Abstract
Heterochromatic homology ensures the segregation of achiasmate chromosomes during meiosis I in Drosophila melanogaster females, perhaps as a consequence of the heterochromatic threads that connect achiasmate homologs during prometaphase I. Here, we ask how these threads, and other possible heterochromatic entanglements, are resolved prior to anaphase I. We show that the knockdown of Topoisomerase II (Top2) by RNAi in the later stages of meiosis results in a specific defect in the separation of heterochromatic regions after spindle assembly. In Top2 RNAi-expressing oocytes, heterochromatic regions of both achiasmate and chiasmate chromosomes often failed to separate during prometaphase I and metaphase I. Heterochromatic regions were stretched into long, abnormal projections with centromeres localizing near the tips of the projections in some oocytes. Despite these anomalies, we observed bipolar spindles in most Top2 RNAi-expressing oocytes, although the obligately achiasmate 4th chromosomes exhibited a near complete failure to move toward the spindle poles during prometaphase I. Both achiasmate and chiasmate chromosomes displayed defects in biorientation. Given that euchromatic regions separate much earlier in prophase, no defects were expected or observed in the ability of euchromatic regions to separate during late prophase upon knockdown of Top2 at mid-prophase. Finally, embryos from Top2 RNAi-expressing females frequently failed to initiate mitotic divisions. These data suggest both that Topoisomerase II is involved in the resolution of heterochromatic DNA entanglements during meiosis I and that these entanglements must be resolved in order to complete meiosis.
Highlights
In most organisms, crossing over between homologs during meiosis ensures their faithful segregation at the first meiotic division
In the oocytes of the fruit fly Drosophila melanogaster, DNA entanglements persist between heterochromatic regions of the chromosomes until after spindle assembly and may facilitate the proper segregation of chromosomes during meiosis
We analyzed Top2 RNAi-expressing oocytes using immunofluorescence with an antibody recognizing a-tubulin to mark the meiotic spindle and one recognizing histone 3 phosphorylated at serine 10 (H3S10p), which marks nuclei that have entered prometaphase I of meiosis and fortuitously highlights the DNA threads connecting achiasmate chromosomes during D. melanogaster female meiosis [8]
Summary
In most organisms, crossing over between homologs during meiosis ensures their faithful segregation at the first meiotic division. During prometaphase I, achiasmate chromosomes move dynamically on the spindle before properly biorienting and congress into a mass with the chiasmate chromosomes at metaphase I [6,7]. During these movements, achiasmate 4th and X chromosomes are connected by heterochromatic threads, which may play a role in the mechanism by which heterochromatin mediates chromosome segregation [6,8]. Achiasmate 4th and X chromosomes are connected by heterochromatic threads, which may play a role in the mechanism by which heterochromatin mediates chromosome segregation [6,8] How these threads are formed is unknown, but they could potentially arise from stalled replication intermediates [9]
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