Synergy between XMAP215 and EB1 increases microtubule growth rates to physiological levels

Self Cite

Background:The reported inhibition of microtubule growth by Stu2 is difficult to reconcile with its cellular phenotypes. Results: Using microscopy assays, we found that Stu2 increases the growth rate and decreases the catastrophe frequency of yeast microtubules. Conclusion: Stu2 promotes microtubule growth, with considerably higher activity on budding yeast microtubules. Significance: The biochemical properties of Stu2 reported here account for the mitotic phenotypes observed in cells.

Section: Stu2 Is a Strong Polymerase On Budding Yeastsupporting

confidence: 78%

Section: Stu2 Is a Weak Polymerase On Porcine Brain Tubulin-tomentioning

confidence: 99%

Stu2, the Budding Yeast XMAP215/Dis1 Homolog, Promotes Assembly of Yeast Microtubules by Increasing Growth Rate and Decreasing Catastrophe Frequency

Podolski

Mahamdeh

Howard

2014

Self Cite

“…EB1 fenestrates the MT lattice, binding between four tubulin heterodimers where it recognizes tubulin in a GTP hydrolysis transition/post-hydrolysis state mimicked by the GTP analog GTP␥S (58 -60). Thus, EB1 binds just distal to the polymerizing MT GTP-cap and via SLAIN2/Sentin localizes the XMAP215 C terminus to this zone (33,34,61,62). In contrast, the XMAP215 N-terminal TOG domain array binds and incorporates ␣␤-tubulin into the growing MT plus-end (28 -31, 38, 40, 41).…”

Section: Discussionmentioning

confidence: 99%

Drosophila melanogaster Mini Spindles TOG3 Utilizes Unique Structural Elements to Promote Domain Stability and Maintain a TOG1- and TOG2-like Tubulin-binding Surface

Howard

Fox

Slep

2015

Background: XMAP215 family members contain arrayed TOG domains that promote microtubule polymerization. Results: TOG3 has a unique architecture that positions conserved residues in a TOG1-and TOG2-like tubulin-binding mode. Conclusion: TOG1, TOG2, and TOG3 have similar architectures and predicted tubulin-binding modes that contrast with TOG4. Significance: XMAP215 family members use a structurally diverse, polarized TOG domain array to promote microtubule polymerization.

“…The two proteins do not interact before binding to the MT end nor through canonical EB1 protein interactions, and they do not affect each other's localization, suggesting that the synergistic effect is allosteric. EB1 could be substituted by the lattice-straightening drug taxol without lowering the synergistic growth rates with XMAP215, providing further evidence that EB1 may affect the structure of the MT lattice (59). Besides direct recruitment via EBs and via ϩTIPs that also bind to EBs, allosteric interactions through the MT lattice are yet another mode of how MAPs can affect each other's activity.…”

Section: Adding Complexity Via ؉ Tipsmentioning

confidence: 97%

“…This effect was investigated in vitro, where EB1 and XMAP215 combined had a much larger effect on MT growth rates than XMAP215 (10-fold increase) or EB1 (1.5-fold increase) alone with rates up to 20 m/min, the highest rates ever observed outside cells (Fig. 2F) (59). This must be close to the maximum possible polymerization rate because the association rate constant for tubulin addition approached the maximum diffusion limited rate with up to 7.6 M Ϫ1 s Ϫ1 per protofilament (59).…”

Section: Adding Complexity Via ؉ Tipsmentioning

confidence: 99%

Building the Microtubule Cytoskeleton Piece by Piece

Alfaro-Aco

Petry

2015

The microtubule (MT) cytoskeleton gives cells their shape, organizes the cellular interior, and segregates chromosomes. These functions rely on the precise arrangement of MTs, which is achieved by the coordinated action of MT-associated proteins (MAPs). We highlight the first and most important examples of how different MAP activities are combined in vitro to create an ensemble function that exceeds the simple addition of their individual activities, and how the Xenopus laevis egg extract system has been utilized as a powerful intermediate between cellular and purified systems to uncover the design principles of selforganized MT networks in the cell. The microtubule (MT)2 cytoskeleton forms the skeletal framework that gives eukaryotic cells their shape and organizes their cytoplasm by positioning organelles, providing tracks for transport, and establishing cell polarity. In an interphase cell, the MT cytoskeleton is also critical for cell motility and a key constituent of cilia and flagella. During cell division, the MT cytoskeleton gets remodeled into a spindle structure that segregates chromosomes. Each of these functions relies on a specific MT architecture, which must be capable of rapid and prolonged change followed by an eventual resumption of a steady state to respond to the cellular environment and morphology changes during growth and differentiation.MTs are made of ␣/␤-tubulin heterodimers, which assemble into a polar, cylindrical structure in the presence of GTP and above the so-called critical concentration in vitro. MT growth phases alternate with swift shrinkage phases (dynamic instability), and their transitions are referred to as catastrophe (switching from growth to shrinkage) and rescue (switching from shrinkage to growth) (1). In cells, a plethora of different MT-associated proteins (MAPs) regulate the MT-inherent abilities of MT nucleation and dynamics ( Fig. 1A) (2). In addition, MT cross-linking proteins connect MTs into networks and molecular motors use MTs as tracks for cargo transport or transport MTs themselves (Fig. 1A). Altogether, different combinations of these four basic groups of MAP activities drive the self-organization of the MT cytoskeleton into discrete three-dimensional patterns (Fig. 1B) (3). Thus, they establish, maintain, and disassemble functional MT structures that are observed on the cellular level.Traditionally, individual MAPs were identified by loss-offunction experiments in cells followed by their detailed in vivo and in vitro characterization. During the past decade, highthroughput genomic and proteomic screens accelerated MAP discovery by cataloging RNAi phenotypes and identifying novel microtubule binders, resulting in comprehensive lists of candidates involved in organizing the MT cytoskeleton in various cell states (4 -7). Now, the challenge is to understand how these MAPs work together to establish the physiological MT architecture of the cell. What specific MAP building blocks can generate the MT networks that shape a dendrite or a polarized epithelial cell (...