Dynamic crystallography reveals early signalling events in ultraviolet photoreceptor UVR8

Abstract

Arabidopsis thaliana UVR8 (AtUVR8) is a long-sought-after photoreceptor that undergoes dimer dissociation in response to UV-B light. Crystallographic and mutational studies have identified two crucial tryptophan residues for UV-B responses in AtUVR8. However, the mechanism of UV-B perception and structural events leading up to dimer dissociation remain elusive at the molecular level. We applied dynamic crystallography to capture light-induced structural events in photoactive AtUVR8 crystals. Here we report two intermediate structures at 1.67Å resolution. At the epicenter of UV-B signaling, concerted motions associated with Trp285/Trp233 lead to ejection of a water molecule, which weakens an intricate network of hydrogen bonds and salt bridges at the dimer interface. Partial opening of the β-propeller structure due to thermal relaxation of conformational strains originating in the epicenter further disrupts the dimer interface and leads to dimer dissociation. These dynamic crystallographic observations provide structural insights into the photo-perception and signaling mechanism of UVR8.

Photoactivation mechanism of a carotenoid-based photoreceptor

“…Given the rod shape of OCP crystals, we were able to collect both the dark and light datasets from separate segments of the same crystal to achieve the best difference signals and to avoid possible effects due to X-ray radiation damage. This T-scan strategy has unraveled remarkable light-induced structural changes in other photoreceptors such as bacteriophytochrome and UVR8 (30,38). These results demonstrate that it is entirely possible to capture large-scale structural motions in the crystal lattice when a photoactive crystal is illuminated at cryogenic temperatures near the glass transition temperature (30,43).…”

“…However, the absence of spectral changes does not rule out photoactivity or existence of any spectroscopically silent structural changes. A good example is the UV-B photoreceptor from plant UVR8, which does not show any detectable light-induced absorption spectral changes in a steady-state experiment, but undergoes UV-B-induced dimer dissociation resulting in visible cracks in crystals (30). We postulate that the spectral change of OCP observed in solution is the consequence rather than the cause of light-induced conformational changes (9,17).…”

“…The crystal lattice evidently imposes an energy barrier that hinders further development of larger conformational changes required for the full activation of OCP. On the other hand, such lattice constraint and cryogenic temperature offer an invaluable kinetic trap for crystallographic observations of those structural events too unstable to capture at room temperature (30,38,43).…”

A Comprehensive Phylogeny Reveals Functional Conservation of the UV-B Photoreceptor UVR8 from Green Algae to Higher Plants

“…Each UVR8 monomer contains the conserved pentapeptide repeat Gly-Trp-Arg-His-Thr (GWRHT) in blades 5, 6, and 7. This motif generates a triad of closely packed tryptophans (W233, W285, and W337) which are key for UV-B photoreception, W285 being the main UV-B sensor (Christie et al, 2012; Wu et al, 2012; Zeng et al, 2015). W233 is also important, both in photoreception and in maintaining exciton coupling, whereas W337 plays an auxiliary role (Christie et al, 2012; Wu et al, 2012).…”

UV-B Perception and Acclimation in Chlamydomonas reinhardtii

“…The plant photoreceptor UV RESISTANCE LOCUS8 (UVR8) specifically perceives UV-B via an intrinsic tryptophan-based mechanism, mainly involving Trp-285 and Trp-233 (Rizzini et al, 2011;Christie et al, 2012;Wu et al, 2012). UVR8 is a seven-bladed b-propeller protein that exists as a homodimer maintained by interactions of charged amino acids across the dimer interaction surface (Rizzini et al, 2011;Christie et al, 2012;Wu et al, 2012;Zeng et al, 2015). The charged amino acids that maintain the homodimer are adjacent to the tryptophans involved in photoreception, suggesting that excitation of these tryptophans by UV-B neutralizes dimer-maintaining interactions Wu et al, 2012;Mathes et al, 2015).…”