Volume 4 Supplement 1
Leishmania IFT140 mutants show normal viability but lack external flagella: a tool for the study of flagellar function through the infectious cycle
© Fowlkes-Comninellis and Beverley. 2015
Published: 13 July 2015
A striking feature of the parasitic protozoan Leishmania is the dramatic remodeling of the flagellum during the infectious cycle; in the sand fly midgut the promastigote form has a long, motile flagellum, while within the phagolysosome of the mammalian macrophage the flagellum barely protrudes past the surface of the amastigote form. The flagellum functions in promastigote motility and likely in cell morphogenesis, division, and maintenance of flagellar pocket structure for both forms. The flagellar pocket is the sole site of endo- and exocytosis in trypanosomatids and a sensory role for the amastigote flagellum has been proposed but a function is not established. Intraflagellar transport (IFT) is required for flagellar assembly and viability in the long, motile flagella of trypanosomes but unknown in Leishmania. We asked whether IFT is essential for viability in 'long' promastigote and 'short' amastigote flagella in L. donovani strain Bob by targeting a core retrograde pathway gene IFT140. Using a plasmid segregation knockout approach , viable knockout promastigotes were readily obtained. Δift140 lack external flagella by light and scanning electron microscopy, and have defective axonemes by transmission electron microscopy, which was reversed by complementation. This is the first example of a Leishmania mutant lacking flagella while retaining normal viability and growth. Thus, Δift140 allows for further studies of promastigote flagellar function. In addition, LdBob can generate amastigotes in vitro and infect mammalian hosts allowing us to probe the effect of Δift140 on the short flagella of amastigotes and parasite virulence.
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