title: Ubiquitin-specific protease USP2-45 acts as a molecular switch to promote α2δ-1-induced downregulation of Cav1.2 channels.
abstract: Availability of voltage-gated calcium channels (Cav) at the plasma membrane is paramount to maintaining the calcium homeostasis of the cell. It is proposed that the ubiquitylation/de-ubiquitylation balance regulates the density of ion channels at the cell surface. Voltage-gated calcium channels Cav1.2 have been found to be ubiquitylated under basal conditions both in vitro and in vivo. In a previous study, we have shown that Cav1.2 channels are ubiquitylated by neuronal precursor cell-expressed developmentally downregulated 4 (Nedd4-1) ubiquitin ligases, but the identity of the counterpart de-ubiquitylating enzyme remained to be elucidated. Regarding sodium and potassium channels, it has been reported that the action of the related isoform Nedd4-2 is counteracted by the ubiquitin-specific protease (USP) 2-45. In this study, we show that USP 2-45 also de-ubiquitylates Cav channels. We co-expressed USPs and Cav1.2 channels together with the accessory subunits β2 and α2δ-1, in tsA-201 and HEK-293 mammalian cell lines. Using whole-cell current recordings and surface biotinylation assays, we show that USP2-45 specifically decreases both the amplitude of Cav currents and the amount of Cav1.2 subunits inserted at the plasma membrane. Importantly, co-expression of the α2δ-1 accessory subunit is necessary to support the effect of USP2-45. We further show that USP2-45 promotes the de-ubiquitylation of both Cav1.2 and α2δ-1 subunits. Remarkably, α2δ-1, but not Cav1.2 nor β2, co-precipitated with USP2-45. These results suggest that USP2-45 binding to α2δ-1 promotes the de-ubiquitylation of both Cav1.2 and α2δ-1 subunits, in order to regulate the expression of Cav1.2 channels at the plasma membrane.
authors.affiliation: Department of Neurosciences, Physiology, and Pharmacology, University College London, London, WC1E 6BT, UK.
authors.forename: Jean-Sebastien
authors.initials: JS
authors.lastname: Rougier
authors.forename: Maxime
authors.initials: M
authors.lastname: Albesa
authors.forename: Ninda
authors.initials: N
authors.lastname: Syam
authors.forename: Guillaume
authors.initials: G
authors.lastname: Halet
authors.forename: Hugues
authors.initials: H
authors.lastname: Abriel
authors.forename: Patricia
authors.initials: P
authors.lastname: Viard
mesh_terms: D000818:Animals; D015153:Blotting, Western; D020746:Calcium Channels, L-Type / Q000096:biosynthesis*; D015536:Down-Regulation; D005786:Gene Expression Regulation / Q000502:physiology*; D057809:HEK293 Cells; D006801:Humans; D047468:Immunoprecipitation; D018408:Patch-Clamp Techniques; D011817:Rabbits; D014162:Transfection; D064570:Ubiquitin-Specific Proteases / Q000378:metabolism*; D054875:Ubiquitination
chemical_list: D020746:Calcium Channels, L-Type; C419385:L-type calcium channel alpha(1C); D064570:Ubiquitin-Specific Proteases