A conserved ATG2 binding site in WIPI4 and yeast Hsv2 is disrupted by mutations causing β-propeller protein-associated neurodegeneration

Abstract

PROPPINs are phosphoinositide-binding β-propeller proteins that mediate membrane recruitment of other proteins and are involved in different membrane remodeling processes. The main role of PROPPINs is their function in autophagy, where they act at different steps in phagophore formation. The human PROPPIN WIPI4 (WDR45) forms a complex with ATG2 involved in phagophore elongation, and mutations in this gene cause β-propeller protein-associated neurodegeneration (BPAN). The yeast functional counterpart of WIPI4 is Atg18, although its closest sequence homolog is another member of the PROPPIN family, Hsv2, whose function remains largely undefined. Here, we provide evidence that Hsv2, like WIPI4 and Atg18, interact with Atg2. We show that Hsv2 and a pool of Atg2 colocalize on endosomes under basal conditions, and at the pre-autophagosomal structure (PAS) upon autophagy induction. We further show that Hsv2 drives the recruitment of Atg2 to endosomes while Atg2 mediates Hsv2 recruitment to the PAS. HSV2 overexpression results in mis-sorting and secretion of carboxypeptidase CPY, suggesting that the endosomal function of this protein is related to the endosome-to-Golgi recycling pathway. Furthermore, we show that the Atg2 binding site is conserved in Hsv2 and WIPI4 but not in Atg18. Notably, two WIPI4 residues involved in ATG2 binding are mutated in patients with BPAN and there is a correlation between the inhibitory effect of these mutations on ATG2 binding and the severity of the disease.

The association of lipid transfer protein VPS13A with endosomes is mediated by sorting nexin SNX5

“…Missense and additional C-terminal truncating mutations were obtained by using mutagenic PCR. pmCherry-SNX5 and pEGFP-SNX5 were generated by cloning the full-length SNX5 coding sequences in the polylinker of pmCherry ( Bueno-Arribas et al, 2021 ) and pEGFP-C3 ( GenBank Accession Number # U57607 ). VPS13A fragments were obtained by PCR and inserted into pEGF-C3.…”

Autophagy in health and disease: From molecular mechanisms to therapeutic target

“…75 Accumulating evidence from recent studies show that both WIPI3 and WIPI4 can interact with ATG2A via the WIR (WIPI-interacting-region) motif. 33,[81][82][83] Further studies show that the WIPI3/WIPI4-ATG2A complex serve to establish the contact sites between ER and phagophore, enabling the transportation of lipids to promote phagophore expansion. [33][34][35] Moreover, WIPI3/WIPI4-ATG2A complex is also involved in autophagosome-lysosome fusion, 84 adding to the complexity of the WIPIs functions.…”

“…Recently, it has been reported that WIPI3, similar to WIPI2, is also able to interact with ATG16L1 to promote LC3B lipidation 75 . Accumulating evidence from recent studies show that both WIPI3 and WIPI4 can interact with ATG2A via the WIR (WIPI‐interacting‐region) motif 33,81–83 . Further studies show that the WIPI3/WIPI4–ATG2A complex serve to establish the contact sites between ER and phagophore, enabling the transportation of lipids to promote phagophore expansion 33–35 .…”

Expanding the view of the molecular mechanisms of autophagy pathway

“…Another member of the yeast PROPPIN family is Hsv2. Recent reports suggest that, under basal conditions, Hsv2 colocalize with a pool of Atg2 on endosomes whereas upon autophagy induction, the colocalization happens at PAS (Bueno‐Arribas et al, 2021, p. 2).…”