Department of Medicinal Chemistry and Molecular Pharmacology, Purdue University, West Lafayette, Indiana.
Department of Medicinal Chemistry and Molecular Pharmacology, Purdue University, West Lafayette, Indiana
Mol Pharmacol. 2020 Dec;98(6):677-685. doi: 10.1124/molpharm.120.000061. Epub 2020 Oct 2.
Regulator of G protein signaling (RGS) proteins are negative modulators of G protein signaling that have emerged as promising drug targets to improve specificity and reduce side effects of G protein-coupled receptor-related therapies. Several small molecule RGS protein inhibitors have been identified; however, enhancing RGS protein function is often more clinically desirable but presents a challenge. Low protein levels of RGS2 are associated with various pathologies, including hypertension and heart failure. For this reason, RGS2 is a prominent example wherein enhancing its function would be beneficial. RGS2 is rapidly ubiquitinated and proteasomally degraded, providing a point of intervention for small molecule RGS2-stabilizing compounds. We previously identified a novel cullin-RING E3 ligase utilizing F-box only protein 44 (FBXO44) as the substrate recognition component. Here, we demonstrate that RGS2 associates with FBXO44 through a stretch of residues in its N terminus. RGS2 contains four methionine residues close to the N terminus that can act as alternative translation initiation sites. The shorter translation initiation variants display reduced ubiquitination and proteasomal degradation as a result of lost association with FBXO44. In addition, we show that phosphorylation of Ser may be an additional mechanism to protect RGS2 from FBXO44-mediated proteasomal degradation. These findings contribute to elucidating mechanisms regulating steady state levels of RGS2 protein and will inform future studies to develop small molecule RGS2 stabilizers. These would serve as novel leads in pathologies associated with low RGS2 protein levels, such as hypertension, heart failure, and anxiety. SIGNIFICANCE STATEMENT: E3 ligases provide a novel point of intervention for therapeutic development, but progress is hindered by the lack of available information about specific E3-substrate pairs. Here, we provide molecular detail on the recognition of regulator of G protein signaling protein 2 (RGS2) by its E3 ligase, increasing the potential for rational design of small molecule RGS2 protein stabilizers. These would be clinically useful in pathologies associated with low RGS2 protein levels, such as hypertension, heart failure, and anxiety.
G 蛋白信号调节蛋白（RGS）是 G 蛋白信号的负调节剂，它们已成为有前途的药物靶点，可以提高特异性并减少与 G 蛋白偶联受体相关治疗的副作用。已经鉴定出几种小分子 RGS 蛋白抑制剂；然而，增强 RGS 蛋白的功能通常更符合临床需求，但这是一个挑战。RGS2 的蛋白质水平低与多种病理有关，包括高血压和心力衰竭。出于这个原因，RGS2 是一个突出的例子，增强其功能将是有益的。RGS2 被迅速泛素化和蛋白酶体降解，为小分子 RGS2 稳定化合物提供了一个干预点。我们之前发现了一种新型的含有环指的 E3 连接酶，利用仅含有 F-box 的蛋白 44（FBXO44）作为底物识别成分。在这里，我们证明 RGS2 通过其 N 端的一段残基与 FBXO44 相关联。RGS2 包含靠近 N 端的四个甲硫氨酸残基，可作为替代翻译起始位点。较短的翻译起始变体由于与 FBXO44 失去关联而导致泛素化和蛋白酶体降解减少。此外，我们还表明丝氨酸的磷酸化可能是保护 RGS2 免受 FBXO44 介导的蛋白酶体降解的另一种机制。这些发现有助于阐明调节 RGS2 蛋白稳定状态的机制，并为开发小分子 RGS2 稳定剂的未来研究提供信息。这些将成为与 RGS2 蛋白水平低相关的病理学的新型先导物，例如高血压、心力衰竭和焦虑症。
E3 连接酶为治疗开发提供了一个新的干预点，但由于缺乏有关特定 E3-底物对的可用信息，进展受到阻碍。在这里，我们提供了 RGS2 与其 E3 连接酶识别的分子细节，增加了合理设计小分子 RGS2 蛋白稳定剂的潜力。这些在与 RGS2 蛋白水平低相关的病理学中具有临床应用价值，例如高血压、心力衰竭和焦虑症。
