Histamine H1 Receptor-Mediated CREB Phosphorylation via Gq Protein Signaling and Arrestin Modulation.

Background/

Objectives

Histamine H1 receptors mediate multiple physiological and pathophysiological processes, including inflammation and allergy, by regulating downstream gene expression via transcription factors. cAMP response element-binding protein (CREB) is a major transcription factor whose phosphorylation is regulated by multiple signaling pathways. Although CREB is closely involved in multiple physiological and pathophysiological processes, the detailed intracellular signaling pathway of H1 receptor-mediated CREB phosphorylation remains to be elucidated. We investigated the roles of Gq proteins and arrestins in H1 receptor-mediated CREB phosphorylation.

Methods

We constructed Chinese hamster ovary (CHO) expressing human wild-type (WT) H1 receptors and two types of C-terminal mutants. One mutant was constructed by truncating the serine 487 residue only at the C-terminus (S487Trunc), and the other was constructed by substituting the serine 487 residue at the C-terminus with alanine (S487A). S487Trunc is a Gq protein-biased while S487A is an arrestin-biased receptor. The expressions of CREB and its phosphorylated form were assessed by immunoblotting.

Results

Histamine promoted CREB phosphorylation in CHO cells expressing WT or S487Trunc receptors, but not in cells expressing S487A. Inhibitors of protein kinase C (PKC), extracellular signal-regulated kinase (ERK), or c-Jun N-terminal kinase (JNK), and Ca2+ chelator suppressed histamine-induced CREB phosphorylation in CHO cells expressing WT or S487Trunc receptors. Basal CREB phosphorylation levels increased following β-arrestin overexpression and decreased after their siRNA-mediated knockdown, thus modulating histamine-stimulated CREB phosphorylation in WT CHO cells.

Conclusions

H1 receptor-mediated CREB phosphorylation is induced through Gq protein/Ca2+/PKC-dependent ERK and JNK activation; arrestins can modulate this process by regulating basal CREB phosphorylation.