Tubulin biochemistry confers intrinsic differences in microtubule dynamics and drug sensitivity between species

William Graham Hirst, Humboldt-Universität Zu Berlin
2021
Microtubules are filamentous intracellular polymers that are fundamental components of subcellular structures including the spindle, the cytoskeleton, and flagella in eukaryotes. This study uses a comparative approach to investigate how the intrinsic dynamic and biochemical characteristics of tubulin vary between species and demonstrates their consequences in two different physiological contexts: 1) Spindle size control in Xenopus frogs, and 2) The specificity of microtubule inhibitors for
more » ... inhibitors for Plasmodium falciparum microtubules over those of their human host. In Xenopus frog eggs, the length of the spindle is biochemically controlled and reaches an upper limit independent of spatial constraints. In this study, in vitro measurements of Xenopus microtubule dynamics show that X. laevis microtubules are both faster-growing and longer-lived X. tropicalis, independent of the influence of microtubule-associated proteins. Furthermore, quantification of Xenopus microtubule length and mass distributions, combined with egg extract spindle assembly reactions, establishes a role for intrinsic microtubule dynamics in modulating spindle length. Microtubules are also established drug targets in fungal and parasitic helminth infections and have in the past decades drawn attention as a potential drug target in the malaria parasite Plasmodium falciparum. In order to characterize P. falciparum microtubule dynamics, structure, and drug specificity, we have used an affinity chromatography-based approach to purify tubulin directly from blood-stage parasites. For the first time, dynamic P. falciparum microtubules have been reconstituted in vitro and parasite-specific suppression of microtubule dynamics by oryzalin and amiprofos methyl has been directly demonstrated. This study establishes an experimental framework to directly test for parasite-specific microtubule inhibition, microtubule structure, and interactions with MAPs that previously have not observed using existing in vitro approaches.
doi:10.18452/22956 fatcat:46sahejeobdnxhunongm4deqye