Lipid-Based Signaling Modulates DNA Repair Response and Survival against Klebsiella pneumoniae Infection in Host Cells and in Mice

Abstract

Klebsiella pneumoniae causes serious infections in the urinary tract, respiratory tract, and blood. Lipid rafts, also known as membrane microdomains, have been linked to the pathogenesis of bacterial infection. However, whether lipid rafts affect K. pneumoniae internalization into host cells remains unknown. Here, we show for the first time that K. pneumoniae was internalized into lung cells by activating lipid rafts. Disrupting lipid rafts by methyl-b-cyclodextrin inhibited pathogen internalization, impairing host defense. A deficient mutant of capsule polysaccharide (CPS) showed a higher internalization rate than a wild-type strain, indicating that CPS may inhibit bacterial entry to host cells. Furthermore, lipid rafts may affect the function of extracellular regulated kinase (ERK)-1/2, and knocking down ERK1/ 2 via short, interfering RNA increased apoptosis in both alveolar macrophages and epithelial cells after infection. To gain insights into bacterial pathogenesis, we evaluated the impact of lipid rafts on DNA integrity, and showed that raft aggregates also affect DNA damage and DNA repair responses (i.e., 8-oxoguanine DNA glycosylase [Ogg1]) through the regulation of reactive oxygen species. Importantly, cells overexpressing Ogg1 demonstrated reduced cytotoxicity during bacterial infection. Taken together, these results suggest that lipid rafts may modulate bacterial internalization, thereby affecting DNA damage and repair, which is critical to host defense against K. pneumoniae.Keywords: Klebsiella pneumoniae infection; internalization; alveolar epithelial cells; lipid rafts; capsule polysaccharide Klebsiella pneumoniae causes serious infections in multiorgan systems, and is the third most commonly isolated bacterium from the blood of patients with sepsis (1). Because K. pneumoniae rapidly develops multidrug-resistant strains, this "superbug" can also cause outbreaks in intensive care units, imposing significant financial burdens and dangerous health threats (2, 3). Despite intensive research during the past few decades, the pathogenesis of K. pneumoniae, including its internalization of alveolar cells, is incompletely understood. K. pneumoniae expresses two critical antigens on its cell surface, LPS (an O antigen) and capsule polysaccharide (CPS; a K antigen) (4). These two antigens contribute to pathogenicity through interactions with host cells in different manners (5). LPS is a major component of Gramnegative bacterial cell walls with high immunogenicity, but its role in pathogenesis remains elusive. Early studies suggest that LPS-deficient strains failed to show significantly different pathogenesis from the wild-type (WT) strain (5). By contrast, CPS demonstrates the most distinguished characteristic of virulence factors for K. pneumoniae, and is implicated in the internalization of pathogens into epithelial cells (6). The capsule may protect the bacterium from phagocytosis by macrophages, through an escape from bactericidal serum factors (6). Other studies also demonstrated that isogenic CPS mutant...

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