The DNA helicase FANCJ (BRIP1) functions in Double Strand Break repair processing, but not crossover formation during Prophase I of meiosis in male mice.
The DNA helicase FANCJ (BRIP1) functions in Double Strand Break repair processing, but not crossover formation during Prophase I of meiosis in male mice.
复制标题
DNA 解旋酶 FANCJ (BRIP1) 在雄性小鼠减数分裂前期 I 期间的双链断裂修复过程中起作用，但在交叉形成中不起作用。
DOI：
10.1101/2023.10.06.561296
复制
发表时间：
2023
期刊：
影响因子：
--
通讯作者：
Cohen,PaulaE
中科院分区：
文献类型：
--
作者：
Horan,TeganS;Ascenção,CarollineFR;Mellor,ChristopherA;Wang,Meng;Smolka,MarcusB;Cohen,PaulaE
同源染色体之间的减数分裂重组是由数百条程序性双链断裂（dsb）的形成引发的。大约10%的dsb导致交叉（COs），同源物之间的物理DNA交换位点对纠正染色体分离至关重要。几乎所有的CO都是由MSH4/MSH5和MLH1/MLH3异源二聚体协同作用形成的，后者代表CO位点的定义标记。CO数量和位置的调控尚不清楚，但无疑需要多种修复途径的协同作用。在之前的报道中，我们发现DNA解旋酶FANCJ （BRIP1/BACH1）的基因陷阱破坏会引起MLH1灶和交叉的数量增加。在体细胞中，FANCJ与包括MLH1在内的许多DNA修复蛋白相互作用，我们假设FANCJ与MLH1一起调节主要的CO通路。为了进一步阐明FANCJ的减数分裂功能，我们通过CRISPR/Cas9基因编辑获得了三个新的FANCJ突变小鼠系：全基因缺失、n端解旋酶结构域截断和c端双标记等位基因。我们还生成了一种针对小鼠FANCJ蛋白c端的抗体。令人惊讶的是，我们的所有幻想突变体都没有显示出在厚膜期MLH1的病灶计数或在前期1期的总CO数发生任何变化。我们发现证据表明，FANCJ和MLH1在减数分裂中不相互作用；此外，FANCJ在减数分裂中不与MSH4、MLH1或MLH3共定位。相反，FANCJ与BRCA1和TOPBP1共定位，在减数分裂前期I早期开始沿着染色体核心形成离散的灶，在颧瘤晚期密集定位于未突触染色体轴，在厚纤维瘤早期密集定位于XY染色体。异型突变体在肺肿中也表现出微妙的dsb持久性。总的来说，这些数据表明FANCJ在早期DSB修复中起作用，但它们排除了FANCJ在mlh1介导的CO事件中的作用。
Meiotic recombination between homologous chromosomes is initiated by the formation of hundreds of programmed double-strand breaks (DSBs). Approximately 10% of these DSBs result in crossovers (COs), sites of physical DNA exchange between homologs that are critical to correct chromosome segregation. Virtually all COs are formed by coordinated efforts of the MSH4/MSH5 and MLH1/MLH3 heterodimers, the latter representing the defining marks of CO sites. The regulation of CO number and position is poorly understood, but undoubtedly requires the coordinated action of multiple repair pathways. In a previous report, we found gene-trap disruption of the DNA helicase, FANCJ (BRIP1/BACH1), elicited elevated numbers of MLH1 foci and chiasmata. In somatic cells, FANCJ interacts with numerous DNA repair proteins including MLH1, and we hypothesized that FANCJ functions with MLH1 to regulate the major CO pathway. To further elucidate the meiotic function of FANCJ, we produced three newFancjmutant mouse lines via CRISPR/Cas9 gene editing: a full-gene deletion, truncation of the N-terminal Helicase domain, and a C-terminal dual-tagged allele. We also generated an antibody against the C-terminus of the mouse FANCJ protein. Surprisingly, none of ourFancjmutants show any change in either MLH1 focus counts during pachynema or total CO number at diakinesis of prophase I. We find evidence that FANCJ and MLH1 do not interact in meiosis; further, FANCJ does not co-localize with MSH4, MLH1, or MLH3 in meiosis. Instead, FANCJ co-localizes with BRCA1 and TOPBP1, forming discrete foci along the chromosome cores beginning in early meiotic prophase I and densely localized to unsynapsed chromosome axes in late zygonema and to the XY chromosomes in early pachynema.Fancjmutants also exhibit a subtle persistence of DSBs in pachynema. Collectively, these data indicate a role for FANCJ in early DSB repair, but they rule out a role for FANCJ in MLH1-mediated CO events.