Identification of two amino acids in the C-terminal domain of mouse CRY2 essential for PER2 interaction.
BACKGROUND
Cryptochromes (CRYs) are a class of flavoprotein blue-light signaling receptors found in plants and animals, and they control plant development and the entrainment of circadian rhythms. They also act as integral parts of the central circadian oscillator in humans and other animals. In mammals, the CLOCK-BMAL1 heterodimer activates transcription of the Per and Cry genes as well as clock-regulated genes. The PER2 proteins interact with CRY and CKIε, and the resulting ternary complexes translocate into the nucleus, where they negatively regulate the transcription of Per and Cry core clock genes and other clock-regulated output genes. Recent studies have indicated that the extended C-termini of the mammalian CRYs, as compared to photolyase proteins, interact with PER proteins.


RESULTS
We identified a region on mCRY2 (between residues 493 and 512) responsible for direct physical interaction with mPER2 by mammalian two-hybrid and co-immunoprecipitation assays. Moreover, using oligonucleotide-based degenerate PCR, we discovered that mutation of Arg-501 and Lys-503 of mCRY2 within this C-terminal region totally abolishes interaction with PER2.


CONCLUSIONS
Our results identify mCRY2 amino acid residues that interact with the mPER2 binding region and suggest the potential for rational drug design to inhibit CRYs for specific therapeutic approaches.
Ozber
Natali
N
Material Science and Engineering, College of Engineering, Koc University, Rumeli Feneri Yolu, 34450 Sariyer, Istanbul, Turkey.
Baris
Ibrahim
I
Tatlici
Gulnaz
G
Gur
Ibrahim
I
Kilinc
Seda
S
Unal
Evrim B
EB
Kavakli
Ibrahim H
IH
D056930:ARNTL Transcription Factors / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D056926:CLOCK Proteins / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D002940:Circadian Rhythm / Q000235:genetics; D056931:Cryptochromes / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D016296:Mutagenesis; D056950:Period Circadian Proteins / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D020798:Two-Hybrid System Techniques
D056930:ARNTL Transcription Factors; C538961:Bmal1 protein, mouse; C538967:Cry2 protein, mouse; D056931:Cryptochromes; C513312:Per2 protein, mouse; D056950:Period Circadian Proteins; D056926:CLOCK Proteins; C538944:Clock protein, mouse