Analysis of meiotic DNAdamage checkpoint

人类减数分裂的分子机制尚不清楚，目前还没有建立有效的分析方法。我们检测了在分裂酵母减数分裂过程中对DNA损伤的检查点系统。在营养CDSL细胞中，检查点Rad(Rad1，Rad3等)羟基脲(HU)依赖激活Chk1使细胞周期停滞于G2期。但在减数分裂过程中，Chk1依赖于Hu的反应减弱。为了研究减数分裂DNA损伤检查点系统的存在，在减数分裂早期用烷化剂甲基甲烷磺酸盐(MMS；0.01%)处理Rad1、Chk1、cds1突变体。这些突变体的染色体分离异常、减数分裂延迟和异常分裂的速度与野生型相似。在患有MMS的减数分裂CDS1细胞中，与营养细胞相比，免疫印迹中观察到Chk1蛋白的轻微磷酸化。在减数分裂的S期，Cdc2-Tyr15也被去磷酸化。本研究中检查点突变体对MMS的损伤反应表明，在分裂酵母减数分裂过程中，DNA损伤检查点的存在或减弱。最近，其他研究小组证明，自发的S期损伤可以通过激活重组而不需要检查点停滞来修复。检验点的异常是否会导致人类减数分裂异常，目前尚无定论。由于已经在人类中发现了一些与DNA重组相关的基因，因此对这些基因进行检测对了解减数分裂的分子机制具有重要意义。

Molecular mechanism in human meiosis is still unknown as no effective methods are established to analyze. We examined checkpoint system to DNA damage during fission yeast meiosis.In vegetative cdsl cells, checkpoint Rad (Rad1, Rad3 etc.) dependent activation of Chk1 in response to hydroxyurea(HU) arrest the cell cycle in G2. But Chk1 dependent response to HU during meiosis is attenuated. To investigate the existence of meiotic DNA damage checkpoint system, rad1, chk1, cds1 mutants are treated with alkylating agent methylmethane sulfonate(MMS; 0.01%) during early meiosis. These mutants undergo aberrant chromosomal segregation, delayed and abnormal meiotic divisions at s similar rate to wild type. In meiotic cds1 cells with MMS, subtle phosphorylation in Chk1 protein is observed in immunoblot, compared to the vegetative cells. Cdc2-Tyr15 in meiotic S phase is also dephosphorylated. The DNA damage response of checkpoint mutants to MMS in this study suggests DNA damage checkpoint in fission yeast meiosis is attenuated or not in existence.Recently, other groups have demonstrated that spontaneous S phase damage is repaired by activating recombination without checkpoint arrest. It is not shown whether the checkpoint aberration causes meiotic abnormality in human. As some genes related to the DNA recombination have already identified in human, it would be significant to examine the genes to know the molecular mechanism in meiosis.