Photopharmacological Manipulation of Mammalian CRY1 for Regulation of the Circadian Clock

Abstract

CRY1 and CRY2 proteins are highly conserved components of the circadian clock that controls daily physiological rhythms. Disruption of CRY functions are related to many diseases, including circadian sleep phase disorder. Development of isoformselective and spatiotemporally controllable tools will facilitate the understanding of shared and distinct functions of CRY1 and CRY2. Here, we developed CRY1-selective compounds that enable lightdependent manipulation of the circadian clock. From phenotypic chemical screening in human cells, we identified benzophenone derivatives that lengthened the circadian period. These compounds selectively interacted with the CRY1 photolyase homology region, resulting in activation of CRY1 but not CRY2. The benzophenone moiety rearranged a CRY1 region called the "lid loop" located outside of the compound-binding pocket and formed a unique interaction with Phe409 in the lid loop. Manipulation of this key interaction was achieved by rationally designed replacement of the benzophenone with a switchable azobenzene moiety whose cis−trans isomerization can be controlled by light. The metastable cis form exhibited sufficiently high half-life in aqueous solutions and structurally mimicked the benzophenone unit, enabling reversible period regulation over days by cellular irradiation with visible light. This study revealed an unprecedented role of the lid loop in CRY-compound interaction and paves the way for spatiotemporal regulation of CRY1 activity by photopharmacology for molecular understanding of CRY1-dependent functions in health and disease.

Cryptochromes: Photochemical and structural insight into magnetoreception

“…Similarly, drug‐discovery efforts in Type II CRYs indicate that elements within the CTT interact with the C‐terminal lid and PBM to gate isoform selectivity in CRY‐regulator compounds (Figure 4(c)). 88,102 Second, in plant CRYs lacking the CTT, ATP binds to the DNA binding cleft (Figure 4(b)), and light induced autophosphorylation of CTT residues is coupled to ATP recognition at this site 103 . In vivo, ATP stabilizes the open conformation where ATP and the CTT compete for binding 73,103 .…”

Structural and Chemical Biology Approaches Reveal Isoform-Selective Mechanisms of Ligand Interactions in Mammalian Cryptochromes

“…Used as tools to activate CRY1 and CRY2, KL101 and TH301 helped to identify CRY-mediated activation of brown adipocyte differentiation (Miller et al, 2020b). Furthermore, a similar approach identified other CRY1 isoform-selective activators, the thienopyrimidine derivative KL201 (Miller et al, 2020a), and benzophenone derivatives TH303 and TH129 (Kolarski et al, 2021a), which together provide molecular tools to investigate CRY1 functions (Figure 1).…”

Active Ester Functionalized Azobenzenes as Versatile Building Blocks

“…Photoswitchable molecules add photoresponsiveness to diverse materials such as polymers [ 1 , 2 , 3 , 4 , 5 , 6 ], dendrimers [ 7 , 8 ] or biomolecules [ 9 , 10 , 11 , 12 , 13 , 14 , 15 , 16 , 17 , 18 , 19 , 20 , 21 , 22 , 23 ]. By precise adjustment of the synthetic strategy the switch can be incorporated into the target material and its switching properties can be utilized.…”

“…Due to its fast and efficient photoswitching characteristics, as well as its high fatigue-resistance [ 24 ], azobenzene is widely used in versatile applications such as molecular machines [ 3 , 25 , 26 , 27 , 28 ], data storage materials [ 29 , 30 , 31 , 32 ], or photopharmacology [ 9 , 10 , 11 , 12 , 13 , 14 , 15 , 16 , 17 , 18 , 19 , 20 , 21 , 22 , 23 ]. Upon irradiation with UV-light, typical azobenzenes undergo isomerization from the thermodynamically favored ( E )-isomer to the less stable ( Z )-isomer ( Figure 1 , left).…”