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pEg7, a new Xenopus protein required for mitotic chromosome condensation in egg extracts.

Cubizolles F, Legagneux V, Le Guellec R, Chartrain I, Uzbekov R, Ford C, Le Guellec K - J. Cell Biol. (1998)

Bottom Line: This maternal mRNA encodes a protein, pEg7, whose expression is strongly increased during oocyte maturation.A similar localization of pEg7 is observed when sperm chromatin is allowed to form mitotic chromosomes in cytostatic factor-arrested egg extracts.Biochemical experiments show that pEg7 associates with Xenopus chromosome-associated polypeptides C and E, two components of the 13S condensin.

View Article: PubMed Central - PubMed

Affiliation: Biologie et Génétique du Développement, CNRS UPR 41, Université de Rennes I, Campus de Beaulieu, 35042 Rennes cedex, France.

ABSTRACT
We have isolated a cDNA, Eg7, corresponding to a Xenopus maternal mRNA, which is polyadenylated in mature oocytes and deadenylated in early embryos. This maternal mRNA encodes a protein, pEg7, whose expression is strongly increased during oocyte maturation. The tissue and cell expression pattern of pEg7 indicates that this protein is only readily detected in cultured cells and germ cells. Immunolocalization in Xenopus cultured cells indicates that pEg7 concentrates onto chromosomes during mitosis. A similar localization of pEg7 is observed when sperm chromatin is allowed to form mitotic chromosomes in cytostatic factor-arrested egg extracts. Incubating these extracts with antibodies directed against two distinct parts of pEg7 provokes a strong inhibition of the condensation and resolution of mitotic chromosomes. Biochemical experiments show that pEg7 associates with Xenopus chromosome-associated polypeptides C and E, two components of the 13S condensin.

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Immunolocalization of  pEg7 on mitotic chromosomes  assembled in vitro. Chromosomes were assembled under  standard conditions (see Materials and Methods) and aliquots  were taken to analyze DNA at  different time points. Samples  were fixed and DNA was stained  with ethidium bromide (a, c, e, g,  i, and k) and pEg7 was detected  by indirect immunofluorescence  using anti–pEg7 G immune serum (b, d, f, h, and j) or preimmune serum as a control (l). (a  and b) 0 min, (c and d) 15 min, (e  and f) 30 min, (g and h) 60 min,  (i–l) 120 min. Bar, 5 μm.
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Figure 4: Immunolocalization of pEg7 on mitotic chromosomes assembled in vitro. Chromosomes were assembled under standard conditions (see Materials and Methods) and aliquots were taken to analyze DNA at different time points. Samples were fixed and DNA was stained with ethidium bromide (a, c, e, g, i, and k) and pEg7 was detected by indirect immunofluorescence using anti–pEg7 G immune serum (b, d, f, h, and j) or preimmune serum as a control (l). (a and b) 0 min, (c and d) 15 min, (e and f) 30 min, (g and h) 60 min, (i–l) 120 min. Bar, 5 μm.

Mentions: Chromosome condensation has been analyzed using an in vitro assay involving mitotic egg extracts. To assess the behavior of these extracts, demembranated sperm nuclei were added and observed for chromosome condensation by staining with Ethidium bromide (Fig. 4, a, c, e, g, i, and k). At the beginning of the incubation, sperm chromatin exhibited a highly compact, snake-like shape (Fig. 4 a). Upon incubation, this compact chromatin rapidly swelled and partly decondensed (Fig. 4 c). Further incubation led to local chromatin condensation (Fig. 4 e) and to the formation of entangled, prophase-like chromatin fibers (Fig. 4 g) that finally resolved into highly condensed individual chromosomes (Fig. 4, i and k). These dynamic changes in chromatin structure are in agreement with previous observations (Hirano and Mitchison, 1993).


pEg7, a new Xenopus protein required for mitotic chromosome condensation in egg extracts.

Cubizolles F, Legagneux V, Le Guellec R, Chartrain I, Uzbekov R, Ford C, Le Guellec K - J. Cell Biol. (1998)

Immunolocalization of  pEg7 on mitotic chromosomes  assembled in vitro. Chromosomes were assembled under  standard conditions (see Materials and Methods) and aliquots  were taken to analyze DNA at  different time points. Samples  were fixed and DNA was stained  with ethidium bromide (a, c, e, g,  i, and k) and pEg7 was detected  by indirect immunofluorescence  using anti–pEg7 G immune serum (b, d, f, h, and j) or preimmune serum as a control (l). (a  and b) 0 min, (c and d) 15 min, (e  and f) 30 min, (g and h) 60 min,  (i–l) 120 min. Bar, 5 μm.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2132990&req=5

Figure 4: Immunolocalization of pEg7 on mitotic chromosomes assembled in vitro. Chromosomes were assembled under standard conditions (see Materials and Methods) and aliquots were taken to analyze DNA at different time points. Samples were fixed and DNA was stained with ethidium bromide (a, c, e, g, i, and k) and pEg7 was detected by indirect immunofluorescence using anti–pEg7 G immune serum (b, d, f, h, and j) or preimmune serum as a control (l). (a and b) 0 min, (c and d) 15 min, (e and f) 30 min, (g and h) 60 min, (i–l) 120 min. Bar, 5 μm.
Mentions: Chromosome condensation has been analyzed using an in vitro assay involving mitotic egg extracts. To assess the behavior of these extracts, demembranated sperm nuclei were added and observed for chromosome condensation by staining with Ethidium bromide (Fig. 4, a, c, e, g, i, and k). At the beginning of the incubation, sperm chromatin exhibited a highly compact, snake-like shape (Fig. 4 a). Upon incubation, this compact chromatin rapidly swelled and partly decondensed (Fig. 4 c). Further incubation led to local chromatin condensation (Fig. 4 e) and to the formation of entangled, prophase-like chromatin fibers (Fig. 4 g) that finally resolved into highly condensed individual chromosomes (Fig. 4, i and k). These dynamic changes in chromatin structure are in agreement with previous observations (Hirano and Mitchison, 1993).

Bottom Line: This maternal mRNA encodes a protein, pEg7, whose expression is strongly increased during oocyte maturation.A similar localization of pEg7 is observed when sperm chromatin is allowed to form mitotic chromosomes in cytostatic factor-arrested egg extracts.Biochemical experiments show that pEg7 associates with Xenopus chromosome-associated polypeptides C and E, two components of the 13S condensin.

View Article: PubMed Central - PubMed

Affiliation: Biologie et Génétique du Développement, CNRS UPR 41, Université de Rennes I, Campus de Beaulieu, 35042 Rennes cedex, France.

ABSTRACT
We have isolated a cDNA, Eg7, corresponding to a Xenopus maternal mRNA, which is polyadenylated in mature oocytes and deadenylated in early embryos. This maternal mRNA encodes a protein, pEg7, whose expression is strongly increased during oocyte maturation. The tissue and cell expression pattern of pEg7 indicates that this protein is only readily detected in cultured cells and germ cells. Immunolocalization in Xenopus cultured cells indicates that pEg7 concentrates onto chromosomes during mitosis. A similar localization of pEg7 is observed when sperm chromatin is allowed to form mitotic chromosomes in cytostatic factor-arrested egg extracts. Incubating these extracts with antibodies directed against two distinct parts of pEg7 provokes a strong inhibition of the condensation and resolution of mitotic chromosomes. Biochemical experiments show that pEg7 associates with Xenopus chromosome-associated polypeptides C and E, two components of the 13S condensin.

Show MeSH
Related in: MedlinePlus