Vimentin Phosphorylation Underlies Myofibroblast Sensitivity to Withaferin A In Vitro and during Corneal Fibrosis.
Vimentin Phosphorylation Underlies Myofibroblast Sensitivity to Withaferin A In Vitro and during Corneal Fibrosis.
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波形蛋白的磷酸化是肌纤维细胞对伴脂蛋白A体外和角膜纤维化期间的敏感性的基础。
DOI：
10.1371/journal.pone.0133399
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发表时间：
2015
期刊：
影响因子：
3.7
通讯作者：
Mohan R
中科院分区：
文献类型：
--
作者：
Bargagna-Mohan P;Lei L;Thompson A;Shaw C;Kasahara K;Inagaki M;Mohan R
波形蛋白是治疗角膜纤维化的新靶点。使用原代兔角膜成纤维细胞和肌成纤维细胞，我们发现肌成纤维细胞不同于成纤维细胞，表现出细胞伸展和细胞极化受损，这与可溶性丝氨酸-38磷酸化波形蛋白(PSer38Vim)水平的增加有关。这种pSer38Vim亚型在细胞扩散过程中被低效地结合到肌成纤维细胞生长的波形蛋白中间丝(IFs)中，因此，与成纤维细胞相比，肌成纤维细胞保持了更高的可溶性pSer38Vim水平。此外，可溶性Vimentin靶向小分子和纤维化抑制因子黄曲霉毒素A(WFA)通过靶向可溶性pSer38Vim的过度磷酸化，有效地阻断了细胞在肌成纤维细胞中的选择性扩散。WFA处理不会诱导成纤维细胞中波形蛋白过度磷酸化。在WFA处理的肌成纤维细胞中，这种过度磷酸化的pSer38Vim物种与接头蛋白细丝蛋白A(Flna)形成复合体，当这些复合体从局部粘连中移位时，看起来像是短的蠕动。细胞外信号调节激酶(ERK)也被WFA磷酸化(PERK)，但令人惊讶的是，PERK不进入细胞核，但在细胞质复合体中仍与pSer38Vim结合。使用角膜碱损伤模型，我们发现野生型小鼠的纤维化角膜具有高水平的PERK表达，而波形蛋白缺陷(Vim KO)小鼠受损的角膜愈合后纤维化程度降低，PERK表达显著降低。最后，WFA治疗导致野生型小鼠愈合的角膜中PERK和pSer38Vim的表达减少。综上所述，这些发现确定了pSer38Vim作为肌肉成纤维细胞对WFA敏感性的重要决定因素的一个迄今未被认识的作用。
Vimentin is a newly recognized target for corneal fibrosis. Using primary rabbit corneal fibroblasts and myofibroblasts we show that myofibroblasts, unlike fibroblasts, display impaired cell spreading and cell polarization, which is associated with increased levels of soluble serine-38 phosphorylated vimentin (pSer38Vim). This pSer38Vim isoform is inefficiently incorporated into growing vimentin intermediate filaments (IFs) of myofibroblasts during cell spreading, and as a result, myofibroblasts maintain higher soluble pSer38Vim levels compared to fibroblasts. Moreover, the soluble vimentin-targeting small molecule and fibrotic inhibitor withaferin A (WFA) causes a potent blockade of cell spreading selectively in myofibroblasts by targeting soluble pSer38Vim for hyperphosphorylation. WFA treatment does not induce vimentin hyperphosphorylation in fibroblasts. This hyperphosphorylated pSer38Vim species in WFA-treated myofibroblasts becomes complexed with adaptor protein filamin A (FlnA), and these complexes appear as short squiggles when displaced from focal adhesions. The extracellular-signal regulated kinase (ERK) is also phosphorylated (pERK) in response to WFA, but surprisingly, pERK does not enter the nucleus but remains bound to pSer38Vim in cytoplasmic complexes. Using a model of corneal alkali injury, we show that fibrotic corneas of wild type mice possess high levels of pERK, whereas injured corneas of vimentin-deficient (Vim KO) mice that heal with reduced fibrosis have highly reduced pERK expression. Finally, WFA treatment causes a decrease in pERK and pSer38Vim expression in healing corneas of wild type mice. Taken together, these findings identify a hereto-unappreciated role for pSer38Vim as an important determinant of myofibroblast sensitivity to WFA.