Integrin α3β1 signaling through MEK/ERK determines alternative polyadenylation of the MMP-9 mRNA transcript in immortalized mouse keratinocytes.
Integrin α3β1 signaling through MEK/ERK determines alternative polyadenylation of the MMP-9 mRNA transcript in immortalized mouse keratinocytes.
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DOI：
10.1371/journal.pone.0119539
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发表时间：
2015
期刊：
影响因子：
3.7
通讯作者：
DiPersio CM
中科院分区：
文献类型：
--
作者：
Missan DS;Mitchell K;Subbaram S;DiPersio CM
整合素α3β1在正常和致瘤性表皮角质形成细胞中高度表达，在正常和病理组织重塑过程中，包括伤口愈合和鳞状细胞癌（SCC）的发展，它调节控制细胞功能和细胞外基质重塑的基因。先前的研究确定了α3β1在永生化和转化的角质形成细胞中在促进肿瘤发生、侵袭和促血管生成对内皮细胞的串扰的基因调节中的作用。其中一个基因，基质金属蛋白酶-9（MMP-9），通过增强mRNA稳定性的转录后机制被α3β1诱导。在本研究中，我们试图研究α3β1控制MMP-9 mRNA稳定性的机制。首先，我们利用荧光素酶报告基因测定来显示位于MMP-9 mRNA的3 '-非翻译区（3'-UTR）内的富含AU的元件（战神）以不依赖于α3β1的方式使转录物不稳定。接下来，我们克隆了MMP-9 mRNA的截短变体，其通过使用替代的上游多聚腺苷酸化信号产生，并且缺少含有去稳定战神的3 '-UTR区域。使用RNA酶保护测定来区分“长”（全长3 '-UTR）和“短”（截短的3'-UTR）MMP-9 mRNA变体，我们证明，与α3β1缺陷（即，α3-空）角质形成细胞。此外，我们确定永生化角质形成细胞获得了α3β1依赖性替代多聚腺苷酸化，因为原代新生角质形成细胞在长转录本和短转录本中没有表现出α3β1依赖性差异。最后，药理学抑制表达α3β1的角质形成细胞中的细胞外信号调节激酶（ERK）/促分裂原活化蛋白激酶（MAPK）通路导致向长变体表达的转变，而Raf-1介导的α3缺失角质形成细胞中ERK的活化显著增强短变体表达，表明ERK/MAPK信号传导在α3β1介导的近端多聚腺苷酸化位点选择中的作用。这些发现确定了整合素α3β1介导的基因调控的一种新模式，通过选择性多聚腺苷酸化。
Integrin α3β1 is highly expressed in both normal and tumorigenic epidermal keratinocytes where it regulates genes that control cellular function and extracellular matrix remodeling during normal and pathological tissue remodeling processes, including wound healing and development of squamous cell carcinoma (SCC). Previous studies identified a role for α3β1 in immortalized and transformed keratinocytes in the regulation of genes that promote tumorigenesis, invasion, and pro-angiogenic crosstalk to endothelial cells. One such gene, matrix metalloproteinase-9 (MMP-9), is induced by α3β1 through a post-transcriptional mechanism of enhanced mRNA stability. In the current study, we sought to investigate the mechanism through which α3β1 controls MMP-9 mRNA stability. First, we utilized a luciferase reporter assay to show that AU-rich elements (AREs) residing within the 3’-untranslated region (3’-UTR) of the MMP-9 mRNA renders the transcript unstable in a manner that is independent of α3β1. Next, we cloned a truncated variant of the MMP-9 mRNA which is generated through usage of an alternative, upstream polyadenylation signal and lacks the 3’-UTR region containing the destabilizing AREs. Using an RNase protection assay to distinguish “long” (full-length 3’-UTR) and “short” (truncated 3’-UTR) MMP-9 mRNA variants, we demonstrated that the shorter, more stable mRNA that lacks 3’-UTR AREs was preferentially generated in α3β1-expressing keratinocytes compared with α3β1-deficient (i.e., α3-null) keratinocytes. Moreover, we determined that α3β1-dependent alternative polyadenylation was acquired by immortalized keratinocytes, as primary neonatal keratinocytes did not display α3β1-dependent differences in the long and short transcripts. Finally, pharmacological inhibition of the extracellular signal-regulated kinase (ERK)/mitogen-activated protein kinase (MAPK) pathway in α3β1-expressing keratinocytes caused a shift towards long variant expression, while Raf-1-mediated activation of ERK in α3-null keratinocytes dramatically enhanced short variant expression, indicating a role for ERK/MAPK signaling in α3β1-mediated selection of the proximal polyadenylation site. These findings identify a novel mode of integrin α3β1-mediated gene regulation through alternative polyadenylation.
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影响因子：
4.8
作者：
Huwiler, A;Akool, ES;Eberhardt, W
通讯作者：
Eberhardt, W
影响因子：
8
作者：
Khabar, Khalid S. A.
通讯作者：
Khabar, Khalid S. A.
DOI：
10.1073/pnas.0900028106
发表时间：
2009-04-28
影响因子：
11.1
作者：
Ji, Zhe;Lee, Ju Youn;Tian, Bin
通讯作者：
Tian, Bin
影响因子：
3.7
作者：
Ernst, Jason;Ghanem, Louis;Steinman, Richard A.
通讯作者：
Steinman, Richard A.
影响因子：
14.9
作者：
Dai W;Zhang G;Makeyev EV
通讯作者：
Makeyev EV