The SMC5/6 Complex - DNA Damage Response Regulation Ensures Meiotic Fidelity

描述（由申请人提供）：我们知道减数分裂期间DNA损伤反应受到干扰将导致不育，妊娠丢失或遗传缺陷。然而，我们对减数分裂过程中DNA损伤反应的调控知之甚少。我的初步数据表明，SMC 5/6复合物是需要介导的DNA损伤反应在减数分裂细胞周期的进展。由于SMC 5/6复合物对细胞活力至关重要，因此尚未进行哺乳动物体内实验，我们对SMC 5/6在哺乳动物中的功能的理解有限。我正在创建一个生殖细胞特异性突变的Smc 5和突变睾丸特异性SMC 5/6基因Eid 3。这些突变体将使我能够对SMC 5/6在减数分裂中的复杂功能（Aim 1）进行首次全面研究。EID 3是SMC 5/6复合物的睾丸特异性Kleisin亚基。然而，体细胞Kleisin SMC 5/6亚基NSE 4也在睾丸内表达。因此，在睾丸内存在两种SMC 5/6复合物，SMC 5/6 NSE 4和SMC 5/6 EID 3。我将确定这两种复合物之间的相似性和差异（目标2）。我已证实SMC 5/6复合物与BAT 3-EP 300复合物相互作用。BAT 3-EP 300复合物是激活TRP 53介导的DNA损伤反应所必需的。根据SMC 5/6和BAT 3-EP 300复合物之间形成的蛋白质相互作用网络，SMC 5/6通过调节TRP 53乙酰化而成为DNA损伤反应途径的关键组分。在目标3中，我将对SMC 5/6和BAT 3-EP 300之间的相互作用进行详细评估，并对EID 3对BAT 3-EP 300介导的TRP 53乙酰化的拮抗作用进行生化分析。我接近K99/R 00奖的假设，SMC 5/6作为一个修复/监视复合物协调DNA修复和TRP 53介导的DNA损伤反应。这一功能确保了减数分裂前期向中期I过渡时的保真度。

DESCRIPTION (provided by applicant): We know a perturbed DNA damage response during meiosis will result in infertility, pregnancy loss or genetic defects. However, we know very little about the regulation of the DNA damage response during meiosis. My preliminary data implies that the SMC5/6 complex is required to mediate the DNA damage response during meiotic cell cycle progression. Because the SMC5/6 complex is essential for cell viability, in vivo experiments in mammals have not been performed and our understanding of the function of SMC5/6 in mammals is limited. I am creating a germ cell-specific mutation of Smc5 and mutating the testis specific SMC5/6 gene Eid3. These mutants will enable me to perform the first comprehensive studies of SMC5/6 complex function in meiosis (Aim1). EID3 is a testis-specific kleisin subunit of the SMC5/6 complex. However, the somatic cell kleisin SMC5/6 subunit, NSE4, is also expressed within the testis. Therefore, there are two SMC5/6 complexes present within the testis, SMC5/6NSE4 and SMC5/6EID3. I will determine the similarities and differences between the two complexes (Aim 2). I have confirmed that the SMC5/6 complex interacts with the BAT3-EP300 complex. The BAT3-EP300 complex is required for the activation of a TRP53-mediated DNA damage response. According to the protein interaction network developed between the SMC5/6 and the BAT3-EP300 complexes, SMC5/6 emerges as a key component of the DNA damage response pathway, by regulating TRP53 acetylation. In Aim 3, I will conduct a detailed assessment of the interaction between SMC5/6 and BAT3-EP300, together with biochemical analysis of the antagonistic function of EID3 on BAT3-EP300-mediated TRP53 acetylation. I approach the K99/R00 award with the hypothesis that SMC5/6 acts as a repair/surveillance complex coordinating DNA repair and the TRP53-mediated DNA damage response. This function ensures fidelity at the prophase to metaphase I transition of meiosis.