Molecular genetic dissection of epidermodysplasia verruciformis

描述(申请人提供)：疣状表皮发育不良(EV)是一种先天性皮肤病，特征是由β-乳头状瘤病毒(EV-HPV)引起的终生播散性和持续性扁平疣皮损。EV与非黑色素瘤皮肤癌(NMSC)的风险增加有关，但在大多数患者中没有其他临床症状(典型的EV)。Ever1和EveR2的病态突变是在2002年发现的。这些蛋白在T淋巴细胞和角质形成细胞中表达，T淋巴细胞参与清除HPV感染的上皮细胞，角质形成细胞是EV-HPV的特异性靶细胞。Ever蛋白与锌转运蛋白ZnT1形成复合体，调节细胞内锌的分布，控制锌依赖的转录活性。在这些患者中缺乏明显的T细胞表型和T细胞相关感染，这表明EV-HPV的易感性与角质形成细胞有关。以往的缺陷约占报告的典型EV病例的75%。我们最近在EV和其他临床特征(“非典型”EV)患者中发现了AR、RhoH和MST1缺陷，这是由于严重的T细胞缺陷所致。这些发现表明，T细胞缺陷可能是持续的EV-HPV感染的基础。为了更好地了解EV的发病机制，并破译EV-HPV免疫的分子和细胞机制，我们旨在利用GW连锁(GWL)和全外显组测序(WES)相结合的无假设、全基因组(GW)筛选方法来鉴定EV的新遗传病因。在我们的15个家系的队列中，我们获得了新的EV致病基因的强有力的初步证据，鉴定了来自两个哥伦比亚家系的6名患者和1名法国患者的CIB1纯合子突变。在这些患者的EBV-B细胞系中，钙和整合素结合蛋白1(CIB1)蛋白的表达缺失。CIB1缺陷患者表现为典型的EV。因此，我们的发现令人惊讶，因为CIB1普遍表达，其众多已知功能中没有一项与对EV-HPV的免疫有关。缺乏CIB1的小鼠表现出血栓形成、血管生成和精子生成受损。我们推测CIB1缺乏会损害角质形成细胞对EV-HPV的固有免疫。我们打算测试CIB1在健康对照组和CIB1缺陷患者的角质形成细胞和T细胞中的表达和功能。我们还将探索CIB1和EVER之间的潜在联系。最后，我们将研究EV-HPV在对照组和患者角质形成细胞中的生长情况。该项目具有很高的创新性，但完全可行，并且已经得到了强有力的初步证据的支持。从生物学的基本观点来看，这项研究将通过阐明CIB1在角质形成细胞中的作用，从而可能确定EV-HPV的细胞学基础，从而为EV-HPV的免疫机制提供相当大的洞察力。从临床的角度来看，对EV致病机制的剖析将为患者提供分子诊断和为家庭提供遗传咨询，并将为其他HPV驱动的病理研究铺平道路。

DESCRIPTION (provided by applicant): Epidermodysplasia verruciformis (EV) is a congenital dermatosis characterized by lifelong disseminated and persistent flat wart lesions caused by beta-papillomaviruses (EV-HPVs). EV is associated with an increase in the risk of non-melanoma skin carcinomas (NMSC), but there are no other clinical signs in most patients ("typical" EV). Morbid mutations in EVER1 and EVER2 were found in 2002. These proteins are expressed in T lymphocytes, which are involved in the clearance of HPV-infected epithelial cells, and in keratinocytes, the specific target cells of EV-HPVs. EVER proteins form a complex with the zinc transporter ZnT1 to regulate the intracellular Zn distribution and to control Zn-dependent transcriptional activity. The lack of pronounced T cell phenotype and T cell-related infections in these patients suggests that the mechanisms by which EVER deficiency confers susceptibility to EV-HPVs involves keratinocytes. EVER deficiencies account for approximately 75% of reported typical EV cases. We have recently identified AR RHOH and MST1 deficiencies in patients with EV and other clinical features ("atypical" EV) due to a profound T cell deficiency. These discoveries showed that a T-cell defect can underlie persistent EV-HPV infections. In order to better understand the pathogenesis of EV, and to decipher the molecular and cellular mechanisms of immunity against EV-HPVs, we aim to identify novel genetic etiologies of EV using a hypothesis-free, genome-wide (GW) screening approach combining GW linkage (GWL) and whole-exome sequencing (WES). In our cohort of 15 kindreds, we obtained strong preliminary evidence of a novel EV-causing gene, with the identification of homozygous mutations in CIB1 in 6 patients from two Colombian kindreds and in one French patient. In these patients' EBV-B cell lines, there is a loss of expression of the calcium and integrin binding protein 1 (CIB1) protein. The CIB1-deficient patients display a typical form of EV. Our findings are therefore surprising, as CIB1 is ubiquitously expressed and none of its numerous known functions is connected with immunity against EV- HPVs. Mice lacking CIB1 display impaired thrombosis, angiogenesis, and spermatogenesis. We hypothesize that CIB1 deficiency impairs keratinocyte intrinsic immunity against EV-HPV. We intend to test the expression and function of CIB1 in both keratinocytes and T cells from healthy controls and CIB1-deficient patients. We will also explore a potential connection between CIB1 and EVER. Finally, we will study the growth of EV-HPV in control and patients' keratinocytes. The project is highly innovative yet perfectly feasible, and already supported by strong preliminary evidence. From a basic biological standpoint, this research will provide considerable insight into the mechanisms of immunity against EV-HPV, notably by clarifying the role of CIB1 in keratinocytes, thereby possibly defining the cellular basis of EV. From a clinical standpoint, the dissection of the pathogenesis of EV will provide molecular diagnoses for patients and genetic counseling for families and will pave the way for the study of other HPV-driven pathologies.