TMEM55B (transmembrane protein 55B) is a phosphatidylinositol-(4,5)-bisphosphate (PI[4,5]P2) phosphatase that regulates cellular cholesterol, modulates LDLR (low-density lipoprotein receptor) decay, and lysosome function. We tested the effects of Tmem55b knockdown on plasma lipids in mice and assessed the roles of LDLR lysosomal degradation and change in (PI[4,5]P2) in mediating these effects. Approach and Results: Western diet-fed C57BL/6J mice were treated with antisense oligonucleotides against Tmem55b or a nontargeting control for 3 to 4 weeks. Hepatic Tmem55b transcript and protein levels were reduced by ≈70%, and plasma non-HDL (high-density lipoprotein) cholesterol was increased ≈1.8-fold (P<0.0001). Immunoblot analysis of fast protein liquid chromatography (FPLC) fractions revealed enrichment of ApoE-containing particles in the LDL size range. In contrast, Tmem55b knockdown had no effect on plasma cholesterol in Ldlr-/- mice. In primary hepatocytes and liver tissues from Tmem55b knockdown mice, there was decreased LDLR protein. In the hepatocytes, there was increased lysosome staining and increased LDLR-lysosome colocalization. Impairment of lysosome function (incubation with NH4Cl or knockdown of the lysosomal proteins LAMP1 or RAB7) abolished the effect of TMEM55B knockdown on LDLR in HepG2 (human hepatoma) cells. Colocalization of the recycling endosome marker RAB11 (Ras-related protein 11) with LDLR in HepG2 cells was reduced by 50% upon TMEM55B knockdown. Finally, knockdown increased hepatic PI(4,5)P2 levels in vivo and in HepG2 cells, while TMEM55B overexpression in vitro decreased PI(4,5)P2. TMEM55B knockdown decreased, whereas overexpression increased, LDL uptake in HepG2 cells. Notably, the TMEM55B overexpression effect was reversed by incubation with PI(4,5)P2. Conclusions: These findings indicate a role for TMEM55B in regulating plasma cholesterol levels by affecting PI(4,5)P2-mediated LDLR lysosomal degradation.

中文翻译：

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磷脂酰肌醇-（4,5）-二磷酸通过LDL（低密度脂蛋白）受体溶酶体降解来调节血浆胆固醇。

TMEM55B（跨膜蛋白55B）是磷脂酰肌醇-（4,5）-双磷酸酯（PI [4,5] P2）磷酸酶，可调节细胞胆固醇，调节LDLR（低密度脂蛋白受体）的衰变和溶酶体功能。我们测试了Tmem55b敲低对小鼠血浆脂质的影响，并评估了LDLR溶酶体降解和（PI [4,5] P2）变化在介导这些作用中的作用。方法和结果：用抗Tmem55b的反义寡核苷酸或非靶向对照治疗西方饮食喂养的C57BL / 6J小鼠3至4周。肝Tmem55b转录和蛋白质水平降低了约70％，血浆非HDL（高密度脂蛋白）胆固醇升高了约1.8倍（P <0.0001）。快速蛋白质液相色谱（FPLC）馏分的免疫印迹分析显示，在LDL大小范围内，富含ApoE的颗粒富集。相反，Tmem55b敲低对Ldlr-/-小鼠的血浆胆固醇没有影响。在Tmem55b敲除小鼠的原代肝细胞和肝组织中，LDLR蛋白降低。在肝细胞中，溶酶体染色增加，LDLR-溶酶体共定位增加。溶酶体功能的损害（与NH4Cl一起孵育或溶酶体蛋白LAMP1或RAB7的敲除）消除了TMEM55B敲除对HepG2（人肝癌）细胞中LDLR的影响。在TMEM55B敲低后，循环内体标记RAB11（Ras相关蛋白11）与LDLR在HepG2细胞中的共定位降低了50％。最后，敲低会增加体内和HepG2细胞中肝PI（4,5）P2的水平，而TMEM55B在体外的过表达降低了PI（4,5）P2。TMEM55B敲低减少，而过表达增加，HepG2细胞中的LDL摄取。值得注意的是，通过与PI（4,5）P2的孵育逆转了TMEM55B的过表达作用。结论：这些发现表明，TMEM55B通过影响PI（4,5）P2介导的LDLR溶酶体降解来调节血浆胆固醇水平。