PEX1G843D remains functional in peroxisome biogenesis but is rapidly degraded by the proteasome.

Connor J Sheedy, Soham P Chowdhury, Bashir A Ali, Julia Miyamoto, Eric Z Pang, Julien Bacal, Katherine U Tavasoli, Chris D Richardson, Brooke M Gardner
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Abstract

The PEX1/PEX6 AAA-ATPase is required for the biogenesis and maintenance of peroxisomes. Mutations in HsPEX1 and HsPEX6 disrupt peroxisomal matrix protein import and are the leading cause of Peroxisome Biogenesis Disorders (PBDs). The most common disease-causing mutation in PEX1 is the HsPEX1G843D allele, which results in a reduction of peroxisomal protein import. Here we demonstrate that in vitro the homologous yeast mutant, ScPex1G700D, reduces the stability of Pex1's active D2 ATPase domain and impairs assembly with Pex6, but can still form an active AAA-ATPase motor. In vivo, ScPex1G700D exhibits only a slight defect in peroxisome import. We generated model human HsPEX1G843D cell lines and show that PEX1G843D is rapidly degraded by the proteasome, but that induced overexpression of PEX1G843D can restore peroxisome import. Additionally, we found that the G843D mutation reduces PEX1's affinity for PEX6, and that impaired assembly is sufficient to induce degradation of PEX1WT. Lastly, we found that fusing a deubiquitinase to PEX1G843D significantly hinders its degradation in mammalian cells. Altogether, our findings suggest a novel regulatory mechanism for PEX1/PEX6 hexamer assembly and highlight the potential of protein stabilization as a therapeutic strategy for PBDs arising from the G843D mutation and other PEX1 hypomorphs.

PEX1 G843D在过氧化物酶体的生物发生中保持功能,但被蛋白酶体迅速降解。
PEX1/PEX6 aaa - atp酶是过氧化物酶体的生物发生和维持所必需的。HsPEX1和HsPEX6的突变破坏过氧化物酶体基质蛋白的输入,是过氧化物酶体生物发生障碍(PBDs)的主要原因。PEX1中最常见的致病突变是Hs PEX1 G843D等位基因,它导致过氧化物酶体蛋白进口减少。本研究表明,在体外,同源酵母突变体Sc Pex1 G700D降低了Pex1活性D2 atp酶结构域的稳定性,并损害了与Pex6的组装,但仍然可以形成一个活性的aaa - atp酶马达。在体内,Sc Pex1 G700D在过氧化物酶体进口方面仅表现出轻微缺陷。我们建立了模型人Hs PEX1 G843D细胞系,发现PEX1 G843D被蛋白酶体快速降解,但诱导过表达PEX1 G843D可以恢复过氧化物酶体的导入。此外,我们发现G843D突变降低了PEX1对PEX6的亲和力,这种受损的组装足以诱导PEX1 WT的降解。最后,我们发现将去泛素酶融合到PEX1 G843D显著阻碍其在哺乳动物细胞中的降解。总之,我们的研究结果提示了PEX1/PEX6六聚体组装的一种新的调控机制,并强调了蛋白质稳定作为G843D突变和其他PEX1亚型引起的pbd的治疗策略的潜力。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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