AP-4 vesicles contribute to spatial control of autophagy via RUSC-dependent peripheral delivery of ATG9A

Abstract

Adaptor protein 4 (AP-4) is an ancient membrane trafficking complex, whose function has largely remained elusive. In humans, AP-4 deficiency causes a severe neurological disorder of unknown aetiology. We apply unbiased proteomic methods, including ‘Dynamic Organellar Maps’, to find proteins whose subcellular localisation depends on AP-4. We identify three transmembrane cargo proteins, ATG9A, SERINC1 and SERINC3, and two AP-4 accessory proteins, RUSC1 and RUSC2. We demonstrate that AP-4 deficiency causes missorting of ATG9A in diverse cell types, including patient-derived cells, as well as dysregulation of autophagy. RUSC2 facilitates the transport of AP-4-derived, ATG9A-positive vesicles from the trans-Golgi network to the cell periphery. These vesicles cluster in close association with autophagosomes, suggesting they are the “ATG9A reservoir” required for autophagosome biogenesis. Our study uncovers ATG9A trafficking as a ubiquitous function of the AP-4 pathway. Furthermore, it provides a potential molecular pathomechanism of AP-4 deficiency, through dysregulated spatial control of autophagy.

Membrane trafficking in health and disease

“…Another group of HSPs is caused by mutations within subunits of the AP4 and AP5 adaptor complexes that function in post-Golgi trafficking (Bauer et al, 2012;Hardies et al, 2015;Moreno-De-Luca et al, 2011;Slabicki et al, 2010;Verkerk et al, 2009), which likely affect endolysosomal function (Sanger et al, 2019). Interestingly, AP4 is important for trafficking of the autophagy-initiating factor ATG9A, suggesting a link between HSP and dysregulated autophagy (Mattera et al, 2017;Davies et al, 2018). HSP also results from mutations in ubiquitinassociated protein 1 (UBAP1) and VPS37A (Farazi Fard et al, 2019;Zivony-Elboum et al, 2012), components of ESCRT-I required for MVB sorting (Schmidt and Teis, 2012).…”

Autophagy in Neurodegenerative Diseases: A Hunter for Aggregates

“…The AP-4 subunits β-1, µ-1, ε-1, and σ-1 are encoded by SPG47, SPG50, SPG51, and SPG52, respectively, and associated with childhood-onset HSP [221]. As the AP-4 complex mediates the sorting of the target proteins from the TGN to another membrane structure, AP-4 deficiency results in the missorting of ATG9A-containing vesicles from the TGN to autophagosome formation sites in the neurons [221,222]. Another AP complex subunit linked with HSP is AP-5 ζ-1, which is encoded by SPG48.…”

“…Therefore, caution should be taken when targeting autophagy for the treatment of neurodegenerative diseases. sorting of the target proteins from the TGN to another membrane structure, AP-4 deficiency results in the missorting of ATG9A-containing vesicles from the TGN to autophagosome formation sites in the neurons [221,222]. Another AP complex subunit linked with HSP is AP-5 ζ-1, which is encoded by SPG48.…”

Syntaxin 16’s Newly Deciphered Roles in Autophagy

“…Autophagosome biogenesis has been shown to occur at locations as varied as endoplasmic reticulum (ER)-mitochondria contact sites [7], ER exit sites [8,9], the trans-Golgi network (TGN) [10], ER-plasma membrane contact sites [11] and the recycling endosomes [12]. The diversity of autophagosomal origin is reflected by the fact that most of the known coat protein complexes in the exocytic and endocytic pathways, including coat protein I (COPI) [13], COPII [14][15][16][17], AP1 [10], AP2 [18] and AP4 [19,20], have all been implicated in the supply of membrane sources for autophagosome formation. In this regard, perhaps the most prominent are ER-derived COPII vesicles [14][15][16][17][21][22][23][24] and Atg9-containing, endosome-derived vesicles [18][19][20][25][26][27][28][29].…”

“…The diversity of autophagosomal origin is reflected by the fact that most of the known coat protein complexes in the exocytic and endocytic pathways, including coat protein I (COPI) [13], COPII [14][15][16][17], AP1 [10], AP2 [18] and AP4 [19,20], have all been implicated in the supply of membrane sources for autophagosome formation. In this regard, perhaps the most prominent are ER-derived COPII vesicles [14][15][16][17][21][22][23][24] and Atg9-containing, endosome-derived vesicles [18][19][20][25][26][27][28][29]. The multiple membrane-spanning Atg9 could be found at membranes of the TGN, recycling endosomes as well as a subpopulation of cytoplasmic membrane vesicles, and upon autophagy induction translocates to sites of autophagosome biogenesis [30,31].…”