@article{697d1747441b4706ba2664b3ab3cceca,
title = "Multiubiquitination of TRPV4 reduces channel activity independent of surface localization",
abstract = "Ubiquitin (Ub)-mediated regulation of plasmalemmal ion channel activity canonically occurs via stimulation of endocytosis. Whether ubiquitination can modulate channel activity by alternative mechanisms remains unknown. Here, we show that the transient receptor potential vanilloid 4 (TRPV4) cation channel is multiubiquitinated within its cytosolic N-terminal and C-terminal intrinsically disordered regions (IDRs). Mutagenizing select lysine residues to block ubiquitination of the N-terminal but not C-terminal IDR resulted in a marked elevation of TRPV4-mediated intracellular calcium influx, without increasing cell surface expression levels. Conversely, enhancing TRPV4 ubiquitination via expression of an E3 Ub ligase reduced TRPV4 channel activity but did not decrease plasma membrane abundance. These results demonstrate Ubdependent regulation of TRPV4 channel function independent of effects on plasma membrane localization. Consistent with ubiquitination playing a key negative modulatory role of the channel, gain-of-function neuropathy-causing mutations in the TRPV4 gene led to reduced channel ubiquitination in both cellular and Drosophila models of TRPV4 neuropathy, whereas increasing mutant TRPV4 ubiquitination partially suppressed channel overactivity. Together, these data reveal a novel mechanism via which ubiquitination of an intracellular flexible IDR domain modulates ion channel function independently of endocytic trafficking and identify a contributory role for this pathway in the dysregulation of TRPV4 channel activity by neuropathy-causing mutations.",
author = "Aisenberg, {William H.} and McCray, {Brett A.} and Sullivan, {Jeremy M.} and Erika Diehl and DeVine, {Lauren R.} and Jonathan Alevy and Bagnell, {Anna M.} and Patrice Carr and Donohue, {Jack K.} and Benedikt Goretzki and Cole, {Robert N.} and Hellmich, {Ute A.} and Sumner, {Charlotte J.}",
note = "Funding Information: Acknowledgments—We thank the Johns Hopkins Institute for Basic Biomedical Sciences Microscope Facility and Hoku West-Foyle for assistance with TIRF image acquisition. This facility was supported by the Office of the Director and the National Institute of General Medical Sciences of the National Institutes of Health under award number S10OD023548. We thank Dr Sandra Gabelli, Dr Phillip Cole, and Dr Zan Chen for their training, advice and reagents for cell-free ubiquitination assays. We thank Dr Thomas Lloyd for providing TRPV4-expressing Drosophila for in vivo ubiquitination assays and Dr Michael Caterina and Dr Hidde Ploegh for sharing TRPV4-null mice and HA-Ub mice, respectively. This study was funded by the Deutsche Forschungsgemeinschaft (DFG, German Research Foundation) under Germany{\textquoteright}s Excellence Strategy—EXC 2051—Project ID 390713860. Funding Information: Funding and additional information—This work was supported by F31 NS105404 (to W. H. A.) and R35 NS122306 (to C. J. S.). E. D. acknowledges a Fulbright PhD Fellowship, a TransMED PhD Fellowship, and a Sibylle Kalkhof-Rose Stiftung Fellowship. B. G. acknowledges a Max Planck Graduate Centre PhD Fellowship. U. A. H. acknowledges support by the Centre for Biomolecular Magnetic Resonance (BMRZ), Goethe University Frankfurt, funded by the state of Hesse. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. Funding Information: We thank the Johns Hopkins Institute for Basic Biomedical Sciences Microscope Facility and Hoku West-Foyle for assistance with TIRF image acquisition. This facility was supported by the Office of the Director and the National Institute of General Medical Sciences of the National Institutes of Health under award number S10OD023548. We thank Dr Sandra Gabelli, Dr Phillip Cole, and Dr Zan Chen for their training, advice and reagents for cell-free ubiquitination assays. We thank Dr Thomas Lloyd for providing TRPV4-expressing Drosophila for in vivo ubiquitination assays and Dr Michael Caterina and Dr Hidde Ploegh for sharing TRPV4-null mice and HA-Ub mice, respectively. This study was funded by the Deutsche Forschungsgemeinschaft (DFG, German Research Foundation) under Germany's Excellence Strategy-EXC 2051-Project ID 390713860. Publisher Copyright: {\textcopyright} 2022 THE AUTHORS",
year = "2022",
month = apr,
day = "1",
doi = "10.1016/j.jbc.2022.101826",
language = "English (US)",
volume = "298",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "4",
}