Genome-wide sequencing of 20 Leishmania gene mutants as part of a wider study to define protein kinases with life cycle transition roles

These 20 gene mutants were generated as part of a larger study investigating protein kinases within Leishmania mexicana. Differentiation between distinct stages is fundamental for the life cycle of intracellular protozoan parasites and for transmission between hosts, requiring stringent spatial and temporal regulation. In the larger study, we applied kinome-wide gene deletion and gene tagging in the Leishmania mexicana promastigote stage to define protein kinases with life cycle transition roles. 44 protein kinase genes were refractory to deletion and are likely core genes required for parasite replication, whilst 162 were dispensable. Phenotyping of pooled gene deletion mutants using bar-seq and projection pursuit clustering revealed functional phenotypic groups of protein kinases involved in differentiation from metacyclic promastigote to amastigote, growth and survival in macrophages and mice, colonisation of the sand fly and motility-associated flagellum function. This unbiased interrogation of protein kinases in Leishmania allows targeted investigation of organelle-associated intrinsic and extrinsic signalling pathways required for successful intracellular parasitism.