Preface

This is the fifth Methods in Cell Biology volume that focuses on microtubules and

microtubule-associated proteins. Two volumes in the 1980s that were publishedwhen

knowledge of the cytoskeleton was in its infancy covered the entire cytoskeleton, TheCytoskeleton (Volume 24 Part A and Volume 25 Part B, ed. L. Wilson). These two

volumes covered a wide range of microtubule, actin, myosin, intermediate filament,

flagella,mitotic spindle, and sea urchin topics. Our knowledge of the cytoskeleton has

expanded greatly and now cytoskeletal methods books have become focused on the

individual filaments and their associated regulatory proteins. With respect to micro-

tubules, Volume 95Microtubules, in vitro (ed. J.J. Correia and L.Wilson) appeared in

2010 and covered five broad microtubule topics: (1) isolation of microtubules and

associated proteins and the biochemistry and characterization of antibodies and tubu-

lin isotypes; (2) microtubule structure and dynamics; (3) drugs; (4) interactions with

motors and maps; and (5) functional extracts and force measurements. Volume 97

Microtubules: in vivo, edited byLynnCassimeris and PhongTran, appeared that same

year and covered a wide range of cellular and species topics including microtubule

dynamics, fission and budding yeast,Drosophila,Giardia,Dictyostelium,C. elegans,plants,melanophores, zebrafish, and cryo-electron tomography.While these volumes

provided an extensive compilation of the methods used for studying microtubules,

microtubule-associated proteins, and antimitotic drugs in numerous contexts, they

were by no means exhaustive and the field has continued progressing rapidly with

the introduction ofmany newmethodologies and approaches. This current and second

volume onMicrotubules, in vitro (ed. J.J. Correia and L.Wilson) contains 24 chapters

that continue our attempts to inform and educate microtubule researchers and new

investigators now entering the field on the rapidly expanding progress in the field.

The 24 chapters in the current volume (Microtubules, in vitro, Part 2, ed. J.J. Correiaand L.Wilson) cover a diverse range of topics. Susan Bane’s group describes biochem-

ical methods for specific fluorescent labeling of tubulin and tubulin/microtubule-

targeting drugs (Chapter 1), while Gary Brouhard’s group describes a fluorescence-

based assay for tubulin structure analysis (Chapter 21). Dan Sackett’s group compares

various modern methods for measuring tubulin polymerization in vitro (Chapter 14).

Three chapters describe methods to evaluate microtubule motion. Douglas Martin’s

group looks at gliding assays for persistence length analysis (Chapter 2), while Kris

Leslie and Niels Galjart describe how to examine microtubule dynamics by TIRF mi-

croscopy (Chapter 8). Megan Valentine’s group describes how to use magnetic twee-

zers to visualize 3D deformations in microtubule networks (Chapter 6). Liam

Cheeseman and collaborators present kinetochore ultrastructure by correlation light-

electron microscopy (Chapter 20). Jessica Lucas’ lab presents imaging methods for

plantmicrotubules (Chapter 15). Two groups describemethods for production and anal-

ysis of microtubule antibodies. Franck Perez’s lab looks at recombinant antibodies for

assaying microtubule structure, conformational states of tubulin, and dynamics

(Chapter 10 and cover art). Carsten Janke and Maria Magiera describe how to use

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antibodies to obtain quantitative information about tubulin posttranslational modifica-

tions with an emphasis on carboxy-terminal modifying enzyme effects (Chapter 16).

Jennifer Ross’s group describes the expression, purification, and biophysical character-

ization of katanin, amicrotubule-severing enzyme (Chapter 13).CarolynMoores’ group

presents biochemical and structural methods for studying the binding of doublecortin to

the microtubule lattice (Chapter 3). Holly Goodson’s group presents computational

methods for simulating various protein or drug bindingmodes to themicrotubule lattice

(Chapter 23). Francois Devred and collaborators present ITC methods for studying

tubulin–MAP interactions (Chapter 18). Jessica Field andhermany international collab-

orators present structural methods for studying the interaction of a novel covalent drug

Zampanolide (isolated from a marine sponge) to a tubulin–stathmin complex

(Chapter 19). Scott Brady describes methods for the analysis of microtubule motor-

driven transport in giant squid axoplasm (Chapter 9). Sharon Lobert’s group describes

methods for studying the complex role of micro-RNA in regulating tubulin isotype

expression and the implications for antimitotic resistance (Chapter 5). Nicholas Cowan

describes the role of a tubulin chaperone machine that assembles a/b heterodimers

(Chapter 11). Mark Rasenick’ group describes the evidence that tubulin interacts with

and regulates G proteins (Chapter 12). Fernando Cabral’s group presents methods for

studying the drug-inducedmicrotubule detachment from centrosomes and spindle poles

(Chapter 4). Phong Tran presents two chapters: one on the dynamics of mitochondria

localization tomicrotubules (Chapter 7) and a second on imaging individual spindlemi-

crotubule dynamic in fission yeast (Chapter 24). Finally, Jose Andreu’s lab describes

methods for the preparation of bacterial tubulin BtubA/B bearing eukaryotic tubulin

sequences (Chapter 17).

This rather eclectic group of methods chapters cover biochemistry, physical

chemistry, binding, antimitotic drugs, enzymology, fluorescence, structure,

microscopy, imaging, assembly, dynamics, accessory proteins, molecular motors,

chaperones, antibodies, signal transduction, centrosomes, kinetochores, spindles,

mitochondria, and bacterial tubulin. As any inquiring microtubule student or investi-

gatorwill attest, this appears to be the appropriate realitywhendescribing themethods

and fields of investigation required to know and understand the general and specific

aspects of microtubule, their regulation, and their varied functions. Attempting to en-

ter this field both broadens your base of cell biology knowledge and stimulates your

scientific imagination.We thank the authors for the hardwork thatwent into preparing

a volume like this collectively successful. (Once again some authors could not meet

the deadline andwe invite them to participate in future volumes, especially if they start

now.)We thank ShaunGamble and ZoeKruze at Elsevier for their excellent technical

support and constant reminders during this process. As usual we also thank the pa-

tience and understanding of our collaborators, colleagues, and family during our

single-minded pursuit of this task—we promise to not do it again, too soon.

John (Jack) CorreiaUMMC, Jackson, MS

Leslie WilsonUCSB, Santa Barbara, CA