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An unexpected localization of basonuclin in the centrosome, mitochondria, and acrosome of developing spermatids.

Yang Z, Gallicano GI, Yu QC, Fuchs E - J. Cell Biol. (1997)

Bottom Line: In epidermis, basonuclin is associated with the nuclei of mitotically active basal cells but not in terminally differentiating keratinocytes.Cell.Its localization with the acrosome suggests that it may also perform a special function during or shortly after fertilization.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Genetics and Cell Biology, The Howard Hughes Medical Institute, The University of Chicago, Illinois 60637, USA.

ABSTRACT
Basonuclin is a zinc finger protein that was thought to be restricted to keratinocytes of stratified squamous epithelia. In epidermis, basonuclin is associated with the nuclei of mitotically active basal cells but not in terminally differentiating keratinocytes. We report here the isolation of a novel form of basonuclin, which we show is also expressed in stratified epithelia. Most unexpectedly, we find both forms in testis, where a surprising localization pattern was uncovered. While basonuclin RNA expression occurs in mitotically active germ cells, protein was not detected until the meiotic stage, where basonuclin localized to the appendage of the distal centriole of spermatocytes and spermatids. Near the end of spermiogenesis, basonuclin also accumulated in the acrosome and mitochondrial sheath surrounding the flagellum. Intriguingly, a perfect six-amino acid residue mitochondrial targeting sequence (Komiya, T., N. Hachiya, M. Sakaguchi, T. Omura, and K. Mihara. 1994. J. Biol. Chem. 269:30893-30897; Shore, G.C., H.M. McBride, D.G. Millar, N.A. Steenaart, and M. Nguyen. 1995. Eur. J. Biochem. 227: 9-18; McBride, H.M., I.S. Goping, and G.C. Shore. 1996. J. Cell. Biol. 134:307-313) is present in basonuclin 1a but not in the 1b form. Moreover, three distinct affinity-purified peptide antibodies gave this unusual pattern of basonuclin antibody staining, which was confirmed by cell fractionation studies. Our findings suggest a unique role for basonuclin in centrosomes within the developing spermatid, and a role for one of the protein forms in germ cell mitochondrial function. Its localization with the acrosome suggests that it may also perform a special function during or shortly after fertilization.

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Anti-BSN antibodies label the acrosomes and middle piece of developing spermatids. Frozen sections of adult mouse testis  were processed as described in the legend to Fig. 7, except that propidium iodide (red) was used instead of DAPI to label chromatin. (A)  Low magnification to show strong staining of spermatids (Sp) near the lumen (Lu) of all seminiferous tubules. Sg, spermatogonia; 1° and  2°, spermatocytes at the first and second meiotic stage, respectively. (B) Higher magnification of spermatids showing αBSN staining of  both the head region and also the middle piece of the tail (arrows). (C) Lower magnification to show that spermatids in all seminiferous  tubules of the adult testis labeled strongly with anti-BSN antibodies; note dotlike staining of spermatocyte centrosomes as well. (D)  Double label of spermatids with propidium iodide and αBSN to show that the majority of αBSN labeling is in the acrosome and not the  nucleus. Bar: (A) 20 μm; (B) 9 μm; (C) 30 μm; (D) 7 μm.
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Figure 8: Anti-BSN antibodies label the acrosomes and middle piece of developing spermatids. Frozen sections of adult mouse testis were processed as described in the legend to Fig. 7, except that propidium iodide (red) was used instead of DAPI to label chromatin. (A) Low magnification to show strong staining of spermatids (Sp) near the lumen (Lu) of all seminiferous tubules. Sg, spermatogonia; 1° and 2°, spermatocytes at the first and second meiotic stage, respectively. (B) Higher magnification of spermatids showing αBSN staining of both the head region and also the middle piece of the tail (arrows). (C) Lower magnification to show that spermatids in all seminiferous tubules of the adult testis labeled strongly with anti-BSN antibodies; note dotlike staining of spermatocyte centrosomes as well. (D) Double label of spermatids with propidium iodide and αBSN to show that the majority of αBSN labeling is in the acrosome and not the nucleus. Bar: (A) 20 μm; (B) 9 μm; (C) 30 μm; (D) 7 μm.

Mentions: In sexually mature adult testes, anti-BSN antibodies strongly stained the spermatid heads (Fig. 8). Costaining with propidium iodide, which labels chromatin, indicated that this labeling was not nuclear. The crescent-shaped staining pattern, coupled with the appearance of this strong staining in the spermatid region of the seminiferous tubules, was reflective of that seen for acrosomal proteins in spermatids (Lepage and Roberts, 1995; Walensky and Snyder, 1995; Yoshiki et al., 1995). Interestingly, despite the fact that spermiogenesis in mouse is initiated by 4 wk, and that acrosomal caps are seen throughout the centers of the 4-wk seminiferous tubules, these caps did not stain with αBSN (not shown). The relatively late acquisition of anti-BSN staining in the acrosome suggested that basonuclin is a component of late-stage sperm acrosomes.


An unexpected localization of basonuclin in the centrosome, mitochondria, and acrosome of developing spermatids.

Yang Z, Gallicano GI, Yu QC, Fuchs E - J. Cell Biol. (1997)

Anti-BSN antibodies label the acrosomes and middle piece of developing spermatids. Frozen sections of adult mouse testis  were processed as described in the legend to Fig. 7, except that propidium iodide (red) was used instead of DAPI to label chromatin. (A)  Low magnification to show strong staining of spermatids (Sp) near the lumen (Lu) of all seminiferous tubules. Sg, spermatogonia; 1° and  2°, spermatocytes at the first and second meiotic stage, respectively. (B) Higher magnification of spermatids showing αBSN staining of  both the head region and also the middle piece of the tail (arrows). (C) Lower magnification to show that spermatids in all seminiferous  tubules of the adult testis labeled strongly with anti-BSN antibodies; note dotlike staining of spermatocyte centrosomes as well. (D)  Double label of spermatids with propidium iodide and αBSN to show that the majority of αBSN labeling is in the acrosome and not the  nucleus. Bar: (A) 20 μm; (B) 9 μm; (C) 30 μm; (D) 7 μm.
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2139879&req=5

Figure 8: Anti-BSN antibodies label the acrosomes and middle piece of developing spermatids. Frozen sections of adult mouse testis were processed as described in the legend to Fig. 7, except that propidium iodide (red) was used instead of DAPI to label chromatin. (A) Low magnification to show strong staining of spermatids (Sp) near the lumen (Lu) of all seminiferous tubules. Sg, spermatogonia; 1° and 2°, spermatocytes at the first and second meiotic stage, respectively. (B) Higher magnification of spermatids showing αBSN staining of both the head region and also the middle piece of the tail (arrows). (C) Lower magnification to show that spermatids in all seminiferous tubules of the adult testis labeled strongly with anti-BSN antibodies; note dotlike staining of spermatocyte centrosomes as well. (D) Double label of spermatids with propidium iodide and αBSN to show that the majority of αBSN labeling is in the acrosome and not the nucleus. Bar: (A) 20 μm; (B) 9 μm; (C) 30 μm; (D) 7 μm.
Mentions: In sexually mature adult testes, anti-BSN antibodies strongly stained the spermatid heads (Fig. 8). Costaining with propidium iodide, which labels chromatin, indicated that this labeling was not nuclear. The crescent-shaped staining pattern, coupled with the appearance of this strong staining in the spermatid region of the seminiferous tubules, was reflective of that seen for acrosomal proteins in spermatids (Lepage and Roberts, 1995; Walensky and Snyder, 1995; Yoshiki et al., 1995). Interestingly, despite the fact that spermiogenesis in mouse is initiated by 4 wk, and that acrosomal caps are seen throughout the centers of the 4-wk seminiferous tubules, these caps did not stain with αBSN (not shown). The relatively late acquisition of anti-BSN staining in the acrosome suggested that basonuclin is a component of late-stage sperm acrosomes.

Bottom Line: In epidermis, basonuclin is associated with the nuclei of mitotically active basal cells but not in terminally differentiating keratinocytes.Cell.Its localization with the acrosome suggests that it may also perform a special function during or shortly after fertilization.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Genetics and Cell Biology, The Howard Hughes Medical Institute, The University of Chicago, Illinois 60637, USA.

ABSTRACT
Basonuclin is a zinc finger protein that was thought to be restricted to keratinocytes of stratified squamous epithelia. In epidermis, basonuclin is associated with the nuclei of mitotically active basal cells but not in terminally differentiating keratinocytes. We report here the isolation of a novel form of basonuclin, which we show is also expressed in stratified epithelia. Most unexpectedly, we find both forms in testis, where a surprising localization pattern was uncovered. While basonuclin RNA expression occurs in mitotically active germ cells, protein was not detected until the meiotic stage, where basonuclin localized to the appendage of the distal centriole of spermatocytes and spermatids. Near the end of spermiogenesis, basonuclin also accumulated in the acrosome and mitochondrial sheath surrounding the flagellum. Intriguingly, a perfect six-amino acid residue mitochondrial targeting sequence (Komiya, T., N. Hachiya, M. Sakaguchi, T. Omura, and K. Mihara. 1994. J. Biol. Chem. 269:30893-30897; Shore, G.C., H.M. McBride, D.G. Millar, N.A. Steenaart, and M. Nguyen. 1995. Eur. J. Biochem. 227: 9-18; McBride, H.M., I.S. Goping, and G.C. Shore. 1996. J. Cell. Biol. 134:307-313) is present in basonuclin 1a but not in the 1b form. Moreover, three distinct affinity-purified peptide antibodies gave this unusual pattern of basonuclin antibody staining, which was confirmed by cell fractionation studies. Our findings suggest a unique role for basonuclin in centrosomes within the developing spermatid, and a role for one of the protein forms in germ cell mitochondrial function. Its localization with the acrosome suggests that it may also perform a special function during or shortly after fertilization.

Show MeSH
Related in: MedlinePlus