Histamine H₁ Receptor-Mediated CREB Phosphorylation via G_q 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 G_q 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 G_q 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 G_q protein/Ca²⁺/PKC-dependent ERK and JNK activation; arrestins can modulate this process by regulating basal CREB phosphorylation.