A Female-specific Role of RGS20 in Transcriptional, Epigenomic and Behavioral Responses to Chronic Pain

The signal transduction protein named Regulator of G-protein Signaling-20 (RGS20) modulates functional responses of several GPCRs in the brain, by acting as a GTPase activator or as an effector antagonist for Ga subunits. The RGS20 variant, RGSz1, is highly expressed in the brain. Using genetic mouse models, genomic and proteomic approaches we identified a female-specific role of RGS20 in sensory hypersensitivity associated with chronic pain states. Constitutive knockout of RGS20, as well as conditional knockdown of RGS20 in the mouse periaqueductal gray (PAG), exacerbate mechanical allodynia and thermal hyperalgesia in models of peripheral inflammation (intraplantar injection of Complete Freud's adjuvant (CFA)) and nerve injury in female but not male mice. Furthermore, we show that prevention of RGS20 action delays recovery from thermal hypersensitivity in female mice after CFA treatment. In PAG tissue, biochemical studies revealed a sex-specific upregulation of RGSz1 levels and Chromatin Immunoprecipitation (ChIP)-qPCR showed decreased binding of ER elements on RGS20 promoter, 10 days after peripheral inflammation. RNA Sequencing analysis of PAG samples from RGS20WT and RGS20KO mice under naïve states or 10 days after CFA, uncovered that exacerbated sensory hypersensitivity upon RGS20 knockdown is associated with changes in the expression of genes and pathways associated with central sensitization and pain maintenance, including molecules involved in serotonin synthesis and release. Proteomic analysis of epigenetic marks in the female PAG, identified that, following CFA treatment, H3K14ac was increased in RGS20WT and decreased in RGS20KO animals. After validating these findings by western blot, we mapped H3K14ac binding with ChIP followed by high-throughput sequencing. We then monitored the expression of multiple target genes, including the histone acetyltransferase KAT2B that we found decreased in RGS20KO after CFA injection. Our studies highlight a novel female-specific intracellular pathway in the PAG which controls the severity of sensory hypersensitivity and recovery from chronic pain states.