EXPLOITATION OF MICROFILAMENT PROTEINS BY LISTERIA-MONOCYTOGENES - MICROVILLUS-LIKE COMPOSITION OF THE COMET TAILS AND VECTORIAL SPREADING IN POLARIZED EPITHELIAL SHEETS

Effective cell-to-cell spreading of the facultative intracellular path ogen Listeria monocytogenes requires the interaction between bacteria and the microfilament system of the host cell. By recruiting actin fil aments into a 'comet tail' localized at one pole of the bacterial cell wall, Listeria become mobile and propel themselves through the cytopl asm. They create protrusions at the plasma membrane that can invaginat e adjacent cells. In this work, we have analysed the structural compos ition of Listeria-recruited microfilaments in various epithelial cell lines by immunofluorescence microscopy. The microfilament-crosslinking proteins alpha-actinin, fimbrin and villin were localized around bact eria as soon as actin filaments could be detected on the bacterial sur face. Surprisingly, the same was found for ezrin/radixin, proteins inv olved in linking microfilaments to the plasma membrane. We found that in a polarized cell line derived from brush border kidney epithelium ( LLC-PK1), the actin filaments surrounding intracytoplasmic motile bact eria show the same immunoreactivity as the brush border-like microvill i, when analysed by a specific actin antibody. The successful invasion of polarized LLC-PK1 islets is vectorial, i.e. it progresses predomin antly from the periphery of the islets towards the centre. Infection o f the peripheral cells is sufficient for infiltration of the entire ce llular islets, without any further contact with the extracellular mili eu. This is in contrast to nonpolarized epithelial sheets, which can b e invaded from the apical surface of any individual cell. The importan ce of active bacterial motility in this vectorial spreading is emphasi zed by our finding that an isogenic Listeria mutant that is unable to recruit actin filaments cannot colonize polarized epithelial layers bu t accumulates in the peripheral cells of the islets.