Browse
Submit Data
Databases
API
Help

Dataset Information

14 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Tau Avoids the GTP Cap at Growing Microtubule Plus-Ends.

ABSTRACT: Plus-end tracking proteins (+TIPs) associate with the growing end of microtubules and mediate important cellular functions. The majority of +TIPs are directed to the plus-end through a family of end-binding proteins (EBs), which preferentially bind the stabilizing cap of GTP-tubulin present during microtubule growth. One outstanding question is whether there may exist other microtubule-associated proteins (MAPs) that preferentially bind specific nucleotide states of tubulin. Here, we report that the neuronal MAP tau preferentially binds GDP-tubulin (K _D = 0.26 ?M) over GMPCPP-tubulin (K _D = 1.1 ?M) in vitro, as well as GTP-tubulin at the tips of growing microtubules, causing tau binding to lag behind the plus-end both in vitro and in live cells. Thus, tau is a microtubule tip avoiding protein, establishing the framework for a possible new class of tip avoiding MAPs. We speculate that disease-relevant tau mutations may exert their phenotype by their failure to properly recognize GDP-tubulin.

SUBMITTER: Castle BT

PROVIDER: S-EPMC7691178 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Tau Avoids the GTP Cap at Growing Microtubule Plus-Ends.

Castle Brian T BT McKibben Kristen M KM Rhoades Elizabeth E Odde David J DJ

iScience 20201106 12

Plus-end tracking proteins (+TIPs) associate with the growing end of microtubules and mediate important cellular functions. The majority of +TIPs are directed to the plus-end through a family of end-binding proteins (EBs), which preferentially bind the stabilizing cap of GTP-tubulin present during microtubule growth. One outstanding question is whether there may exist other microtubule-associated proteins (MAPs) that preferentially bind specific nucleotide states of tubulin. Here, we report that ...[more]

PMID: 33294790

Similar Datasets

Structural model for differential cap maturation at growing microtubule ends.

Project description:Microtubules (MTs) are hollow cylinders made of tubulin, a GTPase responsible for essential functions during cell growth and division, and thus, key target for anti-tumor drugs. In MTs, GTP hydrolysis triggers structural changes in the lattice, which are responsible for interaction with regulatory factors. The stabilizing GTP-cap is a hallmark of MTs and the mechanism of the chemical-structural link between the GTP hydrolysis site and the MT lattice is a matter of debate. We have analyzed the structure of tubulin and MTs assembled in the presence of fluoride salts that mimic the GTP-bound and GDP•Pi transition states. Our results challenge current models because tubulin does not change axial length upon GTP hydrolysis. Moreover, analysis of the structure of MTs assembled in the presence of several nucleotide analogues and of taxol allows us to propose that previously described lattice expansion could be a post-hydrolysis stage involved in Pi release.

| S-EPMC7064335 | biostudies-literature

EBs recognize a nucleotide-dependent structural cap at growing microtubule ends.

Project description:Growing microtubule ends serve as transient binding platforms for essential proteins that regulate microtubule dynamics and their interactions with cellular substructures. End-binding proteins (EBs) autonomously recognize an extended region at growing microtubule ends with unknown structural characteristics and then recruit other factors to the dynamic end structure. Using cryo-electron microscopy, subnanometer single-particle reconstruction, and fluorescence imaging, we present a pseudoatomic model of how the calponin homology (CH) domain of the fission yeast EB Mal3 binds to the end regions of growing microtubules. The Mal3 CH domain bridges protofilaments except at the microtubule seam. By binding close to the exchangeable GTP-binding site, the CH domain is ideally positioned to sense the microtubule's nucleotide state. The same microtubule-end region is also a stabilizing structural cap protecting the microtubule from depolymerization. This insight supports a common structural link between two important biological phenomena, microtubule dynamic instability and end tracking.

| S-EPMC3368265 | biostudies-literature

Organization and dynamics of growing microtubule plus ends during early mitosis.

Project description:A stable cell line expressing EB1-green fluorescent protein was used to image growing microtubule plus ends at the G(2)/M transition. By late prophase growing ends no longer extend to the cell periphery and were not uniformly distributed around each centrosome. Growing ends were much more abundant in the area surrounding the nuclear envelope, and microtubules growing around the nucleus were 1.5 fold longer than those growing in the opposite direction. The growth of longer ends toward the nucleus did not result from a localized faster growth rate, because this rate was approximately 11 microm/min in all directions from the centrosome. Rather, microtubule ends growing toward the nucleus seemed stabilized by dynein/dynactin associated with the nuclear envelope. Injection of p50 into late prophase cells removed dynein from the nuclear envelope, reduced the density of growing ends near the nuclear envelope and resulted in a uniform distribution of growing ends from each centrosome. We suggest that the cell cycle-dependent binding of dynein/dynactin to the nuclear envelope locally stabilizes growing microtubules. Both dynein and microtubules would then be in a position to participate in nuclear envelope breakdown, as described in recent studies.

| S-EPMC151569 | biostudies-literature

Multivalent interactions facilitate motor-dependent protein accumulation at growing microtubule plus-ends.

Project description:Growing microtubule ends organize end-tracking proteins into comets of mixed composition. Here using a reconstituted fission yeast system consisting of end-binding protein Mal3, kinesin Tea2 and cargo Tip1, we found that these proteins can be driven into liquid-phase droplets both in solution and at microtubule ends under crowding conditions. In the absence of crowding agents, cryo-electron tomography revealed that motor-dependent comets consist of disordered networks where multivalent interactions may facilitate non-stoichiometric accumulation of cargo Tip1. We found that two disordered protein regions in Mal3 are required for the formation of droplets and motor-dependent accumulation of Tip1, while autonomous Mal3 comet formation requires only one of them. Using theoretical modelling, we explore possible mechanisms by which motor activity and multivalent interactions may lead to the observed enrichment of Tip1 at microtubule ends. We conclude that microtubule ends may act as platforms where multivalent interactions condense microtubule-associated proteins into large multi-protein complexes.

| S-EPMC9859754 | biostudies-literature

Actin-microtubule coordination at growing microtubule ends.

Project description:To power dynamic processes in cells, the actin and microtubule cytoskeletons organize into complex structures. Although it is known that cytoskeletal coordination is vital for cell function, the mechanisms by which cross-linking proteins coordinate actin and microtubule activities remain poorly understood. In particular, it is unknown how the distinct mechanical properties of different actin architectures modulate the outcome of actin-microtubule interactions. To address this question, we engineered the protein TipAct, which links growing microtubule ends via end-binding proteins to actin filaments. We show that growing microtubules can be captured and guided by stiff actin bundles, leading to global actin-microtubule alignment. Conversely, growing microtubule ends can transport, stretch and bundle individual actin filaments, thereby globally defining actin filament organization. Our results provide a physical basis to understand actin-microtubule cross-talk, and reveal that a simple cross-linker can enable a mechanical feedback between actin and microtubule organization that is relevant to diverse biological contexts.

| S-EPMC4365169 | biostudies-literature

Estimating the microtubule GTP cap size in vivo.

Project description:Microtubules (MTs) polymerize via net addition of GTP-tubulin subunits to the MT plus end, which subsequently hydrolyze to GDP-tubulin in the MT lattice. Relatively stable GTP-tubulin subunits create a "GTP cap" at the growing MT plus end that suppresses catastrophe. To understand MT assembly regulation, we need to understand GTP hydrolysis reaction kinetics and the GTP cap size. In vitro, the GTP cap has been estimated to be as small as one layer (13 subunits) or as large as 100-200 subunits. GTP cap size estimates in vivo have not yet been reported. Using EB1-EGFP as a marker for GTP-tubulin in epithelial cells, we find on average (1) 270 EB1 dimers bound to growing MT plus ends, and (2) a GTP cap size of ?750 tubulin subunits. Thus, in vivo, the GTP cap is far larger than previous estimates in vitro, and ?60-fold larger than a single layer cap. We also find that the tail of a large GTP cap promotes MT rescue and suppresses shortening. We speculate that a large GTP cap provides a locally concentrated scaffold for tip-tracking proteins and confers persistence to assembly in the face of physical barriers such as the cell cortex.

| S-EPMC3461128 | biostudies-literature

Rapid binding to protofilament edge sites facilitates tip tracking of EB1 at growing microtubule plus-ends.

Project description:EB1 is a key cellular protein that delivers regulatory molecules throughout the cell via the tip-tracking of growing microtubule plus-ends. Thus, it is important to understand the mechanism for how EB1 efficiently tracks growing microtubule plus-ends. It is widely accepted that EB1 binds with higher affinity to GTP-tubulin subunits at the growing microtubule tip, relative to GDP-tubulin along the microtubule length. However, it is unclear whether this difference in affinity alone is sufficient to explain the tip-tracking of EB1 at growing microtubule tips. Previously, we found that EB1 binds to exposed microtubule protofilament-edge sites at a ~70 fold faster rate than to closed-lattice sites, due to diffusional steric hindrance to binding. Thus, we asked whether rapid protofilament-edge binding could contribute to efficient EB1 tip tracking. A computational simulation with differential EB1 on-rates based on closed-lattice or protofilament-edge binding, and with EB1 off-rates that were dependent on the tubulin hydrolysis state, robustly recapitulated experimental EB1 tip tracking. To test this model, we used cell-free biophysical assays, as well as live-cell imaging, in combination with a Designed Ankyrin Repeat Protein (DARPin) that binds exclusively to protofilament-edge sites, and whose binding site partially overlaps with the EB1 binding site. We found that DARPin blocked EB1 protofilament-edge binding, which led to a decrease in EB1 tip tracking on dynamic microtubules. We conclude that rapid EB1 binding to microtubule protofilament-edge sites contributes to robust EB1 tip tracking at the growing microtubule plus-end.

| S-EPMC10883673 | biostudies-literature

Tubulin tyrosination is a major factor affecting the recruitment of CAP-Gly proteins at microtubule plus ends.

Project description:Tubulin-tyrosine ligase (TTL), the enzyme that catalyzes the addition of a C-terminal tyrosine residue to alpha-tubulin in the tubulin tyrosination cycle, is involved in tumor progression and has a vital role in neuronal organization. We show that in mammalian fibroblasts, cytoplasmic linker protein (CLIP) 170 and other microtubule plus-end tracking proteins comprising a cytoskeleton-associated protein glycine-rich (CAP-Gly) microtubule binding domain such as CLIP-115 and p150 Glued, localize to the ends of tyrosinated microtubules but not to the ends of detyrosinated microtubules. In vitro, the head domains of CLIP-170 and of p150 Glued bind more efficiently to tyrosinated microtubules than to detyrosinated polymers. In TTL-null fibroblasts, tubulin detyrosination and CAP-Gly protein mislocalization correlate with defects in both spindle positioning during mitosis and cell morphology during interphase. These results indicate that tubulin tyrosination regulates microtubule interactions with CAP-Gly microtubule plus-end tracking proteins and provide explanations for the involvement of TTL in tumor progression and in neuronal organization.

| S-EPMC2064338 | biostudies-literature

The speed of GTP hydrolysis determines GTP cap size and controls microtubule stability.

Project description:Microtubules are cytoskeletal polymers whose function depends on their property to switch between states of growth and shrinkage. Growing microtubules are thought to be stabilized by a GTP cap at their ends. The nature of this cap, however, is still poorly understood. End Binding proteins (EBs) recruit a diverse range of regulators of microtubule function to growing microtubule ends. Whether the EB binding region is identical to the GTP cap is unclear. Using mutated human tubulin with blocked GTP hydrolysis, we demonstrate that EBs bind with high affinity to the GTP conformation of microtubules. Slowing-down GTP hydrolysis leads to extended GTP caps. We find that cap length determines microtubule stability and that the microtubule conformation changes gradually in the cap as GTP is hydrolyzed. These results demonstrate the critical importance of the kinetics of GTP hydrolysis for microtubule stability and establish that the GTP cap coincides with the EB-binding region.

| S-EPMC7018511 | biostudies-literature

Dynein tethers and stabilizes dynamic microtubule plus ends.

Project description:Microtubules undergo alternating periods of growth and shortening, known as dynamic instability. These dynamics allow microtubule plus ends to explore cellular space. The "search and capture" model posits that selective anchoring of microtubule plus ends at the cell cortex may contribute to cell polarization, spindle orientation, or targeted trafficking to specific cellular domains. Whereas cytoplasmic dynein is primarily known as a minus-end-directed microtubule motor for organelle transport, cortically localized dynein has been shown to capture and tether microtubules at the cell periphery in both dividing and interphase cells. To explore the mechanism involved, we developed a minimal in vitro system, with dynein-bound beads positioned near microtubule plus ends using an optical trap. Dynein induced a significant reduction in the lateral diffusion of microtubule ends, distinct from the effects of other microtubule-associated proteins such as kinesin-1 and EB1. In assays with dynamic microtubules, dynein delayed barrier-induced catastrophe of microtubules. This effect was ATP dependent, indicating that dynein motor activity was required. Computational modeling suggests that dynein delays catastrophe by exerting tension on individual protofilaments, leading to microtubule stabilization. Thus, dynein-mediated capture and tethering of microtubules at the cortex can lead to enhanced stability of dynamic plus ends.

| S-EPMC3347920 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data