CRY2

Cryptochromes (from the Greek κρυπτός χρώμα, "hidden colour") are a class of flavoproteins found in plants and animals that are sensitive to blue light. They are involved in the circadian rhythms and the sensing of magnetic fields in a number of species. The name ''cryptochrome'' was proposed as a ''portmanteau'' nature of the photoreceptor, and the ''cryptogamic'' organisms on which many blue-light studies were carried out.

The two genes ''Cry1'' and ''Cry2'' code the two cryptochrome proteins CRY1 and CRY2. In insects and plants, CRY1 regulates the circadian clock in a light-dependent fashion, whereas in mammals, CRY1 and CRY2 act as light-independent inhibitors of CLOCK-BMAL1 components of the circadian clock. In plants, blue-light photoreception can be used to cue developmental signals. Besides chlorophylls, cryptochromes are the only proteins known to form photoinduced radical-pairs ''in vivo''. These appear to enable some animals to detect magnetic fields.

Cryptochromes have been the focus of several current efforts in optogenetics. Employing transfection, initial studies on yeast have capitalized on the potential of Cry2 heterodimerization to control cellular processes, including gene expression, by light.

Discovery

Although Charles Darwin first documented plant responses to blue light in the 1880s, it was not until the 1980s that research began to identify the pigment responsible. In 1980, researchers discovered that the HY4 gene of the plant ''Arabidopsis thaliana'' was necessary for the plant's blue light sensitivity, and, when the gene was sequenced in 1993, it showed high sequence homology with photolyase, a DNA repair protein activated by blue light. By 1995, it became clear that the products of the HY4 gene and its two human homologs did not exhibit photolyase activity and were instead a new class of blue light photoreceptor hypothesized to be circadian photopigments. In 1996 and 1998, ''Cry'' homologs were identified in ''Drosophila'' and mice, respectively.

Evolutionary history and structure

Cryptochromes (CRY1, CRY2) are evolutionarily old and highly conserved proteins that belong to the flavoproteins superfamily that exists in all kingdoms of life. All members of this superfamily have the characteristics of an N-terminal photolyase homology (PHR) domain. The PHR domain can bind to the flavin adenine dinucleotide (FAD) cofactor and a light-harvesting chromophore. Cryptochromes are derived from and closely related to photolyases, which are bacterial enzymes that are activated by light and involved in the repair of UV-induced DNA damage. In eukaryotes, cryptochromes no longer retain this original enzymatic activity. The structure of cryptochrome involves a fold very similar to that of photolyase, with a single molecule of FAD noncovalently bound to the protein. These proteins have variable lengths and surfaces on the C-terminal end, due to the changes in genome and appearance that result from the lack of DNA repair enzymes. The Ramachandran plot shows that the secondary structure of the CRY1 protein is primarily a right-handed alpha helix with little to no steric overlap. The structure of CRY1 is almost entirely made up of alpha helices, with several loops and few beta sheets. The molecule is arranged as an orthogonal bundle.

Function

Phototropism

In plants, cryptochromes mediate phototropism, or directional growth toward a light source, in response to blue light. This response is now known to have its own set of photoreceptors, the phototropins.

Unlike phytochromes and phototropins, cryptochromes are not kinases. Their flavin chromophore is reduced by light and transported into the cell nucleus, where it affects the turgor pressure and causes subsequent stem elongation. To be specific, ''Cry2'' is responsible for blue-light-mediated cotyledon and leaf expansion. ''Cry2'' overexpression in transgenic plants increases blue-light-stimulated cotyledon expansion, which results in many broad leaves and no flowers rather than a few primary leaves with a flower. A double loss-of-function mutation in Arabidopsis thaliana Early Flowering 3 (elf3) and Cry2 genes delays flowering under continuous light and was shown to accelerate it during long and short days, which suggests that Arabidopsis CRY2 may play a role in accelerating flowering time during continuous light.

Photomorphogenesis

Cryptochromes receptors cause plants to respond to blue light via photomorphogenesis. They help control seed and seedling development, as well as the switch from the vegetative to the flowering stage of development. In Arabidopsis, it has been shown that cryptochromes controls plant growth during sub-optimal blue-light conditions.

Light capture

Despite much research on the topic, cryptochrome photoreception and phototransduction in ''Drosophila'' and ''Arabidopsis thaliana'' is still poorly understood. Cryptochromes are known to possess two chromophores: pterin (in the form of 5,10-methenyltetrahydrofolic acid (MTHF)) and flavin (in the form of FAD). Both may absorb a photon, and in ''Arabidopsis'', pterin appears to absorb at a wavelength of 380 nm and flavin at 450 nm. Past studies have supported a model by which energy captured by pterin is transferred to flavin. Under this model of phototransduction, FAD would then be reduced to FADH, which probably mediates the phosphorylation of a certain domain in cryptochrome. This could then trigger a signal transduction chain, possibly affecting gene regulation in the cell nucleus.

A new hypothesis proposes that in plant cryptochromes, the transduction of the light signal into a chemical signal that might be sensed by partner molecules could be triggered by a photo-induced negative charge within the protein - on the FAD cofactor or on the neighbouring aspartic acid. This negative charge would electrostatically repel the protein-bound ATP molecule and thereby also the protein C-terminal domain, which covers the ATP binding pocket prior to photon absorption. The resulting change in protein conformation could lead to phosphorylation of previously inaccessible phosphorylation sites on the C-terminus and the given phosphorylated segment could then liberate the transcription factor HY5 by competing for the same binding site at the negative regulator of photomorphogenesis COP1.

A different mechanism may function in ''Drosophila''. The true ground state of the flavin cofactor in ''Drosophila'' CRY is still debated, with some models indicating that the FAD is in an oxidized form, while others support a model in which the flavin cofactor exists in anion radical form, •. Recently, researchers have observed that oxidized FAD is readily reduced to • by light. Furthermore, mutations that blocked photoreduction had no effect on light-induced degradation of CRY, while mutations that altered the stability of • destroyed CRY photoreceptor function. These observations provide support for a ground state of •. Researchers have also recently proposed a model in which is excited to its doublet