Functional Characterisation of the role of Protein Tyrosine Phosphatase CD148 in Macrophages

Protein tyrosine phosphatases (PTPs) represent a biochemical counterbalance to the activity of protein tyrosine kinases, thus regulating the dynamic phosphorylation state of a cell and controlling diverse physiological processes. In macrophages, tyrosine phosphorylation regulates many signalling pathways including growth, differentiation, activation, phagocytosis and adhesion. This study aimed at the functional characterisation of the receptor protein tyrosine phosphatase CD148 in macrophages. The transcriptional regulation of ptprj (which encodes for CD148) in mouse and human mononuclear phagocytic cells was characterised. The expression of ptprj was found to be highest in macrophages, myelomonocytic cell lines and macrophage-enriched tissues. Ptprj expression was down-regulated by the macrophage growth and differentiation factor macrophage colony stimulating factor-1 (CSF-1) and significantly up-regulated by bacterial components such as lipopolysaccharide (LPS) and CpG DNA. Both ptprj transcript and protein were induced and found to be associated with disease progression in mouse chronic inflammatory disease models. In vitro knockdown studies of ptprj using RNAi constructs confirmed that ptprj regulates macrophage activation and proliferation. Attempts to overexpress ptprj in mice in a macrophage-specific manner were unsuccessful, further suggesting that ptprj inhibits macrophage proliferation and/or differentiation. Following stimulation of bone-marrow derived macrophages (BMMs) with CSF-1 and LPS, there was a redistribution and association of CD148 with membrane ruffles, F-actin and Pyk2 suggesting a role for CD148 in regulating macrophage morphology in response to these stimuli. Treatment of BMMs with anti-CD148 monoclonal antibody inhibited CSF-1 induced macrophage spreading, cytoskeletal changes and chemotaxis, without affecting CSF-1 receptor downstream signalling and cell survival, thereby negatively regulating a subset of CSF-1 receptor responses in macrophages. The results are consistent with the hypothesis that CD148 is a regulator of macrophage activity and inflammation.