Cellular cholesterol involvement in Src, PKC, and p38/JNK transduction pathways by porins

Biological membranes are described as a mosaic of different domains where interactions between membrane components induce the formation of subdomains with different characteristics and functions. Lipids play an important role in the formation of lipid-enriched microdomains where they dynamically associate to form platforms important for membrane protein sorting and construction of signaling complexes. Cholesterol confined in lipid domains is a crucial component required by microorganisms, directly or indirectly, to enter or exit the intracellular compartment. Cellular activation mediated by superficial bacterial component may be modified by local cholesterol depletion. Therefore, new perspectives for unconventional therapeutic intervention in Gram-negative infections may be envisaged. We tested this hypothesis by using methyl-beta-cyclodextrin (mbetaCD) as a cholesterol-complexing agent to alter the U937 plasma membrane cholesterol content. Our results demonstrate that cholesterol depletion of U937 cells inhibited Salmonella enterica serovar Typhimurium porins-mediated phosphorylation of Src kinase family, protein kinase C (PKC), JNK, and p38, while cholesterol repletion restored the phosphorylation. Lipopolysaccharide (LPS) extracted from the same bacterial strain has been used as a control. Our data demonstrate that the lack of activation of signal transduction pathway observed following cholesterol depletion differently modulates the release of interleukin-6 (IL-6) or tumor necrosis factor-alpha (TNF-alpha), suggesting that Src, associated to lipid domains, may represent an important pathway in Gram-negative-induced cellular signal.