Unconventional secretion of alpha-synuclein mediated by palmitoylated DNAJC5 oligomers

Alpha-synuclein (α-syn), a major component of Lewy bodies found in Parkinson’s disease (PD) patients, has been found exported outside of cells and may mediate its toxicity via cell-to-cell transmission. Here, we reconstituted soluble, monomeric α-syn secretion by the expression of DnaJ homolog subfamily C member 5 (DNAJC5) in HEK293T cells. DNAJC5 undergoes palmitoylation and anchors on the membrane. Palmitoylation is essential for DNAJC5-induced α-syn secretion, and the secretion is not limited by substrate size or unfolding. Cytosolic α-syn is actively translocated and sequestered in an endosomal membrane compartment in a DNAJC5-dependent manner. Reduction of α-syn secretion caused by a palmitoylation-deficient mutation in DNAJC5 can be reversed by a membrane-targeting peptide fusion-induced oligomerization of DNAJC5. The secretion of endogenous α-syn mediated by DNAJC5 is also found in a human neuroblastoma cell line, SH-SY5Y, differentiated into neurons in the presence of retinoic acid, and in human-induced pluripotent stem cell-derived midbrain dopamine neurons. We propose that DNAJC5 forms a palmitoylated oligomer to accommodate and export α-syn.α-突触核蛋白（Alpha-synuclein）是帕金森病（PD）患者路易小体的主要组成成分，现已被证实可被分泌至细胞外，并可能通过细胞间传递介导其毒性。本研究通过在HEK293T细胞中表达DnaJ同源家族C亚家族成员5（DNAJC5），重构了可溶性单体α-突触核蛋白的分泌体系。DNAJC5可发生棕榈酰化并锚定在细胞膜上；棕榈酰化对于DNAJC5诱导的α-突触核蛋白分泌至关重要，且该分泌过程不受底物尺寸或解折叠状态的限制。胞质内的α-突触核蛋白可通过DNAJC5依赖的方式被主动转运并捕获于内体膜区室中。由DNAJC5棕榈酰化缺陷突变所导致的α-突触核蛋白分泌减少，可通过膜靶向肽融合诱导的DNAJC5寡聚化得以逆转。在经视黄酸诱导分化为神经元的人神经母细胞瘤细胞系SH-SY5Y，以及人诱导多能干细胞衍生的中脑多巴胺能神经元中，同样存在DNAJC5介导的内源性α-突触核蛋白分泌现象。我们推测，DNAJC5可形成棕榈酰化寡聚体，以容纳并分泌α-突触核蛋白。