Drosophila ATM and ATR have distinct activities in the regulation of meiotic DNA damage and repair.
J Cell Biol. 195:359-67. Abstract
Ataxia telangiectasia-mutated (ATM) and ataxia telangiectasia-related (ATR) kinases are conserved regulators of cellular responses to double strand breaks (DSBs). During meiosis, however, the functions of these kinases in DSB repair and the deoxyribonucleic acid (DNA) damage checkpoint are unclear. In this paper, we show that ATM and ATR have unique roles in the repair of meiotic DSBs in Drosophila melanogaster. ATR mutant analysis indicated that it is required for checkpoint activity, whereas ATM may not be. Both kinases phosphorylate H2AV (gamma-H2AV), and, using this as a reporter for ATM/ATR activity, we found that the DSB repair response is surprisingly dynamic at the site of DNA damage. gamma-H2AV is continuously exchanged, requiring new phosphorylation at the break site until repair is completed. However, most surprising is that the number of gamma-H2AV foci is dramatically increased in the absence of ATM, but not ATR, suggesting that the number of DSBs is increased. Thus, we conclude that ATM is primarily required for the meiotic DSB repair response, which includes functions in DNA damage repair and negative feedback control over the level of programmed DSBs during meiosis.
A Pathway for Synapsis Initiation during Zygotene in Drosophila Oocytes.
Curr Biol. 21:1852-7. Abstract
Formation of the synaptonemal complex (SC), or synapsis, between homologs in meiosis is essential for crossing over and chromosome segregation [1-4]. How SC assembly initiates is poorly understood but may have a critical role in ensuring synapsis between homologs and regulating double-strand break (DSB) and crossover formation. We investigated the genetic requirements for synapsis in Drosophila and found that there are three temporally and genetically distinct stages of synapsis initiation. In "early zygotene" oocytes, synapsis is only observed at the centromeres. We also found that nonhomologous centromeres are clustered during this process. In "mid-zygotene" oocytes, SC initiates at several euchromatic sites. The centromeric and first euchromatic SC initiation sites depend on the cohesion protein ORD. In "late zygotene" oocytes, SC initiates at many more sites that depend on the Kleisin-like protein C(2)M. Surprisingly, late zygotene synapsis initiation events are independent of the earlier mid-zygotene events, whereas both mid and late synapsis initiation events depend on the cohesin subunits SMC1 and SMC3. We propose that the enrichment of cohesion proteins at specific sites promotes homolog interactions and the initiation of euchromatic SC assembly independent of DSBs. Furthermore, the early euchromatic SC initiation events at mid-zygotene may be required for DSBs to be repaired as crossovers.
RanGTP is required for meiotic spindle organization and the initiation of embryonic development in Drosophila.
J Cell Sci. 124:3797-810. Abstract
RanGTP is important for chromosome-dependent spindle assembly in Xenopus extracts. Here we report on experiments to determine the role of the Ran pathway on microtubule dynamics in Drosophila oocytes and embryos. Females expressing a dominant-negative form of Ran have fertility defects, suggesting that RanGTP is required for normal fertility. This is not, however, because of a defect in acentrosomal meiotic spindle assembly. Therefore, RanGTP does not appear to be essential or sufficient for the formation of the acentrosomal spindle. Instead, the most important function of the Ran pathway in spindle assembly appears to be in the tapering of microtubules at the spindle poles, which might be through regulation of proteins such as TACC and the HURP homolog, Mars. One consequence of this spindle organization defect is an increase in the nondisjunction of achiasmate chromosomes. However, the meiotic defects are not severe enough to cause the decreased fertility. Reductions in fertility occur because RanGTP has a role in microtubule assembly that is not directly nucleated by the chromosomes. This includes microtubules nucleated from the sperm aster, which are required for pronuclear fusion. We propose that following nuclear envelope breakdown, RanGTP is released from the nucleus and creates a cytoplasm that is activated for assembling microtubules, which is important for processes such as pronuclear fusion. Around the chromosomes, however, RanGTP might be redundant with other factors such as the chromosome passenger complex.