Streptococcus pyogenes CAMP factor promotes calcium ion uptake in RAW264.7 cells

: Streptococcus pyogenes is a bacterium that causes systemic diseases such as pharyngitis and toxic shock syndrome. S. pyogenes produces molecules that inhibit the function of the human immune system, thus allowing growth and spread of the pathogen in tissues. It is known that S. pyogenes CAMP factor induces vacuolation in macrophages; however, the mechanism remains unclear. In the current study, the mechanism by which CAMP factor induces vacuolation in macrophages was investigated. CAMP factor was found to induce calcium ion uptake in murine macrophage RAW264.7 cells. In addition, EDTA inhibited calcium ion uptake and vacuolation in the cells. The L-type voltage-dependent calcium ion channel blockers nifedipine and verapamil reduced vacuolation. Furthermore, the phosphoinositide 3-kinase inhibitors LY294002 and wortmannin also inhibited the vacuolation induced by CAMP factor. Fluorescent microscopy revealed that clathrin localized to the vacuoles. These results suggest that the vacuolation is related to calcium ion uptake by RAW264.7 cells via L-type voltage-dependent calcium ion channels. Therefore, it was concluded that the vacuoles induced by S. pyogenes CAMP factor in macrophages are clathrin-dependent endosomes induced by activation of the phosphoinositide 3-kinase signaling pathway through calcium ion uptake.