Updated publication reference for PubMed record(s): 34145271. Active host cell invasion by the obligate intracellular apicomplexan parasites relies on the formation of a moving junction (MJ), which joins parasite and host cell plasma membranes during entry. Invading Toxoplasma gondii tachyzoites secrete a complex of RON proteins from the rhoptry organelles that provide the components of the MJ on the cytoplasmic side of the host cell membrane. This study highlights the role of a rhoptry resident kinase, RON13, that plays a crucial role during host cell entry and is a virulence factor in the mouse model of infection. Comparative phosphoproteomics demonstrated RON13-dependent phosphorylation of secreted rhoptry proteins and primarily the RONs complex. Structural and biochemical analysis of RON13 revealed the atypical nature of this active secretory pathway kinase, unique to the Apicomplexa.