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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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In situ hybridization of BSN RNAs in  mouse testis. Testes were isolated from mice at various times after birth, and frozen sections (10 μm)  were hybridized with a 576-nucleotide digoxygeninlabeled cRNA (A–E) or sense (F) control corresponding to the shared region of BSN1a and BSN1b  RNAs. After hybridization, sections were washed extensively and developed for equal times (Yang et al.,  1996). Shown are samples from: A, postnatal day zero  (p0); B, p14; C–F, p35. Bar: (A and B) ∼175 μm; (C)  420 μm; (D) 100 μm; (E and F) 40 μm.
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Figure 5: In situ hybridization of BSN RNAs in mouse testis. Testes were isolated from mice at various times after birth, and frozen sections (10 μm) were hybridized with a 576-nucleotide digoxygeninlabeled cRNA (A–E) or sense (F) control corresponding to the shared region of BSN1a and BSN1b RNAs. After hybridization, sections were washed extensively and developed for equal times (Yang et al., 1996). Shown are samples from: A, postnatal day zero (p0); B, p14; C–F, p35. Bar: (A and B) ∼175 μm; (C) 420 μm; (D) 100 μm; (E and F) 40 μm.

Mentions: To determine where BSN mRNAs are expressed within the testis, we conducted in situ hybridizations on frozen sections of mouse testes isolated at various stages of postnatal development. A digoxygenin-labeled antisense BSN cRNA hybridized strongly in the seminiferous tubules of all testis samples examined (Fig. 5). Hybridization was detected at the periphery of the tubules and appeared to be present even at birth, before spermatogenesis (Fig. 5, A). Hybridization remained high throughout most of spermatogenesis. At 2 wk of postnatal development, hybridization was strongest in the centers of the tubules, where the primary spermatocytes are located, and weaker at the periphery, where the spermatogonia reside (Fig. 5, B). By 4–5 wk of age, spermatid formation, i.e., spermiogenesis, had begun (Rugh, 1990), and BSN RNAs were still detected throughout the tubules (Fig. 5, C–E). The persistence of BSN mRNA in late-stage spermiogenesis suggests that BSN RNAs are stable, as is the case with many mRNAs that are translated at this time (for review see Browder et al., 1991). Basonuclin cRNA hybridization was largely specific for derivatives of the germ cell population within the testis and was not detected in the interstitial Leydig cells. If present at all in Sertoli cells, the signal was reduced over that seen in germ cells. No hybridization was seen with the sense control cRNA (Fig. 5 F).


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)

In situ hybridization of BSN RNAs in  mouse testis. Testes were isolated from mice at various times after birth, and frozen sections (10 μm)  were hybridized with a 576-nucleotide digoxygeninlabeled cRNA (A–E) or sense (F) control corresponding to the shared region of BSN1a and BSN1b  RNAs. After hybridization, sections were washed extensively and developed for equal times (Yang et al.,  1996). Shown are samples from: A, postnatal day zero  (p0); B, p14; C–F, p35. Bar: (A and B) ∼175 μm; (C)  420 μm; (D) 100 μm; (E and F) 40 μm.
© Copyright Policy
Related In: Results  -  Collection

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

Figure 5: In situ hybridization of BSN RNAs in mouse testis. Testes were isolated from mice at various times after birth, and frozen sections (10 μm) were hybridized with a 576-nucleotide digoxygeninlabeled cRNA (A–E) or sense (F) control corresponding to the shared region of BSN1a and BSN1b RNAs. After hybridization, sections were washed extensively and developed for equal times (Yang et al., 1996). Shown are samples from: A, postnatal day zero (p0); B, p14; C–F, p35. Bar: (A and B) ∼175 μm; (C) 420 μm; (D) 100 μm; (E and F) 40 μm.
Mentions: To determine where BSN mRNAs are expressed within the testis, we conducted in situ hybridizations on frozen sections of mouse testes isolated at various stages of postnatal development. A digoxygenin-labeled antisense BSN cRNA hybridized strongly in the seminiferous tubules of all testis samples examined (Fig. 5). Hybridization was detected at the periphery of the tubules and appeared to be present even at birth, before spermatogenesis (Fig. 5, A). Hybridization remained high throughout most of spermatogenesis. At 2 wk of postnatal development, hybridization was strongest in the centers of the tubules, where the primary spermatocytes are located, and weaker at the periphery, where the spermatogonia reside (Fig. 5, B). By 4–5 wk of age, spermatid formation, i.e., spermiogenesis, had begun (Rugh, 1990), and BSN RNAs were still detected throughout the tubules (Fig. 5, C–E). The persistence of BSN mRNA in late-stage spermiogenesis suggests that BSN RNAs are stable, as is the case with many mRNAs that are translated at this time (for review see Browder et al., 1991). Basonuclin cRNA hybridization was largely specific for derivatives of the germ cell population within the testis and was not detected in the interstitial Leydig cells. If present at all in Sertoli cells, the signal was reduced over that seen in germ cells. No hybridization was seen with the sense control cRNA (Fig. 5 F).

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