Studies in yeasts have implicated the importance of Kinl protein kinase, a member of the eukaryotic PARI/MARK/MELK family, in polarized growth, cell division and septation through coordinated activity with the phosphatase, calcineurin. Kinl is also required for virulence of the fungal pathogens Cryptococcus neoformans and Fusarium graminearum. Here we show that kin] deletion in the human fungal pathogen Aspergillus fumigatus does not affect hyphal growth and septation but results in differential susceptibility to antifungals targeting the cell wall and cell membrane. The Akin/ strain remained virulent in a Galleria mellonella model of invasive aspergillosis. Expression of Kinl tagged to GFP or RFP showed its stable localization at the septum. Co-localization experiments revealed calcineurin (CnaA) localization on either side of Kinl at the septum suggesting possible interaction. Bimolecular fluorescence complementation assay confirmed the interaction of Kinl with CnaA at the hyphal tips and septa in the presence of the antifungal caspofungin. Furthermore, phosphoproteomic analyses for the first time revealed Kinl as a substrate of calcineurin providing novel insight into Kinl regulation through calcineurin-mediated dephosphorylation mechanism. (C) 2018 Elsevier Inc. All rights reserved.