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Complementary Superresolution Visualization of Composite Plant Microtubule Organization and Dynamics
Tereza Vavrdová1, Pavel Křenek1, Miroslav Ovečka1
1Department of Cell Biology, Centre of the Region Haná for Biotechnological and Agricultural Research, Faculty of Science, Palacký University Olomouc, Olomouc, Czechia.
Frontiers in Plant Science
|June 26, 2020
Summary
Microtubule-associated protein MAP65-2 dynamics were visualized in Arabidopsis cells using advanced super-resolution microscopy. This study reveals how MAP65-2 crossbridges organize microtubule bundles, crucial for plant cell structure.
Area of Science:
- Cell Biology
- Plant Biology
- Microscopy
Background:
- Microtubule bundling is vital for eukaryotic cell organization, with MAP65 proteins mediating this process in plants.
- MAP65-1 and MAP65-2 are key, redundant microtubule-associated proteins (MAPs) in Arabidopsis thaliana, essential for parallel microtubule array formation.
- High-order microtubule arrays are challenging to image with conventional microscopy, necessitating advanced techniques.
Purpose of the Study:
- To investigate the spatiotemporal localization and dynamics of MAP65-2 in Arabidopsis hypocotyl cells.
- To overcome imaging limitations of dense microtubule bundles using various super-resolution microscopy techniques.
- To understand the relationship between microtubule organization and MAP65-2 crosslinking activity.
Main Methods:
- Utilized Arabidopsis lines expressing fluorescent protein fusions of MAP65-2 and tubulin.
- Employed super-resolution microscopy: Airyscan confocal laser scanning microscopy (ACLSM), structured illumination microscopy (SIM), total internal reflection SIM (TIRF-SIM), and photoactivation localization microscopy (PALM).
- Combined SIM and ACLSM for detailed spatiotemporal analysis; used single molecule localization microscopy (SMLM) for high-resolution distribution.
Main Results:
- Spatiotemporal patterns of MAP65-2 localization and dynamics within microtubule bundles were successfully visualized.
- SIM, TIRF-SIM, and ACLSM proved effective for tracking MAP65-2 dynamics in complex bundles.
- Correlated end-wise microtubule dynamics with MAP65-2 fluctuations using ACLSM.
Conclusions:
- The combination of multiple super-resolution methods provided unprecedented insights into MAP65-2 distribution and dynamics.
- This study elucidates the role of MAP65-2 as a crosslinking protein in organizing microtubule arrays in living Arabidopsis cells.
- Advanced imaging techniques are crucial for dissecting the complex architecture and dynamics of microtubule bundles.
