Cytodifferentiation in the transition cycle of the parasitic protozoan Leishmania mexicana amazonensis was studied in vitro. The flagellated motile promastigotes transform into the nonmotile amastigotes in 7 days at 35 degrees C intracellularly in the murine macrophage line J774G8. In medium 199 plus fetal bovine serum, the reverse transformation occurs extracellularly at 27 degrees C in 2 days. Slab gel electrophoresis of leishmanias labeled with [35S]methionine during transformation revealed changes in protein banding patterns. The intensity of two protein species with apparent molecular weights of approximately equal to 55,000 increased in the amastigote-to-promastigote differentiation and decreased during the reverse transformation. These two protein species comigrated approximately with alpha- and beta-tubulin of Chlamydomonas flagella in two-dimensional gel electrophoresis. The lower band was further identified as beta-tubulin by immunoprecipitation using rabbit antiserum specific to the beta-tubulin of Chlamydomonas axonemes. The biosynthetic change of tubulin was found to correlate with the morphological change of microtubules is leishmanial flagella and cytoskeleton during transformation.