Conditional inactivation of the DNA damage response gene Hus1 in mouse testis reveals separable roles for components of the RAD9-RAD1-HUS1 complex in meiotic chromosome maintenance.
Conditional inactivation of the DNA damage response gene Hus1 in mouse testis reveals separable roles for components of the RAD9-RAD1-HUS1 complex in meiotic chromosome maintenance.
复制标题
DOI：
10.1371/journal.pgen.1003320
复制
发表时间：
2013
期刊：
影响因子：
4.5
通讯作者：
Weiss RS
中科院分区：
文献类型：
--
作者：
Lyndaker AM;Lim PX;Mleczko JM;Diggins CE;Holloway JK;Holmes RJ;Kan R;Schlafer DH;Freire R;Cohen PE;Weiss RS
RAD 9-RAD 1-HUS 1（9-1-1）复合物是一种异源三聚体的PCNA样钳，其通过促进DNA修复以及ATR依赖性DNA损伤检查点信号传导来响应体细胞中的DNA损伤。在酵母、蠕虫和果蝇中，9-1-1复合物也是减数分裂检查点功能和有效完成减数分裂重组所必需的;然而，由于Rad 9、Rad 1和Hus 1是哺乳动物的必需基因，因此对它们在哺乳动物生殖细胞中的功能知之甚少。在这项研究中，我们评估了9-1-1的减数分裂功能，通过分析小鼠生殖细胞特异性缺失的Hus 1，以及通过检查定位的RAD 9和RAD 1的减数分裂染色体在前期I。睾丸生殖细胞中的Hus 1缺失导致减数分裂缺陷、生殖细胞耗竭和生育能力严重受损。Hus 1缺陷的初级精母细胞表现出持续的常染色体γ H2 AX和RAD 51染色，表明未修复的减数分裂DSB，突触缺陷，扩展的XY体结构域通常包含部分或整个常染色体，以及染色体结构异常的增加，例如端对端X染色体-常染色体融合和联会复合体破裂。大多数这些畸变持续在二倍体阶段的精母细胞。与9-1-1复合物在减数分裂DSB修复中的作用一致，RAD 9以Hus 1依赖的方式定位于减数分裂染色体上的点状、含RAD 51的病灶。有趣的是，RAD 1有一个更广泛的分布，只有部分重叠的RAD 9，和本地化的RAD 1和ATR激活TOPBP 1的XY体和非突触常染色体是完整的Hus 1条件性敲除。我们的结论是，哺乳动物HUS 1作为一个典型的9-1-1复合物的组成部分，在减数分裂前期I，以促进DSB修复，并进一步提出，RAD 1和TOPBP 1响应unsynapsed染色质通过一种替代机制，不需要RAD 9或HUS 1。减数分裂是一种特殊的细胞分裂过程，其中生殖细胞经历两次细胞分裂以产生单倍体后代。两个过程，遗传重组和染色体配对/联会，是成功的减数分裂和具有高染色体完整性的配子的产生的关键。已经提出，RAD 9-RAD 1-HUS 1（9-1-1）复合物通过促进ATR检查点激酶的激活在重组以及在染色体突触的检查点依赖性监测中发挥关键作用。我们的数据表明，HUS 1是所需的正常生殖细胞发育和生育能力，有效地完成减数分裂DNA重组事件的子集，并从一个专门的，抑制性染色质结构域包含X和Y染色体的适当排除非性染色体。然而，HUS 1是不需要的ATR的减数分裂功能，在响应染色体突触缺陷。此外，RAD 1定位于沿着不联会的染色体，缺乏可检测的RAD 9的网站，并在没有Hus 1，这意味着RAD 1在一个新的响应非突触染色质是独立的典型的9-1-1复合物。由于生殖细胞中缺乏Hus 1的小鼠表现出染色体异常和生育能力严重降低，这项工作对维持生殖细胞中的基因组稳定性和人类生殖健康具有广泛的意义。
The RAD9-RAD1-HUS1 (9-1-1) complex is a heterotrimeric PCNA-like clamp that responds to DNA damage in somatic cells by promoting DNA repair as well as ATR-dependent DNA damage checkpoint signaling. In yeast, worms, and flies, the 9-1-1 complex is also required for meiotic checkpoint function and efficient completion of meiotic recombination; however, since Rad9, Rad1, and Hus1 are essential genes in mammals, little is known about their functions in mammalian germ cells. In this study, we assessed the meiotic functions of 9-1-1 by analyzing mice with germ cell-specific deletion of Hus1 as well as by examining the localization of RAD9 and RAD1 on meiotic chromosomes during prophase I. Hus1 loss in testicular germ cells resulted in meiotic defects, germ cell depletion, and severely compromised fertility. Hus1-deficient primary spermatocytes exhibited persistent autosomal γH2AX and RAD51 staining indicative of unrepaired meiotic DSBs, synapsis defects, an extended XY body domain often encompassing partial or whole autosomes, and an increase in structural chromosome abnormalities such as end-to-end X chromosome-autosome fusions and ruptures in the synaptonemal complex. Most of these aberrations persisted in diplotene-stage spermatocytes. Consistent with a role for the 9-1-1 complex in meiotic DSB repair, RAD9 localized to punctate, RAD51-containing foci on meiotic chromosomes in a Hus1-dependent manner. Interestingly, RAD1 had a broader distribution that only partially overlapped with RAD9, and localization of both RAD1 and the ATR activator TOPBP1 to the XY body and to unsynapsed autosomes was intact in Hus1 conditional knockouts. We conclude that mammalian HUS1 acts as a component of the canonical 9-1-1 complex during meiotic prophase I to promote DSB repair and further propose that RAD1 and TOPBP1 respond to unsynapsed chromatin through an alternative mechanism that does not require RAD9 or HUS1. Meiosis is a specialized cell division process in which germ cells undergo two cell divisions to produce haploid progeny. Two processes, genetic recombination and chromosome pairing/synapsis, are critical for successful meiosis and the production of gametes with high chromosomal integrity. The RAD9-RAD1-HUS1 (9-1-1) complex has been proposed to play critical roles in recombination as well as in the checkpoint-dependent monitoring of chromosomal synapsis by facilitating activation of the ATR checkpoint kinase. Our data indicate that HUS1 is required for normal germ cell development and fertility, for efficient completion of a subset of meiotic DNA recombination events, and for proper exclusion of the non-sex chromosomes from a specialized, repressive chromatin domain containing the X and Y chromosomes. However, HUS1 is not required for the meiotic functions of ATR in responding to chromosome synapsis defects. Furthermore, RAD1 localizes to sites along asynapsed chromosomes that lack detectable RAD9, and does so in the absence of Hus1, implicating RAD1 in a novel response to unsynapsed chromatin that is independent of the canonical 9-1-1 complex. Since mice lacking Hus1 in germ cells exhibit chromosomal abnormalities and severely reduced fertility, this work has broad implications for the maintenance of genome stability in the germline and for human reproductive health.
登录
查看更多内容
DOI：
10.1126/science.1203430
发表时间：
2011-06-10
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
Cotta-Ramusino C;McDonald ER 3rd;Hurov K;Sowa ME;Harper JW;Elledge SJ
通讯作者：
Elledge SJ
影响因子：
4.8
作者：
Burtelow, MA;Roos-Mattjus, PMK;Karnitz, LM
通讯作者：
Karnitz, LM
影响因子：
14.9
作者：
Guan, Xin;Bai, Haibo;Lu, A-Lien
通讯作者：
Lu, A-Lien
影响因子：
4
作者：
Abdu, Uri;Klovstad, Martha;Schupbach, Trudi
通讯作者：
Schupbach, Trudi
影响因子：
4.4
作者：
Hang, HY;Zhang, YZ;Lieberman, HB
通讯作者：
Lieberman, HB