Misrouting of v-ATPase subunit V0a1 dysregulates lysosomal acidification in a neurodegenerative lysosomal storage disease model

Maria B Bagh,Zhongjian Zhang,Aiyi Liu,Nagarajan Pattabiraman,Goutam Chandra,Anil B Mukherjee,Satya P Singh,Shiyong Peng

doi:10.1038/ncomms14612

Abstract

Defective lysosomal acidification contributes to virtually all lysosomal storage disorders (LSDs) and to common neurodegenerative diseases like Alzheimer’s and Parkinson’s. Despite its fundamental importance, the mechanism(s) underlying this defect remains unclear. The v-ATPase, a multisubunit protein complex composed of cytosolic V1-sector and lysosomal membrane-anchored V0-sector, regulates lysosomal acidification. Mutations in the CLN1 gene, encoding PPT1, cause a devastating neurodegenerative LSD, INCL. Here we report that in Cln1−/− mice, which mimic INCL, reduced v-ATPase activity correlates with elevated lysosomal pH. Moreover, v-ATPase subunit a1 of the V0 sector (V0a1) requires palmitoylation for interacting with adaptor protein-2 (AP-2) and AP-3, respectively, for trafficking to the lysosomal membrane. Notably, treatment of Cln1−/− mice with a thioesterase (Ppt1)-mimetic, NtBuHA, ameliorated this defect. Our findings reveal an unanticipated role of Cln1 in regulating lysosomal targeting of V0a1 and suggest that varying factors adversely affecting v-ATPase function dysregulate lysosomal acidification in other LSDs and common neurodegenerative diseases.

Highlights

Defective lysosomal acidification contributes to virtually all lysosomal storage disorders (LSDs) and to common neurodegenerative diseases like Alzheimer’s and Parkinson’s
Our results show that lysosomal membraneanchored V0 sector isoform a1 (V0a1) subunit of v-ATPase undergoes S-palmitoylation, which is required for its sorting and trafficking to the lysosomal membrane
Since in our study we used Cln[1] À / À mice[23], which mimic infantile NCL (INCL), as the model for neurodegenerative LSDs, we first determined whether lysosomal acidification is dysregulated in these animals

Summary

Introduction

Defective lysosomal acidification contributes to virtually all lysosomal storage disorders (LSDs) and to common neurodegenerative diseases like Alzheimer’s and Parkinson’s. The infantile NCL (INCL)[13], a devastating neurodegenerative LSD, is caused by inactivating mutations in the CLN1 gene[14], which encodes palmitoyl-protein thioesterase-1 (PPT1)[15]. We tested a hypothesis that one or more subunits of v-ATPase requires S-palmitoylation for endosomal sorting, trafficking and reversible assembly of V1/V0 on lysosomal membrane, which is essential for regulating lysosomal pH and that Ppt[1] deficiency disrupts v-ATPase activity impairing its proton transport function, thereby dysregulating acidification of lysosomal lumen. Our results show that lysosomal membraneanchored V0 sector isoform a1 (V0a1) subunit of v-ATPase undergoes S-palmitoylation, which is required for its sorting and trafficking to the lysosomal membrane This process appears to be defective in Ppt1-deficient Cln[1] À / À mice. We demonstrate that treatment of these mice with a thioesterase (Ppt1)mimetic small molecule, N-tert (Butyl) hydroxylamine (NtBuHA), restores near-normal v-ATPase activity and rescues the defective lysosomal acidification phenotype

Methods

Results

Conclusion