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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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Expression of both  basonuclin RNA forms in testis as well as in stratified squamous epithelia. (A) Reversetranscriptase–PCR (RT-PCR)  Analysis I. RNAs were isolated from the various mouse  tissues indicated and subjected to RT-PCR as described above. Primer sets to a  shared portion of BSN1a and BSN1b were used, along with an actin control. HT, heart; LN, lung; MS, muscle; SL, spleen; KD, kidney;  FS, forestomach; ES, esophagus; SK, skin; OV, ovary; TS, testis. (B) RT-PCR Analysis II. RNAs were isolated from mouse skin and testis. These RNAs were subjected to RT-PCR analysis using primer sets specific for each of the two unique exons and for a shared segment  of mouse BSN1a and BSN1b RNAs, respectively. Appropriate primer sets for β-actin were used as controls. DNA fragments generated  were resolved by electrophoresis through 1% agarose gels. Lanes 1–4, skin RNAs; lanes 5–8, testes RNAs. Fragments shown were generated using primers specific for: lanes 1 and 5, BSN1a; lanes 2 and 6, BSN1b; lanes 3 and 7, BSN1a/1b; lanes 4 and 8, actin.
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Figure 3: Expression of both basonuclin RNA forms in testis as well as in stratified squamous epithelia. (A) Reversetranscriptase–PCR (RT-PCR) Analysis I. RNAs were isolated from the various mouse tissues indicated and subjected to RT-PCR as described above. Primer sets to a shared portion of BSN1a and BSN1b were used, along with an actin control. HT, heart; LN, lung; MS, muscle; SL, spleen; KD, kidney; FS, forestomach; ES, esophagus; SK, skin; OV, ovary; TS, testis. (B) RT-PCR Analysis II. RNAs were isolated from mouse skin and testis. These RNAs were subjected to RT-PCR analysis using primer sets specific for each of the two unique exons and for a shared segment of mouse BSN1a and BSN1b RNAs, respectively. Appropriate primer sets for β-actin were used as controls. DNA fragments generated were resolved by electrophoresis through 1% agarose gels. Lanes 1–4, skin RNAs; lanes 5–8, testes RNAs. Fragments shown were generated using primers specific for: lanes 1 and 5, BSN1a; lanes 2 and 6, BSN1b; lanes 3 and 7, BSN1a/1b; lanes 4 and 8, actin.

Mentions: Previously, basonuclin was thought to be restricted in its expression to mitotically active cells of stratified squamous epithelia (Tseng and Green, 1994). In the course of examining basonuclin RNA expression in different mouse tissues, we were surprised to discover that PCR primers corresponding to the shared sequences of BSN1a and BSN1b detected a band in testis mRNA in addition to RNAs isolated from tissues known to contain keratinocytes (Fig. 3 A).


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)

Expression of both  basonuclin RNA forms in testis as well as in stratified squamous epithelia. (A) Reversetranscriptase–PCR (RT-PCR)  Analysis I. RNAs were isolated from the various mouse  tissues indicated and subjected to RT-PCR as described above. Primer sets to a  shared portion of BSN1a and BSN1b were used, along with an actin control. HT, heart; LN, lung; MS, muscle; SL, spleen; KD, kidney;  FS, forestomach; ES, esophagus; SK, skin; OV, ovary; TS, testis. (B) RT-PCR Analysis II. RNAs were isolated from mouse skin and testis. These RNAs were subjected to RT-PCR analysis using primer sets specific for each of the two unique exons and for a shared segment  of mouse BSN1a and BSN1b RNAs, respectively. Appropriate primer sets for β-actin were used as controls. DNA fragments generated  were resolved by electrophoresis through 1% agarose gels. Lanes 1–4, skin RNAs; lanes 5–8, testes RNAs. Fragments shown were generated using primers specific for: lanes 1 and 5, BSN1a; lanes 2 and 6, BSN1b; lanes 3 and 7, BSN1a/1b; lanes 4 and 8, actin.
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Related In: Results  -  Collection

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

Figure 3: Expression of both basonuclin RNA forms in testis as well as in stratified squamous epithelia. (A) Reversetranscriptase–PCR (RT-PCR) Analysis I. RNAs were isolated from the various mouse tissues indicated and subjected to RT-PCR as described above. Primer sets to a shared portion of BSN1a and BSN1b were used, along with an actin control. HT, heart; LN, lung; MS, muscle; SL, spleen; KD, kidney; FS, forestomach; ES, esophagus; SK, skin; OV, ovary; TS, testis. (B) RT-PCR Analysis II. RNAs were isolated from mouse skin and testis. These RNAs were subjected to RT-PCR analysis using primer sets specific for each of the two unique exons and for a shared segment of mouse BSN1a and BSN1b RNAs, respectively. Appropriate primer sets for β-actin were used as controls. DNA fragments generated were resolved by electrophoresis through 1% agarose gels. Lanes 1–4, skin RNAs; lanes 5–8, testes RNAs. Fragments shown were generated using primers specific for: lanes 1 and 5, BSN1a; lanes 2 and 6, BSN1b; lanes 3 and 7, BSN1a/1b; lanes 4 and 8, actin.
Mentions: Previously, basonuclin was thought to be restricted in its expression to mitotically active cells of stratified squamous epithelia (Tseng and Green, 1994). In the course of examining basonuclin RNA expression in different mouse tissues, we were surprised to discover that PCR primers corresponding to the shared sequences of BSN1a and BSN1b detected a band in testis mRNA in addition to RNAs isolated from tissues known to contain keratinocytes (Fig. 3 A).

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