Naegleria’smitotic spindles are built from unique tubulins and highlight core spindle features

Abstract

ABSTRACTNaegleria gruberiis a unicellular eukaryote whose evolutionary distance from animals and fungi has made it useful for developing hypotheses about the last common eukaryotic ancestor.Naegleriaamoebae lack a cytoplasmic microtubule cytoskeleton and assemble microtubules only during mitosis, and thus provides a unique system to study the evolution and functional specificity of mitotic tubulins and the resulting spindle. Previous studies showed thatNaegleriaamoebae express a divergent α-tubulin during mitosis and we now show thatNaegleriaamoebae express a second mitotic α- and two mitotic β-tubulins. The mitotic tubulins are evolutionarily divergent relative to typical α- and β- tubulins, contain residues that suggest distinct microtubule properties, and may represent drug targets for the “brain-eating amoeba”Naegleria fowleri. Using quantitative light microscopy, we find thatNaegleria’s mitotic spindle is a distinctive barrel-like structure built from a ring of microtubule bundles. Similar to those of other species,Naegleria’s spindle is twisted and its length increases during mitosis suggesting that these aspects of mitosis are ancestral features. Because bundle numbers change during metaphase, we hypothesize that the initial bundles represent kinetochore fibers, and secondary bundles function as bridging fibers.