Light-Induced Dimerization Approaches to Control Cellular Processes

Abstract

Light-inducible approaches provide a means to control biological systems with spatial and temporal resolution that is unmatched by traditional genetic perturbations. Recent developments of optogenetic and chemo-optogenetic systems for induced proximity in cells facilitate rapid and reversible manipulation of highly dynamic cellular processes and have become valuable tools in diverse biological applications. New expansions of the toolbox facilitate control of signal transduction, genome editing, "painting" patterns of active molecules onto cellular membranes, and light-induced cell cycle control. A combination of light- and chemically induced dimerization approaches have also seen interesting progress. Herein, an overview of optogenetic systems and emerging chemo-optogenetic systems is provided, and recent applications in tackling complex biological problems are discussed.

Keywords: chemo-optogenetics; dimerization; optogenetics; photochemistry; proteins.

Figure 1

Optogenetic dimerization systems. a) UVR8 proteins homodimerize in the dark, and dissociate to monomers and bind to COP1 upon photoinduced (λ=290–315 nm) conformational change. b) Illumination of LOV2 at λ=440–473 nm causes unwinding of the Jα helix and uncaging of the protein of interest (POI). c) Photoactivation of CRY2 at λ=450–488 nm causes a conformational change, leading to oligomerization of CRY2 or interaction with CIB1. d) The fluorescent Dronpa proteins form tetra‐ or homodimers in the dark and dissociate after illumination at λ=490 nm. The process is reversible by illumination at λ=400 nm. e) CBDs oligomerize in the dark and dissociate upon green‐light illumination. f) PhyB interacts with PIF3/6 under illumination at λ=650 nm, which can be reversed by illumination at λ=750 nm. g) BphP1 interacts with PpsR2 under illumination at λ=750 nm, which can be reversed by illumination at λ=650 nm or in the dark. Chromophores: flavin for LOV2 and CRY2, AdoCbl for CBD, PCB for PhyB and Cph1, and biliverdin for BphP1.

Figure 2

Chemo‐optogenetic dimerization systems and structures of photosensitive dimerizers. Dotted red lines indicate photocleavage sites.

Figure 3

Biological applications of light‐induced dimerization systems. a) The LightOn strategy with the VVD homodimerization system to control gene transcription. b) A combination of the CYR2‐CIB1 system with dCas9 and transcriptional modulators. Blue‐light‐induced dimerization recruited the transcriptional activator to a specific site in the genome targeted by dCas9, leading to upregulation of an endogenous gene. c) The optoSOS approach with the PhyB‐PIF system generated dynamic input stimulus patterns by red light. The altered pathway output downstream of signaling was monitored. d) Nvoc‐TMP‐CI molecules are anchored to the plasma membrane through artificial receptors immobilized on the glass surface. Uncaging with a focused laser beam allows patterns of activated signaling molecules to be generated at the plasma membrane. e) Organelle transport is controlled through the CONC system. Uncaging of the coumarinyl group stimulated recruitment of motor proteins to the cargo and subsequent transport along microtubules. Photocleavage of the NVOC group led to dissociation of motor proteins and cessation of transport. f) Maintaining the spindle checkpoint protein Mad1 on a subpopulation of kinetochores through the photocleavable TNH system was sufficient to halt the transition from metaphase to anaphase. Mad1 was released from a subpopulation of kinetochores within the cell by light. g) Uncaging of NvocTMP‐Cl stimulated recruitment of Rac1 from the cytosol to the plasma membrane through dimerization between HaloTag and eDHFR. Addition of SLF′‐TMP triggered the dissociation of Rac1 and translocation to the nucleus through dimerization between eDHFR and FKBP′. Addition of TMP, which competed for binding to eDHFR, led to release of Rac1 to the cytosol. Multiple cycles could be achieved through washing out.