Impairment of autophagosome-lysosome fusion in the buff mutant mice with the VPS33A^D251E mutation

Abstract

-associated membrane protein 1; LRO, lysosome-related organelle; MAP1LC3/LC3, microtubule-associated protein 1 light chain 3; MEF, mouse embryonic fibroblast; RFP, red fluorescent protein; SNARE, soluble Nethylmaleimide-sensitive attachment protein (SNAP) receptor; SQSTM1/p62, sequestosome 1; STX, syntaxin; TH, tyrosine hydroxylase; VAMP, vesicle associated membrane protein/synaptobrevin; VPS, vacuole protein sorting; WT, wild type.The HOPS (homotypic fusion and protein sorting) complex functions in endocytic and autophagic pathways in both lower eukaryotes and mammalian cells through its involvement in fusion events between endosomes and lysosomes or autophagosomes and lysosomes. However, the differential molecular mechanisms underlying these fusion processes are largely unknown. Buff (bf) is a mouse mutant that carries an Asp251-to-Glu point mutation (D251E) in the VPS33A protein, a tethering protein and a core subunit of the HOPS complex. Bf mice showed impaired spontaneous locomotor activity, motor learning, and autophagic activity. Although the gross anatomy of the brain was apparently normal, the number of Purkinje cells was significantly reduced. Furthermore, we found that fusion between autophagosomes and lysosomes was defective in bf cells without compromising the endocytic pathway. The direct association of mutant VPS33A D251E with the autophagic SNARE complex, STX17 (syntaxin 17)-VAMP8-SNAP29, was enhanced. In addition, the VPS33A D251E mutation enhanced interactions with other HOPS subunits, namely VPS41, VPS39, VPS18, and VPS11, except for VPS16. Reduction of the interactions between VPS33A Y440D and several other HOPS subunits led to decreased association with STX17. These results suggest that the VPS33A D251E mutation plays dual roles by increasing the HOPS complex assembly and its association with the autophagic SNARE complex, which selectively affects the autophagosome-lysosome fusion that impairs basal autophagic activity and induces Purkinje cell loss.

VPS41recessive mutation causes ataxia and dystonia with retinal dystrophy and mental retardation by inhibiting HOPS function and mTORC1 signaling

“…Currently, patients with mutations in HOPS core components VPS11, VPS16 and VPS33A have been reported in literature, which all show a neurodegenerative phenotype (11,(67)(68)(69)(70)(71)(72)(73)(74). Moreover, during our studies, a third patient with compound heterozygote mutations in VPS41 was identified (Personal communication by I. Stolte-Dijkstra, University Medical Center Groningen), bearing the VPS41 R662* mutation as well as a missense mutation (VPS41 H466R ), and displaying a similar neurological phenotype.…”

Crosstalk between the Secretory and Autophagy Pathways Regulates Autophagosome Formation

“…Additionally, HOPS subunits were shown to interact with the autophagosomal SNARE, Stx17, which helps recruit HOPS to the autophagosome (Jiang et al, 2014; Takats et al, 2014). A role for the HOPS-Stx17 interaction in autophagy has subsequently been confirmed in a mouse model (Zhen and Li, 2015). HOPS has also been proposed to be linked to autophagosomes in mammalian cells by another Rab7 effector, Pleckstrin homology domain containing protein family member 1 (PLEKHM1).…”

CORVET, CHEVI and HOPS – multisubunit tethers of the endo-lysosomal system in health and disease

“…This has been shown for Vps3, Vps18, Vps33A and Vps41, and is likely the case for additional subunits. Point mutations and conditional knockouts, however, exhibit less-severe phenotypes, and allow researchers to study defects in animal models (Table S2) (Cai et al, 2016;Peng et al, 2012b;Suzuki et al, 2003;Zhen and Li, 2015). In addition, an increasingly growing range of phenotypes in patients is attributed to mutations in CORVET, CHEVI or HOPS components (Fig.…”

“…Other causative genes for HPS encode AP-3 subunits, which we discussed above in the context of Vps41 function. The bf mouse mutation is particularly interesting because it leads to defects in LRO biogenesis and autophagy, while transport to lysosomes and lysosomal activity remains intact (Zhen and Li, 2015).…”