Novel mechanism underlying fibrinogen biogenesis in the endoplasmic reticulum

大量的疾病现在被认为是“构象疾病”，由蛋白质错误折叠和随后的聚集引起。一个例子是肝纤维蛋白原储存病（HFSD），其中根本原因是纤维蛋白原突变体的内质网（ER）滞留和聚集，导致肝损伤、低纤维蛋白原血症和过度出血。然而，ER中野生型和突变型纤维蛋白原的生物发生和质量控制的分子基础仍然未知。在本申请的初步数据中，我们偶然发现纤维蛋白原的生物合成受主要ER质量控制机制的Sel 1 L-Hrd 1蛋白复合物ER相关降解（ERAD）的调节。Sel 1 L-Hrd 1 ERAD代表了ERAD进化上最保守的分支，靶向错误折叠的ER蛋白，用于胞浆蛋白酶体降解。肝细胞特异性Sel 1 L缺陷小鼠表现出肝细胞ER中含有纤维蛋白原的包涵体，肝损伤和低纤维蛋白原血症，类似于HFSD的人类患者。事实上，纤维蛋白原的野生型和致病突变体都被Sel 1 L-Hrd 1 ERAD降解。这些数据表明肝细胞Sel 1 L-Hrd 1 ERAD在纤维蛋白原生物合成和肝内稳态中的关键作用。因此，本申请的首要假设是Sel 1 L-Hrd 1 ERAD是纤维蛋白原生物合成、凝血和肝脏稳态的关键调节机制，其通过靶向错误折叠的野生型或突变型纤维蛋白原蛋白进行蛋白酶体降解。ERAD对纤维蛋白原的这种作用防止了肝脏中有毒纤维蛋白原内含物和聚集体的形成。我们将实现两个目标：（1）阐明肝脏Sel 1 L-Hrd 1 ERAD在凝血和纤维蛋白原生物合成中的意义和分子机制;（2）阐明Sel 1 L-Hrd 1 ERAD在HFSD中的病理意义。这些研究的完成不仅将阐明ERAD在纤维蛋白原早期生物发生中的意义和分子机制，而且对其他与蛋白质错误折叠相关的构象疾病的机制和治疗策略具有更广泛的影响。

A large number of diseases are now recognized as ‘conformational diseases’, caused by protein misfolding and subsequent aggregation. One example is hepatic fibrinogen storage disease (HFSD), where the underlying cause is the endoplasmic reticulum (ER) retention and aggregation of fibrinogen mutants, leading to liver damage, hypofibrinogenemia, and excessive bleeding. However, the molecular events underlying the biogenesis and quality control of, both wildtype and mutant fibrinogen, in the ER remain unknown. In the preliminary data of this application, we serendipitously found that the biogenesis of fibrinogen is regulated by the Sel1L-Hrd1 protein complex of the principal ER quality-control machinery, ER-associated degradation (ERAD). Sel1L-Hrd1 ERAD represents the most evolutionarily conserved branch of ERAD and targets misfolded ER proteins for cytosolic proteasomal degradation. Hepatocyte-specific Sel1L-deficient mice exhibit fibrinogen-containing inclusions in the ER of hepatocytes, hepatic damage and hypofibrinogenemia, resembling human patients with HFSD. Indeed, both wildtype and disease-causing mutants of fibrinogens are degraded by Sel1L-Hrd1 ERAD. These data point to a critical role of hepatocyte Sel1L-Hrd1 ERAD in fibrinogen biogenesis and live homeostasis. Hence, the overarching hypothesis of this application is that Sel1L-Hrd1 ERAD is a critical regulatory mechanism for fibrinogen biogenesis, coagulation and liver homeostasis by targeting misfolded, either wildtype or mutant, fibrinogen proteins for proteasomal degradation. This action of ERAD towards fibrinogen prevents the formation of toxic fibrinogen inclusions and aggregates in the liver. We will accomplish two Aims: (1) Demonstrate the significance and molecular mechanism of hepatic Sel1L-Hrd1 ERAD in coagulation and fibrinogen biogenesis; and (2) Delineate the pathological importance of Sel1L-Hrd1 ERAD in HFSD. Completion of these studies will not only delineate the significance and molecular mechanism underlying the ERAD function in early biogenesis of fibrinogen, but also have broader impact on the mechanism and therapeutical strategy for other conformational diseases associated with protein misfolding.