A study of the blue-light-dependent phosphorylation, degradation, and photobody formation of Arabidopsis CRY2.
Arabidopsis cryptochrome 2 (CRY2) is a blue-light receptor mediating blue-light inhibition of hypocotyl elongation and photoperiodic promotion of floral initiation. CRY2 is a constitutive nuclear protein that undergoes blue-light-dependent phosphorylation, ubiquitination, photobody formation, and degradation in the nucleus, but the relationship between these blue-light-dependent events remains unclear. It has been proposed that CRY2 phosphorylation triggers a conformational change responsible for the subsequent ubiquitination and photobody formation, leading to CRY2 function and/or degradation. We tested this hypothesis by a structure-function study, using mutant CRY2-GFP fusion proteins expressed in transgenic Arabidopsis. We show that changes of lysine residues of the NLS (Nuclear Localization Signal) sequence of CRY2 to arginine residues partially impair the nuclear importation of the CRY2K541R and CRY2K554/5R mutant proteins, resulting in reduced phosphorylation, physiological activities, and degradation in response to blue light. In contrast to the wild-type CRY2 protein that forms photobodies exclusively in the nucleus, the CRY2K541R and CRY2K554/5R mutant proteins form protein bodies in both the nucleus and cytosol in response to blue light. These results suggest that photoexcited CRY2 molecules can aggregate to form photobody-like structure without the nucleus-dependent protein modifications or the association with the nuclear CRY2-interacting proteins. Taken together, the observation that CRY2 forms photobodies markedly faster than CRY2 phosphorylation in response to blue light, we hypothesize that the photoexcited cryptochromes form oligomers, preceding other biochemical changes of CRY2, to facilitate photobody formation, signal amplification, and propagation, as well as desensitization by degradation.
Institute of Crop Science, Chinese Academy of Agricultural Sciences, Beijing, 100080, China.
Ze-Cheng
ZC
Zuo
Ying-Ying
YY
Meng
Xu-Hong
XH
Yu
Zeng-Lin
ZL
Zhang
De-Shun
DS
Feng
Shih-Fan
SF
Sun
Bin
B
Liu
Chen-Tao
CT
Lin
D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000235:genetics / Q000254:growth & development / Q000378:metabolism* / Q000528:radiation effects*; D029681:Arabidopsis Proteins / Q000737:chemistry* / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism / Q000528:radiation effects; D056931:Cryptochromes / Q000737:chemistry* / Q000378:metabolism*; D018546:Hypocotyl / Q000254:growth & development / Q000528:radiation effects; D008027:Light; D008239:Lysine / Q000378:metabolism; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D019913:Nuclear Localization Signals / Q000737:chemistry / Q000378:metabolism; D010766:Phosphorylation / Q000528:radiation effects; D030821:Plants, Genetically Modified; D020836:Protein Structure, Quaternary; D021381:Protein Transport / Q000528:radiation effects; D059748:Proteolysis / Q000528:radiation effects*
D029681:Arabidopsis Proteins; C103618:CRY2 protein, Arabidopsis; D056931:Cryptochromes; D050505:Mutant Proteins; D019913:Nuclear Localization Signals; D008239:Lysine