsible for its repression on cyclin B-CDK1 kinase activities [Li and Lau 2008].

Figure 2.  Co-localization of TSPY and cyclin B1 in COS7 cells. COS7 cells were transiently transfected with a human TSPY expression vector (pCS2-hTSPY) and were analyzed by immunofluorescence using a mouse monoclonal antibody against human TSPY (red in left panels) and a rabbit polyclonal antibody against human cyclin B1 (green in left panels; red in right panels) [Kido and Lau 2005; Li and Lau 2008]. On the right panel, a mouse monoclonal antibody against α-tubulin was used to detect the microtubule in the mitotic spindle (green). Binding of the primary antibodies were visualized with respective fluorescence dye labelled secondary antibodies. DNA was visualized by DAPI staining (blue). Note that, at M-phase, both transfected TSPY and endogenous cyclin B1 were localized on mitotic spindle. Scale bar = 20 μm. Color figure shown in electronic copy.

Figure 3.  Immunofluorescence analysis of TSPY, TSPX and cyclin B1 in normal adult human testis. (A) Five-micron tissue sections were incubated with anti-TSPY monoclonal antibody (red) [Kido and Lau 2005] or rabbit polyclonal anti-TSPX antibody (green) [Lau et al. 2007]. DNA was visualized by DAPI staining (blue). Bottom panels show the magnified images of boxed area indicated in top panel. TSPY was specifically expressed in spermatogonia and spermatocytes (Spg and Spc, respectively), while TSPX was preferentially expressed in the nuclei of Sertoli cells (Ser) and only at low levels in spermatogonia (Spg). (B) Immunofluorescence of TSPY (red) and cyclin B1 (green) in normal human testis. Pre-mitotic spermatogonia (arrowheads in left panel) and mitotic spermatogonia (arrowheads in right panel) are presented at high magnification. TSPY and cyclin B1 were co-localized in both pre-mitotic and mitotic spermatogonia. Experiments were conducted under an approved protocol by the Institutional Committee on Human Research, VA Medical Center, San Francisco. Scale bar = 20 μm. Color figure shown in electronic copy.

Figure 4.  Relative expression levels of TSPY and cyclin-B1 during different spermatogenic stages in human testis. (A) Immunofluorescence of TSPY (red) and cyclin B1 (green) in spermatogonia, spermatocytes, and spermatids at various spermatogenic stages of a normal human testis [Kido and Lau 2005]. The specific spermatogenic cell types are indicated by brackets with adjacent numbers that correspond to respective stages of spermatogenesis, as illustrated in B. Scale bar = 20 μm. (B) Relative immunofluorescence signals of TSPY and cyclin B1 were diagrammatically illustrated along with schematic representation of spermatogonia, spermatocytes at prophase I and meiosis divisions. Color figure shown in electronic copy.

Li, Y. and Lau, Y.F. (2008) TSPY and its X-encoded homologue interact with cyclin B but exert contrasting functions on cyclin-dependent kinase 1 activities. Oncogene 27:6141–6150.

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Kido, T. and Lau, Y.F. (2005) A Cre gene directed by a human TSPY promoter is specific for germ cells and neurons. Genesis 42:263–275.

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Lau, Y.F., Kido, T. and Li, Y.M. (2007) The TSPY Gene Family. In The Y Chromosome and Male Germ Cell Biology Lau, Y. F. and Chan. W.Y. eds, World Scientific Press, NJ. pp. 73–90.

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