Single-Cell Biophysics: Measurement, Modulation, and Modeling

Single-Cell Biophysics: Measurement, Modulation, and Modeling

Poster Abstracts

43-POS

Board 22

Post-Translational Modifications Regulate Function and Self-Assembly of Axonemal

Tubulin

Yin-wei Kuo

, Ron Orbach

, Jonathon Howard

Yale University, New Haven, CT, USA.

Yale University, New Haven, CT, USA,

Microtubules, cytoskeletal polymers assembled from building blocks comprising α- and β-

tubulin heterodimers, are involved in many important intracellular functions including the

maintenance of cell structure, the formation of the mitotic spindle, the transport of organelles, the

sensation of force and the motility of cells. The diverse functions of microtubules require

delicate control over the dynamics of assembly and disassembly, and the interactions with

microtubule-associated proteins (MAPs). A long standing question is how do the evolutionally

highly conserved tubulin building blocks differentially regulate microtubule dynamics and

functions in different cellular processes.

Highly versatile post-translational modifications (PTMs) are present on tubulin, contributing to

the so-called “tubulin code” and are suggested to be crucial for regulating the diverse functions

of cellular microtubules. However, our current knowledge about microtubule dynamics is

predominantly based on the highly heterogeneous mammalian brain tubulin system, whose

complex isotypes and PTM combinations make it challenging to elucidate the effect and function

of the individual modifications.

To reduce the complication of tubulin isotype, we selected biflagellated green

algae

Chlamydomonas reinhardtii

as our model system. We recently developed a novel method

to purify axonemal tubulin from various strains of

C. reinhardtii

, including a polyglutamylation-

deficient mutant

tpg1

, to study the self-assembly dynamics of axonemal tubulin and how tubulin

post-translational modifications (PTMs) contribute to the distinct properties of axonemal tubulin.

We will do this using total-internal-reflection-fluorescence (TIRF) and other

microscope

techniques to determine the effects of PTMs on dynamics at the single-microtubule level.