Discussion

Kinesins are ATP-dependent microtubule binding molecular motor proteins essential for many key cellular processes in eukaryotic cells. They are involved in the movement of cell organelles, chromosomes, protein complexes, and mRNA to specific destinations within the cell. Kinesins can be modulated by mitogen-activated protein (MAP) kinase signal transduction cascades. Here, we focus on the kinesin homologue, LmxKin29 from Leishmania mexicana. It has been found to be phosphorylated at serine 548, serine 551 and serine 554 using whole cell lysates in phosphoproteomics analyses. A peptide carrying these phosphorylation sites is phosphorylated by activated recombinant LmxMPK3, a Leishmania MAP kinase involved in flagellar length control. A null mutant for LmxMPK3 showed flagella reduced in length to approximately 1/5 of the length of a wild type flagellum. We cloned and expressed LmxKin29 fused to glutathione-S-transferase and a number of putative phosphorylation site mutants. This allowed us to prove that LmxMPK3 can phosphorylate full length LmxKin29 exclusively on serine 554. To understand the function of LmxKin29 in L.mexicana we generated homozygous null mutants. Moreover, we generated cell lines expressing N- or C-terminally GFP-tagged LmxKin29 in the wild type and knock out background. Microscopic analyses indicate that LmxKin29 predominantly localises close to the flagellar pocket. Further phenotypic analyses are under way.