P53 Negatively Regulates RGS13 Protein Expression in Immune Cells

Zhihui Xie,Shoko Iwaki,Yunbiao Lu,Kirk M Druey

doi:10.1074/jbc.m111.228924

Abstract

RGS13, a member of the regulator of G protein signaling (RGS) family, inhibits G protein-coupled receptor signaling in B cells and mast cells (MCs) and suppresses IgE-antigen-induced MC degranulation and anaphylaxis. Although RGS13 expression is induced by immune receptor and chemokine receptor stimulation, the molecular regulation of RGS13 transcription is unknown. Here, we investigated the role of two p53 response elements (REs) in the regulation of RGS13 promoter activity and expression. We found that a 1000-bp DNA fragment upstream of the ATG translation start site (TSS) had promoter activity in reporter gene assays, and deletion or mutation of a p53-binding motif nearest the TSS abolished promoter activity. Notably, p53 bound to both REs in the RGS13 promoter in vivo as assessed by chromatin immunoprecipitation, suggesting that the p53 RE most distal to the TSS is physiologically inactive. We detected reduced RGS13 expression in MCs exogenously expressing p53 or treated with doxorubicin, which induces genotoxic stress and leads to p53 accumulation. RNA silencing of p53 up-regulated RGS13 expression in B lymphocytes, and bone marrow-derived MCs from p53(-/-) mice had increased RGS13 expression. Finally, p53-depleted B cells with increased RGS13 expression had reduced Ca(2+) mobilization in response to sphingosine 1-phosphate. These studies indicate that p53 may modulate immune responses through suppression of RGS13 transcription in MCs and B cells.

Highlights

G protein-coupled receptor (GPCR)3 signaling pathways are initiated by exchange of GDP for GTP on the ␣-subunit of the heterotrimeric G protein, whereas signal termination is mediated by GTP hydrolysis by the G␣ subunit [1]
Given the relatively unknown function of p53 in the regulation of gene transcription in mast cells (MCs) as well as the previous identification of RGS16 as a p53 target gene [17], we focused on the role of p53 in RGS13 transcription in the remainder of this study
RGS13 meets the definition of a p53 response gene according to published criteria [10]: 1) it contains putative p53 response elements (REs) in its upstream promoter region; 2) manipulation of p53 expression or elimination of the p53-binding motif modifies RGS13 promoter activity; 3) p53 binds to the identified RE in vivo as assessed by ChIP; and 4) manipulation of p53 levels leads to changes in the expression of both RGS13 mRNA and protein

Summary

Introduction

G protein-coupled receptor (GPCR)3 signaling pathways are initiated by exchange of GDP for GTP on the ␣-subunit of the heterotrimeric G protein, whereas signal termination is mediated by GTP hydrolysis by the G␣ subunit [1]. Treatment of the human MC line LAD2 with a permeable cAMP analog, which activates the cAMP response element-binding protein pathway, reduces RGS13 mRNA expression [9].

Results

Conclusion