Sterol-induced degradation of HMG CoA reductase depends on interplay of two Insigs and two ubiquitin ligases, gp78 and Trc8
Sterol-induced degradation of HMG CoA reductase depends on interplay of two Insigs and two ubiquitin ligases, gp78 and Trc8
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DOI：
10.1073/pnas.1112831108
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发表时间：
2011-12-20
影响因子：
11.1
通讯作者：
DeBose-Boyd, Russell A.
中科院分区：
文献类型：
--
作者：
Jo, Youngah;Lee, Peter C. W.;DeBose-Boyd, Russell A.
内质网 (ER) 膜中甾醇的积累导致 3-羟基-3-甲基戊二酰辅酶 A 还原酶（胆固醇和非甾醇类异戊二烯合成的限速酶）的泛素化和蛋白酶体降解加速。这种降解是由甾醇诱导的还原酶与内质网膜的 Insig-1 或 Insig-2 蛋白结合引起的。我们之前报道过，在永生化人成纤维细胞（SV-589 细胞）中，Insig-1（而非 Insig-2）招募 gp78（一种膜结合环指泛素连接酶）。我们现在报告 Insig-1 和 Insig-2 都结合另一种膜结合环指泛素连接酶，称为 Trc8。通过 RNA 干扰 (RNAi) 敲低 SV-589 细胞中的 gp78 或 Trc8，可抑制甾醇诱导的还原酶泛素化，并抑制甾醇诱导的降解达 50-60%。 gp78 和 Trc8 的联合敲低产生了更完全的降解抑制 (> 90%)。 gp78 的敲低导致 Trc8 和 Insig-1 蛋白水平增加三到四倍，这与 gp78 的抑制作用相反。相比之下，Trc8 的敲低对 gp78 或 Insig-1 没有影响。目前的结果表明，甾醇诱导的还原酶泛素化和蛋白酶体降解是由至少四种蛋白质的复杂相互作用决定的：Insig-1、Insig-2、gp78 和 Trc8。这些蛋白质中任何一种的浓度变化可能是还原酶降解的细胞和/或组织特异性调节的差异的原因。
Accumulation of sterols in membranes of the endoplasmic reticulum (ER) leads to the accelerated ubiquitination and proteasomal degradation of 3-hydroxy-3-methylglutaryl coenzyme A reductase, a rate-limiting enzyme in synthesis of cholesterol and nonsterol isoprenoids. This degradation results from sterol-induced binding of reductase to the Insig-1 or Insig-2 proteins of ER membranes. We previously reported that in immortalized human fibroblasts (SV-589 cells) Insig-1, but not Insig-2, recruits gp78, a membrane-bound RING-finger ubiquitin ligase. We now report that both Insig-1 and Insig-2 bind another membrane-bound RING-finger ubiquitin ligase called Trc8. Knockdown of either gp78 or Trc8 in SV-589 cells through RNA interference (RNAi) inhibited sterol-induced ubiquitination of reductase and inhibited sterol-induced degradation by 50-60%. The combined knockdown of gp78 and Trc8 produced a more complete inhibition of degradation (> 90%). Knockdown of gp78 led to a three to fourfold increase in levels of Trc8 and Insig-1 proteins, which opposed the inhibitory action of gp78. In contrast, knockdown of Trc8 had no effect on gp78 or Insig-1. The current results suggest that sterol-induced ubiquitination and proteasomal degradation of reductase is dictated by the complex interplay of at least four proteins: Insig-1, Insig-2, gp78, and Trc8. Variations in the concentrations of any one of these proteins may account for differences in cell- and/or tissue-specific regulation of reductase degradation.