Polyglutamylation of Tubulin as a Progressive Regulator of in Vitro Interactions between the Microtubule-Associated Protein Tau and Tubulin

Abstract

The multiple functions of microtubules are mediated by various structural and motor microtubule-associated proteins (MAPs). To harmonize these functions in different places of a single cell, the key problem is to regulate the interactions of these proteins with microtubules. The chemical diversity of tubulin isoforms, which constitute the microtubule wall, could represent a molecular basis for this control. Using an in vitro assay of ligand blotting, we found that the microtubule-associated protein Tau interacts differentially with the diverse posttranslationally-modified isotubulins: its binding is mainly restricted to moderately-modified alpha- and beta-tubulin isoforms. We obtained evidence that the recently-discovered polyglutamylation, which consists of the sequential, posttranslational addition of one to six glutamyl units to both alpha- and beta-tubulin subunits, regulates the binding of Tau as a function of its chain length. The relative affinity of Tau, very low for unmodified tubulin, increases progressively for isotubulins carrying from one to three glutamyl units, reaches an optimal value, and then decreases progressively when the polygutamyl chain lengthens up to six residues. Our results suggest that the unmodified C-terminus of tubulin exerts a constitutive inhibition on Tau binding, probably by locking the MAP-binding site, and that this inhibition could be first released and then restored as the polyglutamyl chain grows. As the posttranslational chain does not appear to interact directly with Tau, it is thought that the growth of this chain from one to six glutamyl units causes a progressive, conformational shift in the structure of the C-terminal domain of tubulin, thus leading to the observed modulation of affinity.

Post-translational modifications of microtubules

“…Overexpression of the TTLL6-type E-ligase (-tubulin elongase) in Tetrahymena thermophila leads to the accumulation of hyperglutamylated nocodazole-resistant cytoplasmic microtubules . The extent to which tubulin is glutamylated might affect the binding affinity of neuronal stabilizing microtubule-associated proteins (MAPs) such as MAP2 and Tau (Bonnet et al, 2001;Boucher et al, 1994). Consistent with this, depletion of the TTLL7 E-ligase in differentiating PC-12 cells by RNA interference (RNAi) inhibits the emergence of MAP2-enriched neurites (Ikegami et al, 2006).…”

The Tubulin Code

“…75 In vitro, Tau, MAP1B, and MAP2 bind preferentially to tubulins with moderate levels of polyglutamylation (~3 glutamyl units) whereas MAP1A shows optimal affinity for highly modified tubulins (~6 glutamyl units). [76][77][78] As a-tubulin glutamylation is abundant in very young neurons whereas b-tubulin glutamylation increases during post-natal development, 50 glutamylation could control transitions in MAP binding during neuronal development. 78 Lys 40 a-tubulin acetylation may also influence MAP binding as overexpression of HDAC6 delocalized p58, a MAP involved in the association of Golgi membranes with microtubules.…”

Loss of α-tubulin polyglutamylation in ROSA22 mice is associated with abnormal targeting of KIF1A and modulated synaptic function

“…3A). Because binding affinities of structural microtubule-associated proteins (MAPs) can also be affected differently by the extent of tubulin polyglutamylation (25,26), we also investigated the ability of MAPs to be coprecipitated with microtubules. In the presence of ATP, which promotes detachment of kinesin motors from microtubules, all MAPs analyzed except MAP2 exhibited weaker binding affinity to microtubules isolated from ROSA22 mutants (Fig.…”