Precise control of microtubule disassembly in living cells

• Published in: The EMBO journal 2022-08, Vol.41 (15), p.e110472-n/a
• Main Authors: Liu, Grace Y, Chen, Shiau‐Chi, Lee, Gang‐Hui, Shaiv, Kritika, Chen, Pin‐Yu, Cheng, Hsuan, Hong, Shi‐Rong, Yang, Wen‐Ting, Huang, Shih‐Han, Chang, Ya‐Chu, Wang, Hsien‐Chu, Kao, Ching‐Lin, Sun, Pin‐Chiao, Chao, Ming‐Hong, Lee, Yian‐Ying, Tang, Ming‐Jer, Lin, Yu‐Chun
• Format: Article
• Language: English
• Subjects: Cell death; Cell division; Cells (biology); Cilia; Contractility; Dimerization; Dismantling; Disruption; EMBO22; EMBO57; Enzymes; Fibers; Genetics; Glutamine; Golgi apparatus; Information processing; intercellular bridge; Interphase; Membrane potential; microtubule; Microtubules; Microtubules - metabolism; Mitochondria; mitotic spindle; Mutation; Optics; Populations; primary cilium; Recruitment; Resource; Rigidity; spastin; Spindle Apparatus - metabolism; Spindles
• ISSN: 0261-4189; 1460-2075; 1460-2075
• DOI: 10.15252/embj.2021110472

Microtubules tightly regulate various cellular activities. Our understanding of microtubules is largely based on experiments using microtubule‐targeting agents, which, however, are insufficient to dissect the dynamic mechanisms of specific microtubule populations, due to their slow effects on the entire pool of microtubules. To overcome this technological limitation, we have used chemo and optogenetics to disassemble specific microtubule subtypes, including tyrosinated microtubules, primary cilia, mitotic spindles, and intercellular bridges, by rapidly recruiting engineered microtubule‐cleaving enzymes onto target microtubules in a reversible manner. Using this approach, we show that acute microtubule disassembly swiftly halts vesicular trafficking and lysosomal dynamics. It also immediately triggers Golgi and ER reorganization and slows the fusion/fission of mitochondria without affecting mitochondrial membrane potential. In addition, cell rigidity is increased after microtubule disruption owing to increased contractile stress fibers. Microtubule disruption furthermore prevents cell division, but does not cause cell death during interphase. Overall, the reported tools facilitate detailed analysis of how microtubules precisely regulate cellular architecture and functions. SYNOPSIS Characterization of microtubules in cells via microtubule‐targeting agents is not ideal for assessing dynamics of specific microtubule populations. Here, a novel approach using an engineered microtubule‐severing enzyme allows spatiotemporal manipulation of microtubule disassembly triggered by either chemicals or illumination. A triple glutamine mutation in Spastin (dNSpastin3Q) reduces its association with microtubules. Engineered dNSpastin3Q acts as a microtubule‐severing enzyme in living cells. Recruitment of dNSpastin3Q to microtubules can be controlled by chemical‐ or light‐induced dimerization. Induced dimerization of dNSpastin3Q leads to acute disassembly of target microtubule subtypes. Graphical Abstract Spatiotemporal manipulation of microtubules by either chemical treatment or light allows assessing their function in cells.