Dynamic palmitoylation of H+-transporting V-ATPase subunit a1 regulates lysosomal acidification and is disrupted in Ppt1-/- mice

Wednesday, September 24, 2014 — Poster Session IV
10:00 a.m. –12:00 p.m.	FAES Academic Center	NICHD	CELLBIO-1

* FARE Award Winner

Authors

• M.B. Bagh
• G. Chandra
• Z. Zhang
• S. Peng
• A.B. Mukherjee

Abstract

Dynamic palmitoylation (palmitoylation-depalmitoylation) regulates the function of numerous proteins. While palmitoyl-acyltransferases catalyze palmitoylation, the thioesterases catalyze depalmitoylation. Genetic deficiency of PPT1, a thioesterase, impairs degradation of palmitoylated proteins by lysosomal hydrolases, which require acidic pH for catalytic activity leading to a neurodegenerative lysosomal storage disease (LSD), INCL. Although elevated lysosomal pH is reported in virtually all LSDs the molecular mechanism(s) remains unexplained. The vacuolar H+-transporting ATPase (V-ATPase), a multi-subunit protein that regulates lysosomal pH requires reversible assembly of its membrane-bound V0-sector and the cytosolic V1-sector. We uncovered that subunit a1 of the V0-sector (V0a1) requires palmitoylation on Cys-25 for localization to lysosomal membrane. Unexpectedly, in brain tissues of Ppt1-/- mice that mimic INCL, V0a1 is mislocalized to plasma membrane. Moreover, PPT1-deficiency impaired clathrin/AP2-mediated V0a1 trafficking to early endosomes inhibiting dissociation of V0a1 from clathrin/AP2/V0a1 complex. This prevented V0a1-AP3 interaction required for lysosomal targeting. Consequently, clathrin/AP2/V0a1 complex was recycled back to plasma membrane disrupting V0-V1 assembly on lysosomal membrane impairing v-ATPase function. Importantly, a PPT1-mimetic, NtBuHA, ameliorated these defects. Our findings demonstrate that PPT1-deficiency alters lysosomal pH in INCL by impairing dynamic palmitoylation of V0a1 and suggest that varying mechanism(s) disrupting V-ATPase function may underlie lysosomal pH abnormality in other LSDs.

back to top