Knockout of DNase1l1l abrogates lens denucleation process and causes cataract in zebrafish
Knockout of DNase1l1l abrogates lens denucleation process and causes cataract in zebrafish
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DNase1l1l 的敲除会破坏斑马鱼晶状体去核过程并导致白内障
DOI：
10.1016/j.bbadis.2020.165724
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发表时间：
2020
影响因子：
6.2
通讯作者：
Hu Yanzhong
中科院分区：
文献类型：
--
作者：
Zhang Jing;Cui Wen-Wen;Du Chunxiao;Huang Yuwen;Pi Xiahui;Guo Wenya;Wang Jungai;Huang Weikang;Chen Danling;Li Jing;Li Hui;Zhang Jun;Ma Yuanfang;Mu Hongmei;Zhang Shuman;Liu Mugen;Cui Xiukun;Hu Yanzhong
在透镜发育过程中，无细胞器区（OFZ）的形成需要去除透镜纤维细胞中的细胞核。核DNA降解缺陷导致白内障形成。DNase 2 β在小鼠透镜分化过程中降解透镜纤维细胞核DNA。热休克因子4是透镜中的主要热休克转录因子，促进了透镜的分化。在小鼠和斑马鱼中敲除Hsf 4导致透镜发育缺陷，其特征在于核保留在透镜纤维细胞中。在以前的体外研究中，我们发现Hsf 4促进人和小鼠透镜上皮细胞中DNase 2 β的表达。在本研究中，发现DNase 1 l1 l代替DNase 2 β在斑马鱼透镜中唯一表达，而在Hsf 4 −/−斑马鱼透镜中不存在。利用CRISPR-Cas9技术构建了DNase 1111基因敲除斑马鱼品系，该品系发生了白内障。DNase 1111的缺失完全阻断了透镜初级和次级纤维细胞的去核过程，而对其他细胞器的清除影响不大。在Hsf 4 −/−斑马鱼透镜中，DNase 1 l1 l的转录调控显著受损。将DNase 1 l11 mRNA拯救到Hsf 4 −/−斑马鱼胚胎中减轻了其在透镜纤维细胞去核中的缺陷。我们的实验结果表明DNase 1 l1 l是透镜纤维细胞中负责核DNA降解的主要DNase，Hsf 4可以转录激活DNase 1 l1 l在斑马鱼中的表达。
Removal of nuclei in lens fiber cells is required for organelle-free zone (OFZ) formation during lens development. Defect in degradation of nuclear DNA leads to cataract formation. DNase2β degrades nuclear DNA of lens fiber cells during lens differentiation in mouse. Hsf4 is the principal heat shock transcription factor in lens and facilitates the lens differentiation. Knockout of Hsf4 in mouse and zebrafish resulted in lens developmental defect that was characterized by retaining of nuclei in lens fiber cells. In previousin vitrostudies, we found that Hsf4 promoted DNase2β expression in human and mouse lens epithelial cells. In this study, it was found that, instead of DNase2β, DNase1l1l is uniquely expressed in zebrafish lens and was absent in Hsf4−/−zebrafish lens. Using CRISPR-Cas9 technology, a DNase1l1l knockout zebrafish line was constructed, which developed cataract. Deletion of DNase1l1l totally abrogated lens primary and secondary fiber cell denucleation process, whereas had little effect on the clearance of other organelles. The transcriptional regulation of DNase1l1l was dramatically impaired in Hsf4−/−zebrafish lens. Rescue of DNase1l1l mRNA into Hsf4−/−zebrafish embryos alleviated its defect in lens fiber cell denucleation. Our resultsin vivodemonstrated that DNase1l1l is the primary DNase responsible for nuclear DNA degradation in lens fiber cells, and Hsf4 can transcriptionally activate DNase1l1l expression in zebrafish.