Microbial signal transduction pathways regulate adaptation to various environmental conditions and enable pathogens to cause infections. Most of these pathways are regulated by protein kinases. The opportunistic fungal pathogen Candida albicans exists as a commensal of the intestinal mycobiota and can cause disseminated candidiasis upon certain circumstances. Both commensalism and pathogenicity require a complex network of signalling pathways. The C. albicans genome was predicted to encode 108 protein kinases yet nearly 50% remain uncharacterised. We aim to dissect the role of C. albicans protein kinases during commensalism and infection. To investigate the pathogenicity of C. albicans, an in vitro intestinal epithelial cell (IEC) infection model was used. A library containing individual mutants lacking each of the 108 protein kinase genes identified in C. albicans was screened for their ability to damage the host tissue by measuring the activity of IEC cytoplasmatic lactate dehydrogenase in the supernatant. Mutants exhibiting an increased or decreased cytotoxicity were validated and their morphology on IECs was assessed.Our kinase mutant screen revealed that deletion of around 30% of all kinases resulted in altered cytotoxicity compared to the wild-type (18 increased and 19 decreased damage). As filamentous growth is a major virulence factor of C. albicans, the morphology of these mutants on IECs was investigated. We identified five mutants (lacking CRK1, HOG1, PBS2, SOK1, or SSK2) which showed increased IEC damage but reduced or wild-type-like filamentation. So far, members of the HOG pathway and CRK1 were described to be indispensable during commensalism and/or systemic infection but seem to negatively impact pathogenicity during epithelial infection. To fully determine the role of these protein kinases during epithelial infection, specific virulence attributes such as adhesion, invasion and translocation as well as metabolic adaptation are assessed for the corresponding mutants.Collectively, we identified a crucial role for several protein kinases during infection of IECs including kinases that negatively and filamentation-independently impact pathogenicity.