IDENTIFICATION OF COFILIN, CORONIN, RAC AND CAPZ IN ACTIN TAILS USINGA LISTERIA AFFINITY APPROACH

Actin assembly is involved in cell motility and intracellular movement of Listeria monocytogenes. Induction of Lister ia actin tails is medi ated by the surface protein ActA. The N-terminal domain of ActA is suf ficient for this function. Cell components known to play a role in the actin-based motility of Listeria are VASP (vasodilatator-stimulated p hosphoprotein), the multiprotein Arp2/3 complex and cofilin, VASP inte racts with the central domain of ActA, Proteins interacting with the N -terminal domain of ActA have not been identified. To identify novel h ost cell components of ActA-induced actin tails, we used bovine brain extracts and an affinity approach with Listeria as matrix. Several kno wn components of Listeria tails were isolated including VASP, Arp3 and cofilin. Cofilin was identified by peptide sequencing, and cofilin re cruitment and Listeria tail length were found to be pH-dependent, in a greement with its recently reported role in enhancing actin filament t urnover. In addition, three proteins not previously known to be associ ated with Listeria tails, coronin, Rac and capZ, were identified in ou r affinity approach. In infected cells, the localization of the identi fied proteins was studied by immunofluorescence. Our findings suggest that these latter proteins, which are known to play critical roles in cellular actin rearrangements, may also be involved in the dynamics of Listeria-induced actin assembly.