Ranieri Danilo, Belleudi Francesca, Magenta Alessandra, Torrisi Maria Rosaria
Istituto Pasteur-Fondazione Cenci Bolognetti, Dipartimento di Medicina Clinica e Molecolare, Sapienza Università di Roma, Italy.
Int J Cancer. 2015 Jul 1;137(1):61-72. doi: 10.1002/ijc.29373. Epub 2014 Dec 13.
The E5 oncoprotein of the human papillomavirus type 16 (HPV16 E5) deregulates epithelial homeostasis through the modulation of receptor tyrosine kinases and their signaling. Accordingly, the fibroblast growth factor receptor 2b (FGFR2b/KGFR), epithelial splicing transcript variant of the FGFR2, is down-modulated by the viral protein expression, leading to impairment of keratinocyte differentiation. Here, we report that, in cell models of transfected human keratinocytes as well as in cervical epithelial cells containing episomal HPV16, the down-regulation of FGFR2b induced by 16E5 is associated with the aberrant expression of the mesenchymal FGFR2c isoform as a consequence of splicing switch: in fact, quantitative RT-PCR analysis showed that this molecular event is transcriptionally regulated by the epithelial splicing regulatory proteins 1 and 2 (ESRP1 and ESRP2) and is able to produce effects synergistic with those caused by TGFβ treatment. Immunofluorescence analysis revealed that this altered FGFR2 splicing leads to changes in the specificity for the ligands FGFs and in the cellular response, triggering epithelial-mesenchymal transition (EMT). Through 16E5 or FGFR2 silencing as well as inhibition of FGFR2 activity we demonstrated the direct role of the viral protein in the receptor isoform switching and EMT, suggesting that these early molecular events during HPV infection might represent additional mechanisms driving cervical transformation and tumor progression.
人乳头瘤病毒16型(HPV16)的E5癌蛋白通过调节受体酪氨酸激酶及其信号传导来破坏上皮细胞的稳态。因此,成纤维细胞生长因子受体2b(FGFR2b/KGFR),即FGFR2的上皮剪接转录变体,会因病毒蛋白表达而下调,导致角质形成细胞分化受损。在此,我们报告,在转染的人角质形成细胞的细胞模型以及含有游离型HPV16的宫颈上皮细胞中,16E5诱导的FGFR2b下调与间充质FGFR2c异构体的异常表达相关,这是剪接转换的结果:事实上,定量RT-PCR分析表明,这一分子事件受上皮剪接调节蛋白1和2(ESRP1和ESRP2)的转录调控,并且能够产生与TGFβ处理所引起的效应协同的作用。免疫荧光分析显示,这种FGFR2剪接改变导致对配体FGFs的特异性改变以及细胞反应改变,从而引发上皮-间质转化(EMT)。通过16E5或FGFR2沉默以及FGFR2活性抑制,我们证明了病毒蛋白在受体异构体转换和EMT中的直接作用,表明HPV感染期间的这些早期分子事件可能代表了驱动宫颈转化和肿瘤进展的额外机制。