AP-1 and NF-κB regulate osteopontin gene expression in anti-CD40 treated B cells (161.16)

Abstract

Abstract Osteopontin (OPN) is a glycoprotein that is implicated in many physiological and pathological processes. Elevated levels of OPN are linked to autoimmunity and cancer and correlate with disease progression. In the immune system, macrophages, dendritic cells, T and B cells express OPN. The regulatory elements involved in B cell OPN expression are poorly understood. Our objective is to determine the regulatory elements critical for OPN expression in B cells. B cells express OPN when stimulated with anti-Ig post treatment with IL-4 or CD40L. To determine the regulatory elements, we made four overlapping OPN promoter-luciferase constructs extending from -2000bp to +80bp. We then transfected 1C10 (anti-CD40 antibody) treated and anti-Ig stimulated primary murine B cells with the OPN promoter constructs. We saw an increase in luciferase activity in the promoter construct containing binding sites for AP-1, Ets and Runx2. Significantly, mutagenesis of the AP-1 site abrogated this increase in luciferase activity. To investigate the role of NF-κB in OPN expression, we used an NF-κB inhibitor to evaluate OPN expression in B cells stimulated with anti-Ig post treatment with1C10. NF-κB inhibition reduced OPN transcript expression in these cells. In conclusion, preliminary data indicate that AP-1 and NF-κB play a role in OPN expression in anti-CD40 treated B cells.