A heterodimeric SNX4:SNX7 SNX-BAR autophagy complex coordinates ATG9A trafficking for efficient autophagosome assembly

Abstract

STATEMENT: A heterodimeric SNX4:SNX7 SNX-BAR complex regulates mammalian autophagosome assembly through the control of ATG9 trafficking. ABSTRACTEfficient mammalian autophagosome biogenesis requires coordinated input from other cellular endomembrane compartments. Such coordination includes the stimulated trafficking to autophagosome assembly sites of the essential autophagy proteins, ATG9 and ATG16L1, via distinct endosomal compartments. Protein trafficking within the endocytic network is directed by a conserved family of sorting nexins (SNXs), with previous studies implicating SNX18 (an SH3 domain-type SNX-BAR protein) in the mobilisation of ATG9A and ATG16L1 from recycling endosomes during autophagy. Using siRNA and CRISPR-Cas9, we demonstrate that a second mammalian SNX-BAR, SNX4, is needed for efficient LC3 lipidation and autophagosome assembly in mammalian cells. SNX-BARs exist as homo-and heterodimers, and we show that SNX4 forms functional heterodimers with either SNX7 or SNX30, and that these associate with tubulovesicular endocytic membranes at steady state. Detailed image-based analysis during the early stages of autophagosome assembly reveal that SNX4:SNX7 is the autophagyspecific heterodimeric SNX-BAR complex, required for efficient recruitment/retention of core autophagy regulators at the nascent isolation membrane. SNX4 partially co-localises with juxtanuclear ATG9Apositive membranes, with our data linking the SNX4 autophagy defect to the mis-trafficking and/or retention of ATG9A in the Golgi region. Together, our findings show that the SNX4:SNX7 heterodimer coordinates ATG9A trafficking within the endocytic network to establish productive autophagosome assembly sites.

Sorting Nexins in Protein Homeostasis

“…The SNX4/SNX7 heterodimer affected autophagosome assembly rather than autophagic flux. influenced autophagosome assembly by controlling ATG9 trafficking [34]. In a similar study, SNX18 along with Dynamin-2 regulates ATG9 trafficking from recycling endosomes to autophagosomes [141].…”

“…Sorting nexins also interact with cargoes outside of endo-lysosomal-TGN pathways (see Figure 1). Here they play significant roles in both selective and non-selective autophagy pathways in yeast and mammalian cells alike [31][32][33][34]. This serves to emphasize the diverse regulatory roles sorting nexins play in maintaining protein homeostasis.…”

“…These are critical for both endosome-to-plasma membrane recycling and endosome-to-TGN retrieval [71]. More recently, they have identified as important players in autophagic processes, which is discussed in more detail below in section 5 [34,72].…”

Selectivity of mRNA degradation by autophagy in yeast

“…Atg24 consists of a sorting nexin complex with Atg20 or Snx41 [32][33][34] . We also tested the impact of Atg20 and Snx41 for vacuolar delivery of mRNAs.…”

Vps501, a novel vacuolar SNX-BAR protein cooperates with the SEA complex to induce autophagy

“…BAR domains allow members of the SNX-BAR subfamily to bind high positive curvature structures, driving the formation of endosomal tubules, while also conferring the ability to form sorting complexes that facilitate cargo selection (Chi et al, 2014;van Weering et al, 2012;van Weering et al, 2010). The Saccharomyces cerevisiae genome encodes seven annotated SNX-BAR proteins, while the human genome encodes twelve (Anton et al, 2020;Teasdale and Collins, 2012;van Weering et al, 2010). In addition to working with complexes such as retromer to mediate retrograde and recycling trafficking of cargos at the tubular endosomal network (TEN) (Burd and Cullen, 2014;Chi et al, 2014;Teasdale and Collins, 2012;van Weering et al, 2012;van Weering et al, 2010), SNX-BAR proteins contribute to other important conserved cellular processes such as macroautophagy (herein referred to as autophagy) and selective autophagy (Ma et al, 2017;Ma et al, 2018;Nemec et al, 2017).…”