Identification and characterization of cellular host kinases involved in Listeria monocytogenes cell-to-cell spread

Host-pathogen interactions that facilitate the intracellular dissemination of Listeria monocytogenes are critical for pathogenesis. L. monocytogenes success in spreading from cell to cell is determined by two key processes: The capacity for cytosolic motility and the ability to generate productive membrane protrusions. Using an RNAi-based genetic approach in combination with computer-assisted image analysis, we screened the entire human kinome for host factors limiting the cell-to-cell spread of L. monocytogenes. Our results have implicated two host serine/threonine kinases that when depleted individually, attenuate L. monocytogenes ability to spread from cell to cell.;CSNK2B-depletion disrupts L. monocytogenes ability to form actin tails and drastically restricts its cell-to-cell spreading potential. We showed that similar to host nucleation promoting factors WASP and WAVE2 CK2-mediated phosphorylation of ActA directly regulates its affinity for the Arp2/3 complex. This reveals that the ActA protein behaves not only as a structural and functional mimic but also as a regulatory mimic. Therefore, CK2-depletion impairs L. monocytogenes actin-based motility and subsequent capacity for efficient cell-to-cell dissemination.;We have additionally identified host kinase CSNK1A1 as having an alternate contribution to L. monocytogenes intracellular spread. Bacteria in CSNK1A1-depleted cells display wild-type actin-tail formation and cytosolic motility. However, upon reaching the plasma membrane, bacteria in CSNK1A1-depleted cells remained confined within membrane protrusions originating from the primary infected cell. Furthermore, the spreading defect observed upon CSNK1A1-depletion is unique to L. monocytogenes as similar experiments with Rickettsia coronii did not generate any observable phenotype. Our results collectively showed that the cell-to-cell spreading defect in CSNK1A1-depleted cells is not due to a impairment of actin-based motility like the one observed for CSNK2B-depletion, but from a failure to generate productive membrane protrusions.