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Review
, 21 (10), 604-14

Regulation of Microtubule Dynamics by TOG-domain Proteins XMAP215/Dis1 and CLASP

Affiliations
Review

Regulation of Microtubule Dynamics by TOG-domain Proteins XMAP215/Dis1 and CLASP

Jawdat Al-Bassam et al. Trends Cell Biol.

Abstract

The molecular mechanisms by which microtubule-associated proteins (MAPs) regulate the dynamic properties of microtubules (MTs) are still poorly understood. We review recent advances in our understanding of two conserved families of MAPs, the XMAP215/Dis1 and CLASP family of proteins. In vivo and in vitro studies show that XMAP215 proteins act as microtubule polymerases at MT plus ends to accelerate MT assembly, and CLASP proteins promote MT rescue and suppress MT catastrophe events. These are structurally related proteins that use conserved TOG domains to recruit tubulin dimers to MTs. We discuss models for how these proteins might use these individual tubulin dimers to regulate dynamic behavior of MT plus ends.

Figures

Figure 1
Figure 1. Assembly and disassembly of dynamic microtubules (MTs)
A) Conformational change of αβ-tubulin accompanying GTP hydrolysis. In the GTP state (in β-tubulin in green), α and β tubulin monomer interfaces result in “straight” tubulin dimer. In the GDP state αβ-tubulin dimer interface is curved by 5 degrees (arrow), leading to a “bent” tubulin dimer. B) Structural changes at MT plus ends. During MT assembly, MT plus ends form a sheet-like group of straight protofilaments. GTP-tubulin dimers (green) assemble on the ends, forming a cap of GTP tubulin. GTP hydrolysis over time converts GTP-tubulin in the lattice to GDP-tubulin (note that the extent of the GTP cap is not known). In the MT disassembly phase, GDP-tubulin protofilaments curl and peel off the MT plus ends. The transitions between growth and shrinkage states are termed catastrophe and rescue.
Figure 2
Figure 2. Localization and activities of XMAP215/Dis and CLASP proteins
A) Drosophila Msps (XMAP215/Dis1) on spindle MTs and poles. B) Drosophila Msps on interphase MT plus ends (Msps, red, arrowhead; tubulin, green) C) Human CLASP1 on anaphase spindle, at spindle midzone MTs and poles (arrows)(CLASP1, green; MT, red; DNA blue). D) Human CLASP1 at kinetochore on a metaphase chromosome (CLASP green; DNA, blue; ACA centromere marker, red). E) Human CLASP2 staining at interphase MT plus ends near plasma membrane (CLASP, red; MT, green). F) Fission yeast CLASP Cls1p in clusters on interphase MT bundles near nuclear envelope (Cls1p, green; MT, red). G) Human CLASP1 on the lattice of interphase MTs near the leading edge of the cell (arrowhead). H) Schematic showing dynamic behavior of pure MTs (grey lines) I) XMAP215 (green) at the MT plus end accelerates MT assembly and leads to formation of long MTs. J) S. pombe CLASP, Cls1p (red) binds to the MT lattice and promotes local MT rescue, preventing MTs from shrinking completely. Images in A–G are reproduced with permission from [40] [24, 48, 53, 61, 63].
Figure 3
Figure 3. Domain organization of XMAP215/Dis1 and CLASP families from yeast, worms and mammals and their binding partners
A) XMAP215/Dis1 proteins contain conserved TOG domains and SK rich domain. Domain organization of yeast orthologs S.cereviae Stu2, S.pombe Dis1 and Alp-14 with two TOG domains, C. elegans Zyg9 with three TOG domains and D. melanogaster MSPS, X. laevis XMAP215 and human ch-TOG with five TOG domains. All molecules contain regions with stretches of sequences rich in Serine, Glycine, Lysine (SK-rich domains). TOG domains are colored based on the conserved phylogenetic classes from sequence alignments (Figure 4): TOG1 class (Blue), TOG2 class (Cyan), TOG4 class (sky blue), TOG4 (purple), TOG5 class (maroon). Protein binding partners (Blue) described in the text, are shown below each protein, with arrows denoting approximate binding sites. An absence of an arrow denotes an interaction in which binding domains have not yet been mapped. B) CLASP proteins contain conserved TOG-Like (TOGL) domains and SR-rich domains Similar to A, domain organization of S. cerevisae Stu1 and S. pombe Cls1 with two TOGL domains, C. elegans Cls2 with two TOGL domains and D.melanogaster MAST/orbit, human and Xenpus CLASP1 with three TOGL domains. All molecules contain regions with stretches of sequences rich in Serine, Proline and Arginine (SR-rich domains). TOGL domains are also colored based on the conserved phylogenetic classes from sequence alignments (Figure 4): TOGL1 class (orange), TOGL2 class (red), TOGL3 class (purple). Protein binding partners (Blue), described in the text, are shown below each protein with arrows denoting approximate binding sites based on studies described in the text. An absence of an arrow denotes an interaction in which the interacting domain has not yet been mapped.
Figure 4
Figure 4. XMAP215/Dis1 and CLASP proteins bind to a tubulin dimer to form a globular complex
A) XMAP215/Dis1 molecules wrap around soluble tubulin dimers with their TOG domains to form globular complexes. Models for tubulin binding are shown on the left, and electron microscopy images are shown on the right (Stu2 or XMAP215 alone, above; tubulin complex, below). Yeast Stu2 is a homodimer with two sets of TOG domains that wrap around a single tubulin dimer, while Xenopus XMAP215 is a monomer with two internal “halves” each consisting of two TOG domains that interact with a single tubulin dimer. EM images of XMAP215 and Stu2 are reproduced with permission from [12, 14]. White Arrows denote the open conformation of XMAP215 and Stu2 molecules B) S.pombe CLASP, Cls1 is a homodimer that wrap around soluble tubulin dimers with two sets of TOGL domains. Model for Cls1 dimer binding to the tubulin dimer is shown on the left; electron microscopy images are shown on the right (Cls1 alone, above; cls1-tubulin complex, below). EM of Cls1 were reproduced with permission from [15] White Arrows denote the open conformation of XMAP215 and Stu2 molecules
Figure 5
Figure 5. XMAP215/Dis1 TOG and CLASP TOGL domains and the tubulin dimer-binding interface
A) Phylogenetic tree based on sequence alignment of TOG and TOGL domains: Separate XMAP215/Dis1 family TOG domains and CLASP family TOGL domains from S. cereviase, S. Kluyveri, S .pombe, C. elegans, D. melangaster, A. thaliana, X. laevis, mouse, and human orthologs were aligned using MUSCLE alignment server (further described in Figure S1,[78]). Distance matrices for aligned sequences were used to calculate a phylogenetic tree to classify TOG/TOGL sequences. The tree shows that aligned sequences of TOG and TOGL domains (shown in figure S1) are grouped in conserved phylogenetic classes based on TOG domain position in the protein. This analysis indicates that XMAP215/Dis1 TOG and CLASP family TOGL domain sequences have a common origin. The degree of separation between the different classes describes further divergence in the sequences. The colors of the different branches of the phylogenetic tree are consistent with the coloring scheme of TOG/TOGL domains in Figure 3. The classes are highlighted as follows: XMAP215/Dis1 TOG1 (aqua blue), XMAP215/Dis1 TOG2 (cyan), XMAP215 TOG3 (light blue), XMAP215 TOG4 (purple), XMAP215 TOG5 (brown), Cls1 TOGL1 and CLASP TOGL1 (beige), CLASP/Cls1 TOGL2 (orange), CLASP TOGL3 (purple). B) Structures of TOG domains and mutational analyses reveal the site for tubulin dimer binding. Top panel: structures of three TOG domains from Zyg9, MSPS, and Stu2 TOG domains show that TOG domains have a flat paddle shape with six conserved HEAT repeats [13, 16]). The overlaid structures are shown narrow side of the paddle overlooking the tubulin binding loops, and the wide side of the paddle overlooking the HEAT repeat helices. Lower panel: sequence conservation and mutational analyses show that TOG domains bind tubulin dimers using intra-HEAT repeat Loops T1–T5 (shown in red) in similar views as described in the upper panel. C) Detailed sequences of the tubulin binding loops in XMAP215/Dis1 TOG domain and CLASP TOGL domain classes. These aligned tubulin-binding loops describe the amino acid variations in the tubulin-binding sites of TOG and TOGL domains. All the tubulin-binding loop (T1–T5) sequences contain strictly conserved residues (purple), moderately conserved residues (blue), and weakly conserved residues (either in cyan or not colored). Although strictly conserved residues are maintained throughout TOG domains, there are class-conserved residues that differ between classes. For example, XMAP215/Dis1 TOG1 and TOG2 classes differ in many residues neighboring strictly conserved residues within T1 and T2; those variations are maintained in each class. Some classes such as XMAP215 TOG3, XMAP215 TOG4, CLASP TOGL1 and CLASP TOGL3 contain divergent variations in strictly conserved residues, suggesting they may have a weaker affinity for binding tubulin dimer.
Figure 6
Figure 6. Models for XMAP215/Dis1 and CLASP mechanisms
A) XMAP215/Dis1 family proteins are MT polymerases. Upper panel: XMAP215 binds tubulin dimers with TOG domains and the MT lattice with its SK domain (left). XMAP215-tubulin complexes bind and diffuse along MT lattices. In the absence of XMAP215-tubulin MT assembly is slow. At MT plus ends, XMAP215-tubulin complexes accumulate at the MT plus end accelerates MT assembly (right). Lower panel shows two models for how XMAP215-tubulin complexes increase MT assembly rate: Model 1, XMAP215 TOG domains cycles to load its bound tubulin dimer at growing MT ends. It promotes multiple cycles of tubulin dimer binding, accompanied by conformational change to release tubulin at MT plus ends. Model 2, XMAP215 stabilizes the assembly conformation of a microtubule by binding and stabilizing polymerized-tubulin conformation (yellow) with its TOG domains. The bound tubulin may be a soluble tubulin dimer, or a dimer located at specific site on the MT, such as the very MT end or seam. The conformation of the tubulin dimer or its nucleotide state while bound to XMAP215 is not known and is shown as GDP. B) CLASP family proteins promote MT rescues and inhibit MT catastrophes. Upper panel: CLASP binds tubulin dimer with its TOGL domains and binds MT lattices with high affinity with its SR-rich domain (left). CLASP high affinity leads to sites of high concentration along MTs. When a dynamic MT undergoes catastrophe, MT disassembly occurs until the plus end reaches such a site of high local CLASP concentration (middle). There, CLASP locally promotes rescue events, in which MT depolymerization halts and MT assembly reinitiates (right). Lower panel shows two models of how CLASP molecules induce MT rescue: Model 1, CLASP molecules releases their loaded tubulin dimer into the MT plus end, to reinitiate polymerization. In this model, CLASP molecules act as local polymerase, similar to model 1 of XMAP215. Model 2, CLASP molecules utilize their loaded tubulin to prevent MT disassembly and restore MT to the assembly phase. During this activity, CLASP molecules may or may not release their bound tubulin. CLASP-tubulin complexes may halt depolymerization, while MT assembly may be reinitiated by tubulin dimers polymerizing from solution.

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