Dataset Information

Stu2 acts as a microtubule destabilizer in metaphase budding yeast spindles.

ABSTRACT: The microtubule-associated protein Stu2 (XMAP215) has the remarkable ability to act either as a polymerase or as a destabilizer of the microtubule plus end. In budding yeast, it is required for the dynamicity of spindle microtubules and also for kinetochore force generation. To understand how Stu2 contributes to these distinct activities, we analyzed the contributions of its functional domains to its localization and function. We find that Stu2 colocalizes with kinetochores using its TOG domains, which bind GTP-tubulin, a coiled-coil homodimerization domain, and a domain that interacts with plus-end interacting proteins. Stu2 localization is also promoted by phosphorylation at a putative CDK1 phosphorylation site located within its microtubule-binding basic patch. Surprisingly, however, we find that kinetochore force generation is uncorrelated with the amount of kinetochore-colocalized Stu2. These and other data imply that Stu2 colocalizes with kinetochores by recognizing growing microtubule plus ends within yeast kinetochores. We propose that Stu2 destabilizes these plus ends to indirectly contribute to the "catch-bond" activity of the kinetochores.

SUBMITTER: Humphrey L

PROVIDER: S-EPMC5996951 | biostudies-literature | 2018 Feb

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Stu2 acts as a microtubule destabilizer in metaphase budding yeast spindles.

Humphrey Lauren L Felzer-Kim Isabella I Joglekar Ajit P AP

Molecular biology of the cell 20171129 3

The microtubule-associated protein Stu2 (XMAP215) has the remarkable ability to act either as a polymerase or as a destabilizer of the microtubule plus end. In budding yeast, it is required for the dynamicity of spindle microtubules and also for kinetochore force generation. To understand how Stu2 contributes to these distinct activities, we analyzed the contributions of its functional domains to its localization and function. We find that Stu2 colocalizes with kinetochores using its TOG domains ...[more]

PMID: 29187578

Similar Datasets

Project description:Fluorescently labeled proteins absorb and emit light, appearing as Gaussian spots in fluorescence imaging. When fluorescent tags are added to cytoskeletal polymers such as microtubules, a line of fluorescence and even non-linear structures results. While much progress has been made in techniques for imaging and microscopy, image analysis is less well-developed. Current analysis of fluorescent microtubules uses either manual tools, such as kymographs, or automated software. As a result, our ability to quantify microtubule dynamics and organization from light microscopy remains limited. Despite the development of automated microtubule analysis tools for in vitro studies, analysis of images from cells often depends heavily on manual analysis. One of the main reasons for this disparity is the low signal-to-noise ratio in cells, where background fluorescence is typically higher than in reconstituted systems. Here, we present the Toolkit for Automated Microtubule Tracking (TAMiT), which automatically detects, optimizes, and tracks fluorescent microtubules in living yeast cells with sub-pixel accuracy. Using basic information about microtubule organization, TAMiT detects linear and curved polymers using a geometrical scanning technique. Images are fit via an optimization problem for the microtubule image parameters that are solved using non-linear least squares in Matlab. We benchmark our software using simulated images and show that it reliably detects microtubules, even at low signal-to-noise ratios. Then, we use TAMiT to measure monopolar spindle microtubule bundle number, length, and lifetime in a large dataset that includes several S. pombe mutants that affect microtubule dynamics and bundling. The results from the automated analysis are consistent with previous work and suggest a direct role for CLASP/Cls1 in bundling spindle microtubules. We also illustrate automated tracking of single curved astral microtubules in S. cerevisiae, with measurement of dynamic instability parameters. The results obtained with our fully-automated software are similar to results using hand-tracked measurements. Therefore, TAMiT can facilitate automated analysis of spindle and microtubule dynamics in yeast cells.

Dataset Information

Stu2 acts as a microtubule destabilizer in metaphase budding yeast spindles.

Publications

Stu2 acts as a microtubule destabilizer in metaphase budding yeast spindles.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets