G-protein coupled receptor kinases (GRKs) are serine-threonine protein kinases which phosphorylate agonist bound G-protein coupled receptors (GPCRs) leading to their desensitization. Although originally discovered with regard to GPCR desensitization, recent studies have shown that GRKs play much wider roles than previously appreciated. In this regard, studies have shown that GRKs can phosphorylate non GPCR receptor as well as non receptor substrates. There is an ample amount of evidence in the literature showing that the expression and activity of GRKs is altered in several inflammatory disease conditions. For instance, the activity and expression levels of GRK2 were found to be significantly decreased in peripheral blood mononuclear cells (PBMC) of patients with rheumatoid arthritis whereas the expression levels of GRK2 and GRK5 were found to be enhanced in the neutrophils of sepsis patients. Moreover in our previous studies we found that stimulation of primary peritoneal macrophages with LPS causes an increase in the expression of GRK2. These studies point to a crucial role of GRKs in inflammatory signaling, however, the physiological importance of changes in the expression levels of GRKs is not well established. To enhance our understanding of the role of GRK2 and 5 in inflammatory signaling, we first investigated the mechanism by which GRK2 and 5 regulate TNFα-induced inflammatory signaling in mouse macrophage cell line.

Our results in this study demonstrated that both GRK2 and 5 positively regulate TNFα-induced NFκB signaling. Knockdown of GRK2 and 5 inhibited TNFα-induced NFκB signaling whereas overexpression of GRK2 and 5 substantially enhanced TNFα-induced NFκB activity. Furthermore, GRK2 and 5 were found to interact with and phosphorylate IκBα, an inhibitor of NFκB thus regulating NFκB signaling.

To further elucidate the role of GRKs in inflammation under in vivo conditions, we utilized mice with homozygous GRK5 gene deletion. Primary peritoneal macrophages from GRK5^−/− mice stimulated with LPS showed an inhibition of NFκB activity as compared to cells from GRK5^+/+ mice. Secretion of several LPS-induced inflammatory cytokines was found to be reduced in cell culture supernatants of GRK5^−/− mice. Plasma levels of cytokines and chemokines were also found to be reduced which was associated with reduced liver injury in GRK5^−/− mice compared to GRK5 ^+/+ mice suggesting that GRK5 positively regulates LPS-induced inflammatory signaling in vivo by modulating transcription factor NFκB. Since homozygous gene deletion of GRK2 is lethal in mice, we utilized Cre-loxP system to achieve a cell specific deletion of GRK2 whereby GRK2 was specifically deleted in the cells of myeloid lineage. LPS injection into these mice caused an increased expression of cytokines/chemokines in the plasma as well as in the primary peritoneal macrophage cell culture supernatants. GRK2 deficient mice also exhibited an increased lung and liver injury. Mechanistically, IKKβ-NFκB1 p105/TPL2-MEK-ERK pathway was found to be negatively regulated by GRK2 which resulted in an increased expression of cytokines/chemokines in GRK2 deficient mice. Taken together, these results show that both GRK2 and 5 play a crucial role in LPS-induced inflammatory signaling under in vivo conditions by regulating NFκB and ERK signaling pathways.