Study on biosynthesis of laminin-5 and keratinocyte attachment

层粘连蛋白-5是皮肤基底膜区锚定细丝的一种成分，对角质形成细胞的附着至关重要。大疱性交界性表皮松解症(JEB)是一种常染色体隐性遗传性皮肤水疱病，有致死性和非致死性两种形式，大多数患者都有层粘连蛋白-5缺陷。我们分析了6例JEB患者中层粘连蛋白-5异源三聚体的突变位置、基因表达水平和蛋白质链组装，以确定遗传损伤类型如何影响JEB的病理生理学。层粘连蛋白-5基因内的突变定位多样，最严重的形式JEB与提前终止密码子关联最好，而不是映射到任何特定的蛋白结构域。在所有6例JEB患者中，我们观察到的层粘连蛋白-5组装中间体与我们之前的工作预测的一样，表明α3β2Gamma2异源三聚体通过Beta2Gamma2异源二聚体在细胞内组装。由于组装过程是分泌过程，破坏组装所需的蛋白质-蛋白质中断的突变预计会限制层粘连蛋白-5的分泌，并可能干扰功能。然而，我们的数据表明，通常最严重的突变会降低mRNA的稳定性，并通过导致层粘连蛋白-5异三聚体组装所需的一条链的缺失(在非致命性的小鼠品种中)或完全缺失(在致命性的Herlitz-Jeb中)，作为区块链组装的功能性零等位基因。

Laminin-5 is a component of the anchoring filament in the basement membrane zone of the skin and crucial for keratinocyte attachment. Junctional epidermolysis bullosa (JEB) is an autosomal recessive skin blistering disease with both lethal and non-lethal forms, with most patients shown to have defects in laminin-5. We analyzed the location of mutations, gene expression levels, and protein chain assembly of laminin-5 heterotrimer in six JEB patients to determine how the type of genetic lesion influences the pathophsiology of JEB.Mutations within laminin-5 genes were diversely located, with the most severe forms of JEB correlating best with premature termination codons, rather than mapping to any particular protein domain. In all six JEB patients, the laminin-5 assembly intermediates we observed were as predicuted by our previous work indicating that the alpha3beta2gamma2 heterotrimer assembles intracellularly via a beta2gamma2 heterodimer intermediate. Since assembly procedes secretion, mutations that disrupt protein-protein inteructions needed for assembly are predicuted to limit the secretion of laminin-5, and likely to interfere with function. However, our data indicate that typically the most severe mutations diminish mRNA stabilty, and serve as functional null alleles that block chain assembly by resulting in either a deficiency (in the non-lethal mitis variety) or a complete abscence (in lethal Herlitz-JEB) of one of the chains needed for laminin-5 heterotrimer assembly.