Differential regulation of histamine H receptor-mediated ERK phosphorylation by G proteins and arrestins.

G protein-coupled histamine H receptors play crucial roles in allergic and inflammatory reactions, in which the phosphorylation of extracellular signal-regulated kinase (ERK) appears to mediate the production of inflammatory cytokines. ERK phosphorylation is regulated by G protein- and arrestin-mediated signal transduction pathways. Here, we aimed to explore how H receptor-mediated processes of ERK phosphorylation might be differentially regulated by G proteins and arrestins. For this purpose, we evaluated the regulatory mechanism(s) of H receptor-mediated ERK phosphorylation in Chinese hamster ovary cells expressing G protein- and arrestin-biased mutants of human H receptors, S487TR and S487A, in which the Ser487 residue in the C-terminal was truncated and mutated to alanine, respectively. Immunoblotting analysis indicated that histamine-induced ERK phosphorylation was prompt and transient in cells expressing G protein-biased S487TR, whereas it was slow and sustained in cells expressing arrestin-biased S487A. Inhibitors of G proteins (YM-254890) and protein kinase C (PKC) (GF109203X), and an intracellular Ca chelator (BAPTA-AM) suppressed histamine-induced ERK phosphorylation in cells expressing S487TR, but not those expressing S487A. Conversely, inhibitors of G protein-coupled receptor kinases (GRK2/3) (cmpd101), β-arrestin2 (β-arrestin2 siRNA), clathrin (hypertonic sucrose), Raf (LY3009120), and MEK (U0126) suppressed histamine-induced ERK phosphorylation in cells expressing S487A, but not those expressing S487TR. These results suggest that H receptor-mediated ERK phosphorylation might be differentially regulated by the G protein/Ca/PKC and GRK/arrestin/clathrin/Raf/MEK pathways to potentially determine the early and late phases of histamine-induced allergic and inflammatory responses, respectively.