Transient Receptor Potential Melastatin 2 Regulates Phagosome Maturation and Is Required for Bacterial Clearance in Escherichia coli Sepsis

Abstract

Background Transient receptor potential melastatin 2 is a Ca2+-permeable cation channel abundantly expressed in macrophages. Trpm2−/− mice showed exacerbated infection and mortality during polymicrobial sepsis, which is associated with inefficient bacterial killing in macrophages. However, the mechanism of transient receptor potential melastatin 2 regulating bacterial killing remains unknown. Methods Trpm2 −/− mice were intraperitoneally injected with Escherichia coli. The survival rate (n = 21) and bacterial burden (n = 5) were assessed. The processes of phagosome maturation and phagosome–lysosome fusion in peritoneal macrophages were extensively studied. The impact of increasing intracellular Ca2+ concentration on bacterial clearance in macrophages (n = 3) and on survival rate of Trpm2−/− mice infected with E. coli (n = 21) was investigated. Results Trpm2 −/− mice exhibited increased mortality (85% vs. 54%; P < 0.01) and aggravated bacterial burden during E. coli sepsis. Trpm2−/− peritoneal macrophages infected with E. coli showed dampened recruitment of lysosomal-associated membrane protein 1 and impaired phagosome maturation evidenced by a decrease in the accumulation of early endosome antigen 1, whereas a normal acquisition of Ras-related protein in brain 5. Increasing the cytosolic Ca2+ concentration in Trpm2−/− peritoneal macrophages via ionomycin treatment facilitated early endosome antigen 1 recruitment to Ras-related protein in brain 5 and phagosomal localization of lysosomal-associated membrane protein 1 and consequently enhanced bactericidal activity. Adoptive transfer of ionomycin-treated Trpm2−/− peritoneal macrophages improved bacterial clearance and survival (67% vs. 29%; P < 0.01) in Trpm2−/− mice challenged with E. coli. Conclusions Transient receptor potential melastatin 2 plays a critical role in host defense against invading bacteria via promoting phagosome maturation through facilitation of early endosome antigen 1 recruitment.

Role of the phagosomal redox-sensitive TRP channel TRPM2 in regulating bactericidal activity of macrophages

“…Zhan et al have also observed defective E. coli killing in stimulated peritoneal Trpm2 −/− macrophages, which they ascribed to defective phagosome-lysosome fusion resulting from impaired Ca 2+ influx via the plasma membrane-localized TRPM2 (Zhang et al, 2017). In the present study however, on carefully examining the distribution and function of TPRM2, we observed localization of TRPM2 in phagosomes but not in lysosomes of macrophages derived from bone marrow.…”

“…The reasons for the discrepancy in the findings are not clear. One possibility is that the stimulated peritoneal macrophages used by Zhang et al (2017) may have been activated prior to their use, and thus this favored fusion of phagosomes and lysosomes.…”

“…This includes recruiting the vATPase that pumps H + into phagosomes, and assembly and activation of phagosomal membrane NADPH oxidases (NOXs) that produce ROS (Russell et al, 2009). In light of the role of ROS in bacteria killing (Pollock et al, 1995;Segal, 1996;Segal et al, 2000), the previous observation that the NOXinhibiting function of TRPM2 promoted bacterial killing during phagocytosis at the plasma membrane (Di et al, 2011;Zhang et al, 2017) led us to consider the possibility that TRPM2 could also mediate bacterial killing through a phagosomal acidification mechanism. Phagosomal acidification in macrophages is essential for activation of lysosomal proteases, microbe killing and protein degradation (Aronson and De Duve, 1968;Coffey and De Duve, 1968;Pillay et al, 2002;Savina et al, 2006).…”

“…Impaired production of cytokines in Trpm2 −/− mice led to high mortality following Listeria monocytogenes (LM) challenge (Knowles et al, 2011). TRPM2 may also control the killing of internalized Escherichia coli through its role as a plasma membrane Ca 2+ -influx channel and induce phagosome-lysosome fusion (Zhang et al, 2017). Because of the importance of macrophages in killing bacteria, here, we addressed the question of whether TRPM2 is expressed in phagosomal membrane of macrophages and whether it functions as an efflux of cation channel to regulate phagosomal pH and thereby bacterial killing.…”

TRPM2 participates the transformation of acute pain to chronic pain during injury‐induced neuropathic pain

“…TRPM2-mediated calcium influx controls the ROS-induced signaling cascade, results in chemokine production in monocytes and neutrophil infiltration, and leads to aggravated inflammation in dextran sulfate sodium colitis (Yamamoto et al, 2008). TRPM2 also participated in various physiological processes, such as insulin secretion from pancreatic beta cells, phagosome maturation in sepsis model, and cellular depolarization in phagocytes (Di et al, 2011;Togashi et al, 2006;Zhang, Cui, Zhang, Chen, & Fang, 2017).…”