SNX14 mutations affect endoplasmic reticulum-associated neutral lipid metabolism in autosomal recessive spinocerebellar ataxia 20
SNX14 mutations affect endoplasmic reticulum-associated neutral lipid metabolism in autosomal recessive spinocerebellar ataxia 20
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DOI：
10.1093/hmg/ddy101
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发表时间：
2018-06-01
影响因子：
3.5
通讯作者：
Stanier, Philip
中科院分区：
文献类型：
--
作者：
Bryant, Dale;Liu, Yang;Stanier, Philip
SNX 14突变导致常染色体隐性小脑共济失调20（SCAR 20）。突变通常导致蛋白质的丢失，尽管也报道了几种编码区缺失。患者来源的成纤维细胞显示出破坏的自噬，但SNX 14的确切功能尚不清楚。酵母同源物Mdm 1在内质网（ER）-溶酶体/空泡细胞器间束缚中起作用，但在哺乳动物中仍然需要功能保守。在这里，我们表明SNX 14的丢失改变但不阻止自噬通量。此外，我们发现SNX 14是一种ER相关蛋白，在中性脂质稳态和细胞器间串扰中发挥作用。SNX 14需要其N-末端跨膜螺旋用于ER定位，而Phox同源（PX）结构域用于亚细胞定位。SNX 14突变成纤维细胞和SNX 14（KO）HEK 293细胞均积聚异常胞质空泡，表明内溶酶体稳态缺陷。然而，ER-晚期内体/溶酶体接触位点在SNX 14（KO）细胞中维持，表明其不是ER-内溶酶体束缚的先决条件。SNX 14缺乏症的进一步研究表明中性脂质代谢的一般缺陷。SNX 14（KO）细胞在LAMP 1阳性溶酶体结构中显示出明显的菲律宾蛋白核周蓄积，表明胆固醇蓄积。与此一致，SNX 14（KO）细胞显示胆固醇酯水平轻微但可检测的降低，这在U18666 A中加剧。最后，SNX 14与ER衍生的脂滴（LD）油酸酯处理后，表明在ER-LD串扰的作用。因此，我们确定了SNX 14在ER、溶酶体和LD之间的中性脂质稳态中的重要作用，这可能提供早期干预靶点以减轻SCAR 20的临床症状。
Mutations in SNX14 cause the autosomal recessive cerebellar ataxia 20 (SCAR20). Mutations generally result in loss of protein although several coding region deletions have also been reported. Patient-derived fibroblasts show disrupted autophagy, but the precise function of SNX14 is unknown. The yeast homolog, Mdm1, functions in endoplasmic reticulum (ER)-lysosome/vacuole inter-organelle tethering, but functional conservation in mammals is still required. Here, we show that loss of SNX14 alters but does not block autophagic flux. In addition, we find that SNX14 is an ER-associated protein that functions in neutral lipid homeostasis and inter-organelle crosstalk. SNX14 requires its N-terminal transmembrane helices for ER localization, while the Phox homology (PX) domain is dispensable for subcellular localization. Both SNX14-mutant fibroblasts and SNX14(KO) HEK293 cells accumulate aberrant cytoplasmic vacuoles, suggesting defects in endolysosomal homeostasis. However, ER-late endosome/lysosome contact sites are maintained in SNX14(KO) cells, indicating that it is not a prerequisite for ER-endolysosomal tethering. Further investigation of SNX14-deficiency indicates general defects in neutral lipid metabolism. SNX14(KO) cells display distinct perinuclear accumulation of filipin in LAMP1-positive lysosomal structures indicating cholesterol accumulation. Consistent with this, SNX14(KO) cells display a slight but detectable decrease in cholesterol ester levels, which is exacerbated with U18666A. Finally, SNX14 associates with ER-derived lipid droplets (LD) following oleate treatment, indicating a role in ER-LD crosstalk. We therefore identify an important role for SNX14 in neutral lipid homeostasis between the ER, lysosomes and LDs that may provide an early intervention target to alleviate the clinical symptoms of SCAR20.