Department of Cell Biology, The Johns Hopkins School of Medicine, Baltimore, MD 21205, USA.
Department of Chemistry, Purdue University, West Lafayette, IN 47907, USA.
Dis Model Mech. 2018 Jul 13;11(7):dmm033670. doi: 10.1242/dmm.033670.
The human zinc metalloprotease ZMPSTE24 is an integral membrane protein crucial for the final step in the biogenesis of the nuclear scaffold protein lamin A, encoded by After farnesylation and carboxyl methylation of its C-terminal CAAX motif, the lamin A precursor (prelamin A) undergoes proteolytic removal of its modified C-terminal 15 amino acids by ZMPSTE24. Mutations in or that impede this prelamin A cleavage step cause the premature aging disease Hutchinson-Gilford progeria syndrome (HGPS), and the related progeroid disorders mandibuloacral dysplasia type B (MAD-B) and restrictive dermopathy (RD). Here, we report the development of a 'humanized yeast system' to assay ZMPSTE24-dependent cleavage of prelamin A and examine the eight known disease-associated missense mutations. All mutations show diminished prelamin A processing and fall into three classes, with defects in activity, protein stability or both. Notably, some ZMPSTE24 mutants can be rescued by deleting the E3 ubiquitin ligase Doa10, involved in endoplasmic reticulum (ER)-associated degradation of misfolded membrane proteins, or by treatment with the proteasome inhibitor bortezomib. This finding may have important therapeutic implications for some patients. We also show that ZMPSTE24-mediated prelamin A cleavage can be uncoupled from the recently discovered role of ZMPSTE24 in clearance of ER membrane translocon-clogged substrates. Together with the crystal structure of ZMPSTE24, this humanized yeast system can guide structure-function studies to uncover mechanisms of prelamin A cleavage, translocon unclogging, and membrane protein folding and stability.
人类锌金属蛋白酶 ZMPSTE24 是一种膜内在蛋白，对于核骨架蛋白 lamin A 生物发生的最后一步至关重要，lamin A 由编码。在其 C 末端 CAAX 基序的法呢基化和羧基甲基化之后，lamin A 前体（prelamin A）通过 ZMPSTE24 进行其修饰的 C 末端 15 个氨基酸的蛋白水解切除。或 中的突变，阻碍了这种 prelamin A 切割步骤，导致过早衰老疾病 Hutchinson-Gilford 早衰综合征（HGPS），以及相关的早衰障碍下颌面骨发育不良 B 型（MAD-B）和限制性皮肤病（RD）。在这里，我们报告了开发“人源化酵母系统”来测定 ZMPSTE24 依赖性的 prelamin A 切割，并检查已知的八个与疾病相关的 错义突变。所有突变显示出减少的 prelamin A 处理，并分为三类，具有活性、蛋白质稳定性或两者的缺陷。值得注意的是，一些 ZMPSTE24 突变体可以通过删除涉及内质网（ER）相关的错误折叠膜蛋白降解的 E3 泛素连接酶 Doa10 或用蛋白酶体抑制剂硼替佐米来挽救。这一发现可能对一些患者具有重要的治疗意义。我们还表明，ZMPSTE24 介导的 prelamin A 切割可以与 ZMPSTE24 在清除 ER 膜转位器堵塞底物中最近发现的作用脱耦。与 ZMPSTE24 的晶体结构一起，该人源化酵母系统可以指导结构-功能研究，以揭示 prelamin A 切割、转位器疏通以及膜蛋白折叠和稳定性的机制。
