The Wilms Tumor Gene, Wt1, Maintains Testicular Cord Integrity by Regulating the Expression of Col4a1 and Col4a2
The Wilms Tumor Gene, Wt1, Maintains Testicular Cord Integrity by Regulating the Expression of Col4a1 and Col4a2
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肾母细胞瘤基因 Wt1 通过调节 Col4a1 和 Col4a2 的表达来维持睾丸索完整性
DOI：
10.1095/biolreprod.112.105379
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发表时间：
2013-03-01
影响因子：
3.6
通讯作者：
Liu, Yi-Xun
中科院分区：
文献类型：
--
作者：
Chen, Su-Ren;Chen, Min;Liu, Yi-Xun
Wt 1在发育中的睾丸支持细胞中特异性表达。先前的一项研究表明，Wt 1在维持睾丸索的完整性方面起着关键作用。然而，其潜在机制尚不清楚。在这项研究中，我们发现，层粘连蛋白阳性的基底层内衬睾丸索是破碎的，完全没有在某些地区的Wt 1（-/flox）; Amh-Cre睾丸，表明睾丸索中断可以归因于基底膜的破坏。为了探讨这种效应的分子机制，我们研究了细胞粘附分子（CAMs）和睾丸索基底层成分的实时RT-PCR，Western印迹和免疫染色的表达。与对照组相比，Wt 1（-/flox）; Amh-Cre睾丸中CAM（如E-cadherin、N-cadherin、claudin 11、occludin、beta-catenin和ZO-1）的表达无明显变化。然而，Col 4a 1和Col 4a 2的mRNA水平在Wt 1缺陷的睾丸中显著降低。免疫染色分析进一步证实，胶原蛋白IV蛋白水平显着降低Wt 1（-/flox）; Amh-Cre睾丸。此外，荧光素酶和点突变分析显示，Col 4a 1和Col 4a 2启动子被WT 1和SOX 9加成反式激活。鉴于这一发现和以前的结果表明，SOX 9表达下降迅速Wt 1删除后，我们得出结论，Wt 1的支持细胞的重要的基底层组件的下调，这反过来又会导致基底层和随后的睾丸索中断中断的损失。
Wt1 is specifically expressed in Sertoli cells in the developing testis. A previous study has demonstrated that Wt1 plays a critical role in maintaining the integrity of testicular cords. However, the underlying mechanism is unclear. In this study, we found that the laminin-positive basal lamina lining the testicular cords was fragmented and completely absent in some areas of Wt1(-/flox); Amh-Cre testes, indicating that the testicular cord disruption can be attributed to the breakdown of the basement membrane. To explore the molecular mechanism underlying this effect, we examined the expression of cell adhesion molecules (CAMs) and testicular cord basal lamina components by real-time RT-PCR, Western blotting, and immunostaining. Compared with control testes, the expression of CAMs (such as E-cadherin, N-cadherin, claudin11, occludin, beta-catenin, and ZO-1) was not obviously altered in Wt1(-/flox); Amh-Cre testes. However, the mRNA level of Col4a1 and Col4a2 was significantly decreased in Wt1-deficient testes. Immunostaining assays further confirmed that the collagen IV protein levels were dramatically reduced in Wt1(-/flox); Amh-Cre testes. Moreover, luciferase and point mutation analyses revealed that the Col4a1 and Col4a2 promoters were additively transactivated by WT1 and SOX9. Given this finding and previous results showing that SOX9 expression declines rapidly after Wt1 deletion, we conclude that the loss of Wt1 in Sertoli cells results in the downregulation of the important basal lamina component, which in turn causes the breakdown of the basal lamina and subsequent testicular cord disruption.