Ubiquitin ligase RNF138 promotes episodic ataxia type 2-associated aberrant degradation of human Ca_v2.1 (P/Q-type) calcium channels

Voltage-gated Ca_V2.1 channels comprise a pore-formingα_1A subunit with auxiliaryα₂δ and β subunits. Ca_V2.1 channels play an essential role in regulating synaptic signaling. Mutations in the human gene encoding the Ca_V2.1 subunit are associated with the cerebellar disease episodic ataxia type 2 (EA2). Several EA2-causing mutants exhibit impaired protein stability and exert dominant-negative suppression of Ca_V2.1 wild-type (WT) protein expression via aberrant proteasomal degradation. Here, we set out to delineate the protein degradation mechanism of human Ca_V2.1 subunit by identifying RNF138, an E3 ubiquitin ligase, as a novel Ca_V2.1-binding partner. In neurons, RNF138 and Ca_V2.1 coexist in the same protein complex and display notable subcellular colocalization at presynaptic and postsynaptic regions. Overexpression of RNF138 promotes polyubiquitination and accelerates protein turnover of Ca_V2.1. Disrupting endogenous RNF138 function with a mutant (RNF138-H36E) or shRNA infection significantly upregulates the Ca_V2.1 protein level and enhances Ca_V2.1 protein stability. Disrupting endogenous RNF138 function also effectively rescues the defective protein expression of EA2 mutants, as well as fully reversing EA2 mutant-induced excessive proteasomal degradation of Ca_V2.1WTsubunits. RNF138-H36E coexpression only partially restores the dominant-negative effect of EA2 mutants on Ca_V2.1WTfunctional expression, which can be attributed to defective membrane trafficking of Ca_V2.1 WT in the presence of EA2 mutants. We propose that RNF138 plays a critical role in the homeostatic regulation of Ca_V2.1 protein level and functional expression and that RNF138 serves as the primary E3 ubiquitin ligase promoting EA2-associated aberrant degradation of human Ca_V2.1 subunits.