Phospholipid cofactor solubilization inhibits formation of native prions.
Phospholipid cofactor solubilization inhibits formation of native prions.
复制标题
磷脂辅助因子溶解抑制天然朊病毒的形成。
DOI：
10.1111/jnc.15930
复制
发表时间：
2023
影响因子：
4.7
通讯作者：
Supattapone,Surachai
中科院分区：
文献类型：
--
作者：
Schwind,AbigailM;Walsh,DanielJ;Burke,CassandraM;Supattapone,Surachai
辅因子分子是体外产生感染性哺乳动物朊病毒所必需的。小鼠和仓鼠朊病毒似乎有不同的辅因子偏好：虽然小鼠和仓鼠朊病毒都可以使用磷脂酰乙醇胺（PE）作为朊病毒辅因子，但只有仓鼠朊病毒也可以使用单链RNA作为替代辅因子。在这里，我们研究了去污剂增溶对体外啮齿动物朊病毒形成的影响。我们发现，可以溶解PE的去污剂（正辛基葡萄糖苷、正辛基半乳糖苷和CHAPS）在使用田鼠脑匀浆底物的系列蛋白质错误折叠循环扩增（sPMCA）反应中抑制小鼠朊病毒的形成，而不能溶解PE的去污剂（Triton X-100和IPEGAL）则没有影响。对于所有三种PE增溶洗涤剂，仅在临界胶束浓度（CMC）以上观察到对RML小鼠朊病毒形成的抑制。另外两种小鼠朊病毒株Me 7和301 C也被三种PE溶解洗涤剂抑制，但不被Triton X-100或IPEGAL抑制。相比之下，没有洗涤剂抑制仓鼠朊病毒的形成在平行的sPMCA反应使用相同的银行田鼠脑匀浆基板。在使用纯化底物的复溶sPMCA反应中，当免疫纯化的银行田鼠PrPC底物补充脑磷脂而不是RNA时，正辛基葡糖苷抑制仓鼠朊病毒形成。有趣的是，磷脂辅因子增溶在使用细菌表达的重组PrP底物的sPMCA反应中没有影响，表明增溶的抑制作用需要PrP翻译后修饰。总体而言，这些体外结果表明，PE促进天然而非重组朊病毒形成的能力需要磷脂双层的完整性，这表明膜结构可能在体内朊病毒形成中起重要作用。
Cofactor molecules are required to generate infectious mammalian prions in vitro. Mouse and hamster prions appear to have different cofactor preferences: Whereas both mouse and hamster prions can use phosphatidylethanolamine (PE) as a prion cofactor, only hamster prions can also use single‐stranded RNA as an alternative cofactor. Here, we investigated the effect of detergent solubilization on rodent prion formation in vitro. We discovered that detergents that can solubilize PE (n‐octylglucoside, n‐octylgalactoside, and CHAPS) inhibit mouse prion formation in serial protein misfolding cyclic amplification (sPMCA) reactions using bank vole brain homogenate substrate, whereas detergents that are unable to solubilize PE (Triton X‐100 and IPEGAL) have no effect. For all three PE‐solubilizing detergents, inhibition of RML mouse prion formation was only observed above the critical micellar concentration (CMC). Two other mouse prion strains, Me7 and 301C, were also inhibited by the three PE‐solubilizing detergents but not by Triton X‐100 or IPEGAL. In contrast, none of the detergents inhibited hamster prion formation in parallel sPMCA reactions using the same bank vole brain homogenate substrate. In reconstituted sPMCA reactions using purified substrates, n‐octylglucoside inhibited hamster prion formation when immunopurified bank vole PrPCsubstrate was supplemented with brain phospholipid but not with RNA. Interestingly, phospholipid cofactor solubilization had no effect in sPMCA reactions using bacterially expressed recombinant PrP substrate, indicating that the inhibitory effect of solubilization requires PrPCpost‐translational modifications. Overall, these in vitro results show that the ability of PE to facilitate the formation of native but not recombinant prions requires phospholipid bilayer integrity, suggesting that membrane structure may play an important role in prion formation in vivo.