Ku N O, Michie S, Oshima R G, Omary M B
Department of Medicine, VA Palo Alto Health Care System, California 94304, USA.
J Cell Biol. 1995 Dec;131(5):1303-14. doi: 10.1083/jcb.131.5.1303.
The two major intermediate filament proteins in glandular epithelia are keratin polypeptides 8 and 18 (K8/18). To evaluate the function and potential disease association of K18, we examined the effects of mutating a highly conserved arginine (arg89) of K18. Expression of K18 arg89-->his/cys and its normal K8 partner in cultured cells resulted in punctate staining as compared with the typical filaments obtained after expression of wild-type K8/18. Generation of transgenic mice expressing human K18 arg89-->cys resulted in marked disruption of liver and pancreas keratin filament networks. The most prominent histologic abnormalities were liver inflammation and necrosis that appeared at a young age in association with hepatocyte fragility and serum transaminase elevation. These effects were caused by the mutation since transgenic mice expressing wild-type human K18 showed a normal phenotype. A relative increase in the phosphorylation and glycosylation of detergent solubilized K8/18 was also noted in vitro and in transgenic animals that express mutant K18. Our results indicate that the highly conserved arg plays an important role in glandular keratin organization and tissue fragility as already described for epidermal keratins. Phosphorylation and glycosylation alterations in the arg mutant keratins may account for some of the potential changes in the cellular function of these proteins. Mice expressing mutant K18 provide a novel animal model for human chronic hepatitis, and for studying the tissue specific function(s) of K8/18.
腺上皮细胞中的两种主要中间丝蛋白是角蛋白多肽8和18(K8/18)。为了评估K18的功能及与潜在疾病的关联,我们研究了K18中一个高度保守的精氨酸(精氨酸89)发生突变的影响。与野生型K8/18表达后形成的典型丝状结构相比,K18精氨酸89突变为组氨酸/半胱氨酸及其正常的K8伴侣在培养细胞中的表达产生了点状染色。表达人K18精氨酸89突变为半胱氨酸的转基因小鼠导致肝脏和胰腺角蛋白丝网络明显破坏。最显著的组织学异常是肝脏炎症和坏死,在幼年时出现,伴有肝细胞脆性增加和血清转氨酶升高。这些影响是由突变引起的,因为表达野生型人K18的转基因小鼠表现出正常表型。在体外以及表达突变型K18的转基因动物中,还发现去污剂可溶解的K8/18的磷酸化和糖基化相对增加。我们的结果表明,这个高度保守的精氨酸在腺角蛋白组织和组织脆性中起重要作用,这与表皮角蛋白的情况已被描述的一致。精氨酸突变角蛋白中的磷酸化和糖基化改变可能解释了这些蛋白质细胞功能的一些潜在变化。表达突变型K18的小鼠为人类慢性肝炎以及研究K8/18的组织特异性功能提供了一种新的动物模型。