PACSIN1 is indispensable for amphisome-lysosome fusion during basal autophagy and subsets of selective autophagy.
PACSIN1 is indispensable for amphisome-lysosome fusion during basal autophagy and subsets of selective autophagy.
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DOI：
10.1371/journal.pgen.1010264
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发表时间：
2022-06
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影响因子：
4.5
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中科院分区：
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自噬是降解细胞质物质以维持细胞稳态的重要过程。在自噬过程中，双膜自噬体包围细胞质物质，要么与核内体（称为两性体）融合，然后再与溶酶体融合，要么直接与溶酶体融合，在这两种情况下都产生由溶酶体水解酶降解其内容物的自噬体。然而，目前尚不清楚是否有特定的机制和/或条件来区分这些替代途径。在这里，我们发现PACSIN1是一种新的自噬调节因子。在基础营养丰富的条件下，PACSIN1缺失显著降低了自噬活性，而在饥饿条件下则没有。电镜和共定位分析表明，PACSIN1缺失导致了两性体的积累，表明溶酶体融合受到抑制。PACSIN1与自噬SNARE SNAP29相互作用，是STX17和YKT6复合物正确组装所必需的。此外，溶噬、聚合不需要PACSIN1，这提示了货物特异性融合机制。在秀丽隐杆线虫中，PACSIN1的同源物sdn -1的缺失抑制了基础自噬和聚集蛋白的清除，这表明PACSIN1的保守作用。综上所述，我们的研究结果表明，两性体-溶酶体融合过程在营养状态和应激条件下受到优先调节，而PACSIN1是自噬过程中特异性的关键。自噬是一种进化保守的细胞质降解系统，在该系统中，称为自噬体的双膜结构将一些细胞质物质隔离，然后转运到溶酶体进行降解。以往主要基于电镜的研究表明，自噬体要么直接与溶酶体融合，要么与核内体/多泡体（MVB）融合，产生两性体，然后与溶酶体融合。然而，目前尚不清楚这些过程是如何调节的，由于缺乏涉及这两种途径中的任何一种的关键分子，这两种途径的生理相关性阻碍了详细的表征。在本研究中，我们发现PACSIN1是一种新的自噬调节因子，其功能对两性体-溶酶体融合过程至关重要。通过对PACSIN1的分析，我们发现基础自噬和选择性自噬亚群需要通过两性体的PACSIN1依赖融合过程，这表明根据环境和/或货物的不同，有两种自噬途径被利用。
Autophagy is an indispensable process that degrades cytoplasmic materials to maintain cellular homeostasis. During autophagy, double-membrane autophagosomes surround cytoplasmic materials and either fuse with endosomes (called amphisomes) and then lysosomes, or directly fuse with lysosomes, in both cases generating autolysosomes that degrade their contents by lysosomal hydrolases. However, it remains unclear if there are specific mechanisms and/or conditions which distinguish these alternate routes. Here, we identified PACSIN1 as a novel autophagy regulator. PACSIN1 deletion markedly decreased autophagic activity under basal nutrient-rich conditions but not starvation conditions, and led to amphisome accumulation as demonstrated by electron microscopic and co-localization analysis, indicating inhibition of lysosome fusion. PACSIN1 interacted with SNAP29, an autophagic SNARE, and was required for proper assembly of the STX17 and YKT6 complexes. Moreover, PACSIN1 was required for lysophagy, aggrephagy but not mitophagy, suggesting cargo-specific fusion mechanisms. In C. elegans, deletion of sdpn-1, a homolog of PACSINs, inhibited basal autophagy and impaired clearance of aggregated protein, implying a conserved role of PACSIN1. Taken together, our results demonstrate the amphisome-lysosome fusion process is preferentially regulated in response to nutrient state and stress, and PACSIN1 is a key to specificity during autophagy. Autophagy is an evolutionally conserved cytoplasmic degradation system in which double membrane structure called autophagosomes sequester several cytoplasmic materials and then transport to lysosomes for degradation. Previous studies mainly based on electron microscopy indicates autophagosomes either fuse with lysosomes directly or fuse with endosomes/MVB (Multi Vesicular Body), producing amphisomes then fuse with lysosomes. However, it remains unknown how these processes are regulated and the physiological relevance of these two routes due to lack of key molecules involved in either of two routes precludes the detailed characterization. In the current study, we identified PACSIN1 as a novel regulator of autophagy, whose function is essential for amphisome-lysosome fusion process specifically. Through the analysis of PACSIN1, we revealed that PACSIN1-dependent fusion process via amphisomes is required for basal autophagy and subsets of selective autophagy, suggesting that two autophagic routes are utilized depending on the context and/or cargos.