RGS2 protein degradation is mediated by a novel Cullin 4B/F‐box 44 E3 ligase complex (843.8)

Hypertension and heart failure are major health issues and there is a need to identify more effective treatments and novel drug targets. R egulator of G protein S ignaling 2 (RGS2) is highly expressed in both heart and vascular smooth muscle and regulates Gα q signaling. RGS2 ‐/‐ mice are hypertensive, show enhanced responses to vasoconstrictors and lower tolerability to pressure overload. Over ‐expression of RGS2 reduces cardiac hypertrophy and we have shown that pharmacologically enhanced RGS2 protein expression has functional effects on G protein signaling. Therefore, we hypothesize that enhancing RGS2 protein expression is a novel route in treating cardiovascular disease. RGS2 is rapidly degraded through the proteasome, but the enzymes involved are not known. Thus, the goal of the current study was to identify the molecular machinery responsible for RGS2 protein degradation. Utilizing a β‐galactosidase complementation assay that efficiently quantifies levels of a protein we performed an siRNA screen to identify genes that regulate RGS2 protein levels. We identified components of a putative cullin‐RING E3 ligase (CRL). siRNA knock‐down of either the substrate recognition component F‐box 44 or cullin 4B (CUL4B), but not cullin 4A, resulted in a significant increase in RGS2 protein levels and stability and overexpression of F‐box 44 CUL4B decreased RGS2 protein levels. We also demonstrated co‐immunoprecipitation of F‐box 44 with both RGS2 and CUL4B. RGS proteins are difficult drug targets due to their mode of action being through protein‐protein interactions. Inhibiting a more druggable protein within the degradation pathway could be a way to increase RGS2 protein levels and function. Grant Funding Source : Supported by NIH‐DA023252 and The Swedish Heart‐ and Lung Foundation