Sterol-regulated HMG-CoA reductase (HMGCR) degradation and SREBP-2 cleavage are two major feedback regulatory mechanisms governing cholesterol biosynthesis. Reportedly, lanosterol selectively stimulates HMGCR degradation, and cholesterol is a specific regulator of SREBP-2 cleavage. However, it is unclear whether other endogenously generated sterols regulate these events. Here, we investigated the sterol intermediates from the mevalonate pathway of cholesterol biosynthesis using a CRISPR/Cas9-mediated genetic engineering approach. With a constructed HeLa cell line expressing the mevalonate transporter, we individually deleted genes encoding major enzymes in the mevalonate pathway, used lipidomics to measure sterol intermediates, and examined HMGCR and SREBP-2 statuses. We found that the C4-dimethylated sterol intermediates, including lanosterol, 24,25-dihydrolanosterol, follicular fluid meiosis activating sterol, testis meiosis activating sterol, and dihydro-testis meiosis activating sterol, were significantly upregulated upon mevalonate loading. These intermediates augmented both degradation of HMGCR and inhibition of SREBP-2 cleavage. The accumulated lanosterol induced rapid degradation of HMGCR, but did not inhibit SREBP-2 cleavage. The newly synthesized cholesterol from the mevalonate pathway is dispensable for inhibiting SREBP-2 cleavage. Together, these results suggest that lanosterol is a bona fide endogenous regulator that specifically promotes HMGCR degradation, and that other C4-dimethylated sterol intermediates may regulate both HMGCR degradation and SREBP-2 cleavage.

中文翻译：

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甲羟戊酸途径的内源性固醇中间体调节HMGCR降解和SREBP-2加工。

甾醇调节的HMG-CoA还原酶（HMGCR）降解和SREBP-2裂解是控制胆固醇生物合成的两个主要反馈调节机制。据报道，羊毛甾醇选择性刺激HMGCR降解，胆固醇是SREBP-2裂解的特定调节剂。但是，尚不清楚其他内源性固醇是否调节这些事件。在这里，我们使用CRISPR / Cas9介导的基因工程方法研究了胆固醇生物合成的甲羟戊酸酯途径中的甾醇中间体。使用构建的表达甲羟戊酸转运蛋白的HeLa细胞系，我们分别删除了编码甲羟戊酸途径中主要酶的基因，使用脂质组学测量固醇中间体，并检查了HMGCR和SREBP-2的状态。我们发现C4-二甲基化的固醇中间体，包括羊毛甾醇，24，25-二氢羊毛甾醇，卵泡液减数分裂激活固醇，睾丸减数分裂激活固醇和二氢睾丸减数分裂激活固醇在甲羟戊酸负载后显着上调。这些中间体既增加了HMGCR的降解又抑制了SREBP-2的裂解。积累的羊毛甾醇诱导HMGCR快速降解，但不抑制SREBP-2裂解。来自甲羟戊酸途径的新合成的胆固醇对于抑制SREBP-2裂解是必要的。总之，这些结果表明羊毛甾醇是真正促进HMGCR降解的真正的内源性调节剂，而其他C4-二甲基化固醇中间体可能同时调节HMGCR降解和SREBP-2裂解。甲羟戊酸负载后，二氢睾丸减数分裂激活固醇和二氢睾丸减数分裂激活固醇显着上调。这些中间体既增加了HMGCR的降解又抑制了SREBP-2的裂解。积累的羊毛甾醇诱导HMGCR快速降解，但不抑制SREBP-2裂解。来自甲羟戊酸途径的新合成的胆固醇对于抑制SREBP-2裂解是必要的。总之，这些结果表明羊毛甾醇是真正促进HMGCR降解的真正的内源性调节剂，而其他C4-二甲基化固醇中间体可能同时调节HMGCR降解和SREBP-2裂解。甲羟戊酸负载后，二氢睾丸减数分裂激活固醇和二氢睾丸减数分裂激活固醇显着上调。这些中间体既增加了HMGCR的降解又抑制了SREBP-2的裂解。积累的羊毛甾醇诱导HMGCR快速降解，但不抑制SREBP-2裂解。来自甲羟戊酸途径的新合成的胆固醇对于抑制SREBP-2裂解是必要的。总之，这些结果表明，羊毛甾醇是真正促进HMGCR降解的真正的内源性调节剂，而其他C4-二甲基化固醇中间体可以调节HMGCR降解和SREBP-2裂解。积累的羊毛甾醇诱导HMGCR快速降解，但不抑制SREBP-2裂解。来自甲羟戊酸途径的新合成的胆固醇对于抑制SREBP-2裂解是必要的。总之，这些结果表明羊毛甾醇是真正促进HMGCR降解的真正的内源性调节剂，而其他C4-二甲基化固醇中间体可能同时调节HMGCR降解和SREBP-2裂解。积累的羊毛甾醇诱导HMGCR快速降解，但不抑制SREBP-2裂解。来自甲羟戊酸途径的新合成的胆固醇对于抑制SREBP-2裂解是必要的。总之，这些结果表明羊毛甾醇是真正促进HMGCR降解的真正的内源性调节剂，而其他C4-二甲基化固醇中间体可能同时调节HMGCR降解和SREBP-2裂解。