Flagella methylation promotes bacterial adhesion and host cell invasion

Abstract

35The flagellum is the motility device of many bacteria and the long external filament is 36 made of several thousand copies of a single protein, flagellin. While posttranslational 37 modifications of flagellin are common among bacterial pathogens, the role of lysine 38 methylation remained unknown. Here, we show that both flagellins of Salmonella 39 enterica, FliC and FljB, are methylated at surface-exposed lysine residues. A 40 Salmonella mutant deficient in flagellin methylation was outcompeted for gut 41 colonization in a gastroenteritis mouse model. In support, methylation of flagellin 42 promoted invasion of epithelial cells in vitro. Lysine methylation increased the surface 43 hydrophobicity of flagellin and enhanced flagella-dependent adhesion of Salmonella to 44 phosphatidylcholine vesicles and epithelial cells. In summary, posttranslational flagellin 45 methylation constitutes a novel mechanism how flagellated bacteria facilitate adhesion 46 to hydrophobic host cell surfaces and thereby contributes to efficient gut colonization 47 and successful infection of the host. 48 49 50 51 52 53 54 Introduction: 57 The Gram-negative enteropathogen Salmonella enterica uses a variety of strategies to 58 successfully enter and replicate within a host. In this respect, bacterial motility enables 59 the directed movement of the bacteria towards nutrients or the target site of infection. A 60 rotary nanomachine, the flagellum, mediates motility of many bacteria, including 61 Salmonella enterica 1 . Flagella also play a central role in other infection processes, 62 involving biofilm formation, immune system modulation and adhesion 2-4 . 63 The eukaryotic plasma membrane plays an important role in the interaction of 64 flagellated bacteria with host cells during the early stages of infection 5 . The flagella of S. 65 enterica, Escherichia coli and Pseudomonas aeruginosa can function as adhesion 66 molecules 6-8 mediating the contact to various lipidic plasma membrane components, 67 including cholesterol, phospholipids, sulfolipids and the gangliosides GM1 and aGM1 9-68 12 . 69 Structurally, the flagellum consists out of three main parts: the basal body embedded 70 within the inner and outer membranes of the bacterium, a flexible linking structure -the 71 hook, and the long, external filament, which functions as the propeller of the motility 72 device 13 . The filament is formed by more than 20,000 subunits of a single protein, 73 flagellin. Many S. enterica serovars express either of two distinct flagellins, FliC or FljB, 74 in a process called flagellar phase variation 14,15 . FliC-expressing bacteria display a 75 distinct motility behavior on host cell surfaces and a competitive advantage in 76 colonization of the intestinal epithelia compared to FljB-expressing bacteria 16 . However, 77while the structure of FliC has been determined previously 17 , the structure of FljB 78 remained unknown. 130We next aligned the amino acid sequences of FljB and FliC up-and downstream of the 131 identified ɛ-N-methyl-lysine residues (...