The Wnt5A/protein kinase C pathway mediates motility in melanoma cells via the inhibition of metastasis suppressors and initiation of an epithelial to mesenchymal transition
The Wnt5A/protein kinase C pathway mediates motility in melanoma cells via the inhibition of metastasis suppressors and initiation of an epithelial to mesenchymal transition
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DOI：
10.1074/jbc.m700075200
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发表时间：
2007-06-08
影响因子：
4.8
通讯作者：
Weeraratna, Ashani T.
中科院分区：
文献类型：
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作者：
Dissanayake, Samudra K.;Wade, Michael;Weeraratna, Ashani T.
我们已经表明，Wnt 5A增加黑色素瘤细胞的运动性。为了探索涉及Wnt 5A的细胞途径，我们通过微阵列分析比较了WNT 5A的功能获得（WNT 5A稳定转染子）与功能丧失（siRNA敲低）。增加WNT 5A抑制了几个基因的表达，这些基因在小干扰RNA介导的WNT 5A敲低后重新表达。受WNT 5A影响的基因包括KISS-1（一种转移抑制因子）和CD 44（参与转移期间肿瘤细胞归巢）。这可以使用小干扰RNA和重组Wnt 5A（rWnt 5A）在蛋白质水平上验证。在WNT 5A上调的基因中，有一个基因波形蛋白，与上皮细胞向间质细胞转化（EMT）有关，EMT涉及E-钙粘蛋白的减少，这是由于转录抑制因子Snail的上调。rWnt 5A处理增加Snail和波形蛋白表达，降低E-钙粘蛋白，即使在显性负性TCF 4存在下，表明这种激活不依赖于Wnt/β-连环蛋白信号。由于Wnt 5A可以通过蛋白激酶C（PKC）发出信号，因此还使用PKC抑制和活化研究来评估PKC在Wnt 5A介导的运动性和EMT中的作用。用佛波酯处理低水平表达Wnt 5A的细胞增加Snail表达，抑制高水平表达Wnt 5A的细胞中的PKC
We have shown that Wnt5A increases the motility of melanoma cells. To explore cellular pathways involving Wnt5A, we compared gain-of-function (WNT5A stable transfectants) versus loss-of-function (siRNA knockdown) of WNT5A by microarray analysis. Increasing WNT5A suppressed the expression of several genes, which were re-expressed after small interference RNA-mediated knockdown of WNT5A. Genes affected by WNT5A include KISS-1, a metastasis suppressor, and CD44, involved in tumor cell homing during metastasis. This could be validated at the protein level using both small interference RNA and recombinant Wnt5A (rWnt5A). Among the genes up-regulated by WNT5A was the gene vimentin, associated with an epithelial to mesenchymal transition (EMT), which involves decreases in E-cadherin, due to up-regulation of the transcriptional repressor, Snail. rWnt5A treatment increases Snail and vimentin expression, and decreases E-cadherin, even in the presence of dominant-negativeTCF4, suggesting that this activation is independent of Wnt/ss-catenin signaling. Because Wnt5A can signal via protein kinase C (PKC), the role of PKC in Wnt5A- mediated motility and EMT was also assessed using PKC inhibition and activation studies. Treating cells expressing low levels of Wnt5A with phorbol ester increased Snail expression inhibiting PKC in cells expressing high levels of Wnt5A