Opto-Katanin: An Optogenetic Tool for Localized Microtubule Disassembly

Abstract

Microtubules are major cytoskeletal filaments that drive chromosome separation during cell division, serve as rails for intracellular transport and as a scaffold for organelle positioning. Experimental manipulation of microtubules is widely used in cell and developmental biology, but tools for precise subcellular spatiotemporal control of microtubule integrity are currently lacking. Here, we exploit the dependence of the mammalian microtubule-severing protein katanin on microtubule-targeting co-factors to generate a light-activated system for localized microtubule disassembly that we named opto-katanin. Targeted illumination with blue light induces rapid and localized opto-katanin recruitment and local microtubule depolymerization, which is quickly reversible after stopping light-induced activation. Opto-katanin can be employed to locally perturb microtubule-based transport and organelle morphology in dividing cells and differentiated neurons with high spatiotemporal precision. We show that different microtubule-associated proteins can be used to recruit opto-katanin to microtubules and induce severing, paving the way for spatiotemporally precise manipulation of specific microtubule subpopulations.

In Vivo Photocontrol of Microtubule Dynamics and Integrity, Migration and Mitosis, by the Potent GFP-Imaging-Compatible Photoswitchable Reagents SBTubA4P and SBTub2M

“…However, until recently, only two optogenetic tools to modulate MT dynamics and network architecture were known: Wittmann’s photo-inactivatable MT-polymerizing tool π-EB1, and Slep’s photoactivated plus tip recruitment system . Two further optogenetic tools to photocontrol enzymatic severing of MTs have recently appeared as preprints, , highlighting the interest in methods to photocontrol microtubule dynamics, network organization, and interaction partners.…”

Photoswitchable Epothilone‐Based Microtubule Stabilisers Allow GFP‐Imaging‐Compatible, Optical Control over the Microtubule Cytoskeleton**

“…[13] However, they are not widely used, likely due to slow kinetics of the hydrolytic step in the photouncaging cascades, and photolability of the CA4 stilbene. [14] Two optogenetic tools to depress MT polymerisation under light are reported: Wittmann's photo-inactivat-able π-EB1, [15] and Slep's photoactivated plus tip recruitment system; [16] and in recent weeks, two preprints of photocontrolled MT-severing enzymes have also appeared, [17,18] highlighting broad interest in methods to photocontrol microtubule dynamics and network organisation.…”