Heat shock factor 4 regulates lysosome activity by modulating the αB-crystallin-ATP6V1A-mTOR complex in ocular lens

• Published in: Biochimica et biophysica acta. General subjects Vol. 1864; no. 3; p. 129496
• Main Authors: Cui, Xiukun, Feng, Ruiping, Wang, Jungai, Du, Chunxiao, Pi, Xiahui, Chen, Danling, Li, Jing, Li, Hui, Zhang, Jun, Zhang, Jing, Mu, Hongmei, Zhang, Fengyan, Liu, Mugen, Hu, Yanzhong
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.03.2020
• Subjects: acidification; ATP6V1A; cataract; cell lines; Danio rerio; dissociation; epithelial cells; eye lens; germ cells; H-transporting ATP synthase; half life; heat shock proteins; immunoblotting; Lens epithelial cells; Lysosome acidification; lysosomes; mice; mTOR; mutation; phosphorylation; proteasome endopeptidase complex; protein-protein interactions; proteolysis; rapamycin; reverse transcriptase polymerase chain reaction; small interfering RNA; ubiquitination; Zebrafish; αB-crystallin; ATP6V1A; mTOR; Zebrafish; Lens epithelial cells; Lysosome acidification; αB-crystallin
• ISSN: 0304-4165; 1872-8006; 1872-8006
• DOI: 10.1016/j.bbagen.2019.129496

Germline mutations in heat shock factor 4 (HSF4) cause congenital cataracts. Previously, we have shown that HSF4 is involved in regulating lysosomal pH in mouse lens epithelial cell in vitro. However, the underlying mechanism remains unclear. HSF4-deficient mouse lens epithelial cell lines and zebrafish were used in this study. Immunoblotting and quantitative RT-PCR were used for expression analysis. The protein-protein interactions were tested with GST-pull downs. The lysosomes were fractioned by ultracentrifugation. HSF4 deficiency or knock down of αB-crystallin elevates lysosomal pH and increases the ubiquitination and degradation of ATP6V1A by the proteasome. αB-crystallin localizes partially in the lysosome and interacts solely with the ATP6V1A protein of the V1 complex of V-ATPase. Furthermore, αB-crystallin can co-precipitate with mTORC1 and ATP6V1A in GST pull down assays. Inhibition of mTORC1 by rapamycin or siRNA can lead to dissociation of αB-crystallin from the ATP6V1A and mTORC1complex, shortening the half-life of ATP6V1A and increasing the lysosomal pH. Mutation of ATP6V1A/S441A (the predicted mTOR phosphorylation site) reduces its association with αB-crystallin. In the zebrafish model, HSF4 deficiency reduces αB-crystallin expression and elevates the lysosomal pH in lens tissues. HSF4 regulates lysosomal acidification by controlling the association of αB-crystallin with ATP6V1A and mTOR and regulating ATP6V1A protein stabilization. This study uncovers a novel function of αB-crystallin, demonstrating that αB-crystallin can regulate lysosomal ATP6V1A protein stabilization by complexing to ATP6V1A and mTOR. This highlights a novel mechanism by which HSF4 regulates the proteolytic process of organelles during lens development. •Germline mutations in HSF4 gene causes congenital cataracts failing in fiber cell differentiation.•Deficiency of HSF4 down regulates alpha B-crystallin resulting in elevating lysosomal pH.•Alpha B-crystallin associates with and regulates ATP6V1A protein stabilization.•mTOR is involved in regulating the association of alpha B-crystallina and ATP6V1A.