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Fig. 7. Direct comparison of the levels and localization of specific histone modifications in mouse immature oocyte nuclei and on the metaphase II plate of ovulated eggs. Immature germinal vesicle stage oocytes (GV A-T) and metaphase II arrested eggs (MII A-T) are stained with a panel of histone modification antibodies (Red) and Sytox DNA stain (Green). Transmitted light micrographs are also shown (Light). Regions of DNA that co-localize with histone modifications are seen in yellow (Overlay). Results show that in both immature oocytes and metaphase II arrested eggs, the Me(Lys4)H3 modification strongly co-localizes with DNA (see GV D and MII D). The Me(Lys9)H3 modification also strongly co-localizes with DNA in both immature oocytes and in metaphase II arrested eggs (see GV H and MII H). The hyperacetylated H4 modification also strongly co-localizes with DNA in immature oocyte nuclei (see GV L), however, staining for this modification is negative on metaphase II arrested chromatin (MII K). The Me(Arg17)H3 modification stains strongly in immature oocyte nuclei in a punctate manner (GV O) and appears to co-localize with DNA both around the nucleolus (GV P) and at several other locations in the nucleus (GV P, arrow). Staining for this modification in the ovulated egg, however, is limited to punctate cytoplasmic spots (MII O) and is absent from the chromatin (MII P). Me(Arg3)H4 modification staining is weak and punctate in immature oocyte nuclei (GV S) and does not appear to co-localize with DNA except at one or two locations (GV T, arrow). Staining for this modification in the ovulated egg, however, is absent from the metaphase II chromatin (MII S, MII T). These results show that levels of the hyperacetylated H4, Me(Arg17)H3 and Me(Arg3)H4 modifications are dramatically reduced in ovulated egg chromatin whereas levels of Me(Lys4)H3 and Me(Lys9)H3 modifications do not appear to be affected.
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