Methods in Tubulin Proteomics
Leah M. Miller,Hui Xiao,Berta Burd,Susan Band Horwitz,Ruth Hogue Angeletti,Pascal Verdier-Pinard +5 more
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TLDR
In this article, the authors present protocols that are used for the analysis of tubulin isotypes and PTMs present in tubulin isolated from cells in culture or tissues and for identifying tubulin regions altered by microtubule-stabilizing agents.Abstract:
New analytical methods are needed for the successful outcome of experiments aimed at characterizing mechanisms of microtubule dynamics and at understanding the effects of drugs on microtubules. The identification of tubulin isotypes and of regions of the microtubule involved in drug interactions has been advanced by proteomic methodologies. The diversity of tubulin sequences and posttranslational modifications (PTMs) can generate a complex mixture of heterodimers with unique molecular dynamics driving specific functions. Mass spectrometry (MS)-based approaches have been developed, and in combination with chromatographic and/or electrophoretic separation of tubulin polypeptides or peptides, they have contributed to our understanding of tubulin proteomics. We present protocols that we have used for the analysis of tubulin isotypes and PTMs present in tubulin isolated from cells in culture or tissues and for the identification of tubulin regions altered by microtubule-stabilizing agents. Tubulin proteomics complements structural and computer modeling information for a high-resolution view of microtubule dynamics and its alteration by drugs. These methodologies will help in providing insights into tubulin isotype-specific functions and in the design of drugs targeting either all tubulin heterodimers indiscriminately or only those containing specific isotypes.read more
Citations
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Journal ArticleDOI
Deciphering post-translational modification codes.
TL;DR: This manuscript highlights several examples of combinatorial PTMs in proteins, and describes recent technological developments, which are driving the ability to understand how PTM patterns may “code” for biological outcomes.
Journal ArticleDOI
The chemical complexity of cellular microtubules: tubulin post-translational modification enzymes and their roles in tuning microtubule functions
TL;DR: This review focuses on the astonishing chemical complexity introduced in the tubulin heterodimer at the post‐translational level and summarizes the recent advances in identifying the enzymes responsible for these modifications and deciphering the consequences of tubulin's chemical diversity on the function of molecular motors and microtubule associated proteins.
Journal ArticleDOI
Succination is Increased on Select Proteins in the Brainstem of the NADH dehydrogenase (ubiquinone) Fe-S protein 4 (Ndufs4) Knockout Mouse, a Model of Leigh Syndrome *
Gerardo G. Piroli,Allison M. Manuel,Anna C. Clapper,Michael D. Walla,John E. Baatz,Richard D. Palmiter,Albert Quintana,Albert Quintana,Norma Frizzell +8 more
TL;DR: It is demonstrated for the first time that protein succination is increased in the brainstem (BS) of the Ndufs4 knockout mouse, a model of Leigh syndrome, and fumarate is a novel biochemical link that may contribute to the progression of the neuropathology in this mitochondrial disease model.
Journal ArticleDOI
Mechano-induced cell metabolism promotes microtubule glutamylation to force metastasis.
Stéphanie Torrino,Eloise M. Grasset,Stéphane Audebert,Ilyes Belhadj,Caroline Lacoux,Meagan Haynes,Sabrina Pisano,Sophie Abelanet,Frédéric Brau,Stephen Y. Chan,Bernard Mari,William M. Oldham,Andrew J. Ewald,Thomas Bertero +13 more
TL;DR: The authors showed that matrix stiffening rewires glutamine metabolism to promote microtubule glutamylation and force MT stabilization, thereby promoting cell invasion and decreased microtubules' mechanical stability.
Journal ArticleDOI
Identification of protein succination as a novel modification of tubulin.
Gerardo G. Piroli,Allison M. Manuel,Michael D. Walla,Matthew J. Jepson,Jonathan W. C. Brock,Mathur Rajesh,Ross M. Tanis,William E. Cotham,Norma Frizzell +8 more
TL;DR: The results demonstrate that succination is a novel post-translational modification of tubulin and suggest that extensive modification by fumarate, either physiologically or pharmacologically, may alter microtubule dynamics.
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