Wilms tumor protein-dependent transcription of VEGF receptor 2 and hypoxia regulate expression of the testis-promoting gene Sox9 in murine embryonic gonads
Wilms tumor protein-dependent transcription of VEGF receptor 2 and hypoxia regulate expression of the testis-promoting gene Sox9 in murine embryonic gonads
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DOI：
10.1074/jbc.m117.816751
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发表时间：
2017-12-08
影响因子：
4.8
通讯作者：
Scholz, Holger
中科院分区：
文献类型：
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作者：
Kirschner, Karin M.;Sciesielski, Lina K.;Scholz, Holger
Wilms肿瘤蛋白1 （WT1）参与了性发育过程中几个基因的调控，但其在性腺形成中的功能尚不清楚。在这里，我们报道WT1刺激小鼠胚胎性腺中编码VEGF受体2的基因Kdr的表达。我们发现WT1和KDR在睾丸的支持细胞和胚胎卵巢的体细胞中共表达。体内morolinos介导的WT1敲低可降低培养胚胎性腺在多个发育阶段的Kdr转录本。此外，WT1与胚胎睾丸和卵巢染色质中的Kdr启动子结合。强制表达作为转录因子的WT1（-KTS）异构体会增加KDR mRNA水平，而可能在转录后水平起作用的WT1（+KTS）异构体则不会。ChIP表明WT1（-KTS）与KDR启动子结合，而WT1（+KTS）与KDR启动子不结合。用KDR酪氨酸激酶抑制剂SU1498或KDR配体VEGFA处理发现，KDR信号抑制胚胎XX性腺促睾丸基因Sox9。WT1敲除消除了su1498介导的KDR抑制对Sox9表达的刺激作用。暴露于1% O-2以模拟胚胎中的低氧条件，增加了Vegfa的表达，但不影响性腺外植体中Sox9 mRNA的水平。然而，在1%的O-2中，在SU1498的存在下孵育，可显著降低培养睾丸中的Sox9转录物，并增加卵巢中的Sox9水平。这些发现表明，局部氧环境和增强KDR表达的WT1都有助于小鼠性腺中Sox9的性别特异性表达。
Wilms tumor protein 1 (WT1) has been implicated in the control of several genes in sexual development, but its function in gonad formation is still unclear. Here, we report that WT1 stimulates expression of Kdr, the gene encoding VEGF receptor 2, in murine embryonic gonads. We found that WT1 and KDR are co-expressed in Sertoli cells of the testes and somatic cells of embryonic ovaries. Vivo-morpholino-mediated WT1 knockdown decreased Kdr transcripts in cultured embryonic gonads at multiple developmental stages. Furthermore, WT1 bound to the Kdr promoter in the chromatin of embryonic testes and ovaries. Forced expression of the WT1(-KTS) isoform, which functions as a transcription factor, increased KDR mRNA levels, whereas the WT1(+KTS) isoform, which acts presumably on the post-transcriptional level, did not. ChIP indicated that WT1(-KTS), but not WT1(+KTS), binds to the KDR promoter. Treatment with the KDR tyrosine kinase inhibitor SU1498 or the KDR ligand VEGFA revealed that KDR signaling represses the testis-promoting gene Sox9 in embryonic XX gonads. WT1 knockdown abrogated the stimulatory effect of SU1498-mediated KDR inhibition on Sox9 expression. Exposure to 1% O-2 to mimic the low-oxygen conditions in the embryo increased Vegfa expression but did not affect Sox9 mRNA levels in gonadal explants. However, incubation in 1% O-2 in the presence of SU1498 significantly reduced Sox9 transcripts in cultured testes and increased Sox9 levels in ovaries. These findings demonstrate that both the local oxygen environment and WT1, which enhances KDR expression, contribute to sex-specific Sox9 expression in developing murine gonads.