ZMPSTE24 Missense Mutations that Cause Progeroid Diseases Decrease Prelamin A Cleavage Activity and/or Protein Stability
ZMPSTE24 Missense Mutations that Cause Progeroid Diseases Decrease Prelamin A Cleavage Activity and/or Protein Stability
复制标题
导致早衰性疾病的 ZMPSTE24 错义突变会降低 Prelamin A 裂解活性和/或蛋白质稳定性
DOI：
10.1101/198069
复制
发表时间：
2017
期刊：
影响因子：
--
通讯作者：
Spear E
中科院分区：
文献类型：
--
作者：
Spear E
锌金属蛋白酶ZMPSTE 24是一种完整的膜蛋白，对核支架蛋白核纤层蛋白A（由LMNA基因编码）的生物发生至关重要。核纤层蛋白A作为前体核纤层蛋白A合成，其经历其C-末端CAAX基序的翻译后修饰，包括法尼基化和羧基甲基化，然后通过ZMPSTE 24蛋白水解去除修饰的C-末端15个氨基酸。LMNA或ZMPSTE 24中阻碍前层蛋白A裂解的突变导致一系列相关的早老性疾病，包括过早衰老障碍Hutchinson-Gilford早衰综合征（HGPS）、下颌骨肢端发育不良B型（MAD-B）和限制性皮肤病（RD），这是由于前层蛋白A的永久法尼基化形式的积累所致。在这里，我们已经开发了一种体内试验，以检查ZMPSTE 24依赖的裂解prelamin A在酵母酿酒酵母，一种生物体，缺乏核纤层。我们证明，prelamin A加工在酵母中具有相同的要求，在哺乳动物细胞中，包括法尼基化的CAAX基序半胱氨酸，一个完整的底物切割位点，和ZMPSTE 24酶活性。使用这种“人源化酵母”系统来检查八种已知的与疾病相关的ZMPSTE 24错义突变，所有突变都显示出减少的前层蛋白A加工，并分为三类，活性缺陷（I类），稳定性缺陷（II类），或两者兼而有之（III类）。II类ZMPSTE 24突变体可以通过删除E3泛素连接酶Doa 10来拯救，Doa 10是许多错误折叠的膜蛋白的ER相关蛋白降解的关键因子，这可能具有治疗意义。我们还表明，ZMPSTE 24介导的前层蛋白A裂解可以从最近发现的ZMPSTE 24在清除ER膜translocon阻塞底物中的作用中解偶联。与ZMPSTE 24的晶体结构一起，我们的“人源化酵母系统”可以指导结构-功能研究，以揭示前层蛋白A切割，转位子疏通以及膜蛋白折叠和稳定性的机制。
The zinc metalloprotease ZMPSTE24 is an integral membrane protein critical for the biogenesis of the nuclear scaffold protein lamin A (encoded by theLMNAgene). Lamin A is synthesized as a precursor, prelamin A, that undergoes posttranslational modification of its C-terminal CAAX motif, including farnesylation and carboxyl methylation, followed by proteolytic removal of the modified C-terminal 15 amino acids by ZMPSTE24. Mutations inLMNAorZMPSTE24that impede prelamin A cleavage cause a spectrum of related progeroid diseases, including the premature aging disorder Hutchinson-Gilford Progeria Syndrome (HGPS), mandibuloacral dysplasia type B (MAD-B), and restrictive dermopathy (RD), due to the accumulation of a permanently farnesylated form(s) of prelamin A. Here, we have developed anin vivoassay to examine the ZMPSTE24-dependent cleavage of prelamin A in the yeastSaccharomyces cerevisiae, an organism that lacks a nuclear lamina. We demonstrate that prelamin A processing has the same requirements in yeast as in mammalian cells, including farnesylation of the CAAX motif cysteine, an intact substrate cleavage site, and ZMPSTE24 enzymatic activity. Using this “ humanized yeast” system to examine the eight known disease-associatedZMPSTE24missense mutations, all show diminished prelamin A processing and fall into three classes, with defects in activity (Class I), stability (Class II), or both (Class III). Class II ZMPSTE24 mutants can be rescued by deleting the E3 ubiquitin ligase Doa10, a key factor in ER-associated protein degradation of many misfolded membrane proteins, which may have therapeutic implications. We also show that ZMPSTE24-mediated prelamin A cleavage can be uncoupled from the recently discovered role of ZMPSTE24 in the clearance of ER membrane translocon-clogged substrates. Together with the crystal structure of ZMPSTE24, our “ humanized yeast system” can guide structure-function studies to uncover the mechanisms of prelamin A cleavage, translocon unclogging, and membrane protein folding and stability.