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" Characterization of Tau and Anticancer Drug Interactions with Microtubules via Cryo-Electron Microscopy Studies "
Hejab, Nisreen M A
Downing, Kenneth HLiu, Fenyong
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Latin Dissertation
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English
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898035
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TL7zb0x85p
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Bruner, Justin Pearce
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Characterization of Tau and Anticancer Drug Interactions with Microtubules via Cryo-Electron Microscopy Studies\ Hejab, Nisreen M ADowning, Kenneth HLiu, Fenyong
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2016
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2016
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| Abstract
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Microtubules (MTs) are essential components of the eukaryotic cytoskeleton. They form by the polymerization of tubulin into cylindrical polymers composed of protofilaments. MTs are involved in a diverse array of cellular functions due to their dynamic instability, which is modulated by many factors in vivo and in vitro. Agents that modulate MT dynamics include MT associated proteins (MAPs) and naturally occurring compounds known as MT stabilizing agents (MSAs). A number of MSAs have demonstrated or predicted potential as anticancer agents, but a detailed structural basis for their mechanism of action is still lacking. We have used cryo-electron microscopy (cryo-EM) to study the structural basis of action of zampanolide, a taxane-binding site (TBS) MSA, as well as several proteins known to stabilize MTs, with main focus on tau. We have obtained a high-resolution (4.2 Å) cryo-EM reconstruction of microtubules stabilized by zampanolide and have compared it to Taxol, which binds to the same pocket on β-tubulin. We find that each TBS MSA has distinct structural effects on the microtubule (MT) lattice. Binding of Taxol or zampanolide both induce MT heterogeneity, but each affects the longitudinal interface differently. Tau is the prominent neuronal MAP and it has been implicated with Alzheimer disease (AD). Tau is known to bind and stabilize MTs in the axons but its binding mode and stabilizing mechanism are not well understood. Our study shows that tau binds exclusively to the external surface of MTs along protofilaments and that it remains in an extended confirmation. In addition, in the presence of kinesin, full length tau (FL-tau) dissociates from MTs allowing kinesin binding. Surprisingly, sub-stoichiometric amounts of tau result in the formation of GDP-tubulin double rings that disappear upon increasing the amounts of tau added to MTs. FL-tau and minimal 2-R or 4-R tau constructs result in the formation of sleeve-like structures around MTs, suggesting that tau binds to tubulin dimers as well as to fully polymerized MTs. Thus, we hypothesize that tau functions by oligomerizing tubulin dimers leading to a compacted MT lattice that could reinforce the interactions across the longitudinal MT interface. Furthermore, we identified amino acids on the surface of tubulin that could interact with tau. Tau binds to MTs near the C-terminus, and the sequestering of the acidic C-terminus of tubulin could contribute to the stability of MTs in the presence of tau.
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Hejab, Nisreen M A
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UC Berkeley
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