A small conserved domain of Drosophila PERIOD is important for circadian phosphorylation, nuclear localization, and transcriptional repressor activity.
We identify in this study a 27-amino-acid motif which is conserved between the Drosophila melanogaster period protein (PER) and the three mammalian PERs. Characterization of PER lacking this motif (PER Delta) shows that it is important for phosphorylation of Drosophila PER by casein kinase I epsilon (CKI epsilon; doubletime protein or DBT) and CKII. S2 cell assays indicate that the domain also contributes significantly to PER nuclear localization as well as to PER transcriptional repressor activity. These two phenomena appear linked, since PER Delta transcriptional repressor activity in S2 cells was restored when nuclear localization was facilitated. Two less direct assays of PER Delta activity in flies can be interpreted similarly. The separate assay of nuclear import and export suggests that the domain functions in part to facilitate PER phosphorylation within the cytoplasm, which in turn promotes nuclear entry. As there is evidence that the kinases also function within the nucleus to promote transcriptional repression, we suggest that there is a subsequent collaboration between phosphorylated PER and the kinases to repress CLK-CYC activity, probably through the phosphorylation of CLK. This is then followed by additional PER phosphorylation, which occurs within the nucleus and leads to PER degradation.
Nawathean
Pipat
P
Howard Hughes Medical Institute and Department of Biology, Brandeis University, Waltham, MA 02454, USA.
Stoleru
Dan
D
Rosbash
Michael
M
D021581:Active Transport, Cell Nucleus; D000595:Amino Acid Sequence; D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism*; D002940:Circadian Rhythm / Q000502:physiology*; D017124:Conserved Sequence; D029721:Drosophila Proteins; D004331:Drosophila melanogaster / Q000235:genetics* / Q000502:physiology; D008954:Models, Biological; D008969:Molecular Sequence Data; D009043:Motor Activity; D016296:Mutagenesis; D050505:Mutant Proteins / Q000378:metabolism; D009687:Nuclear Proteins / Q000737:chemistry* / Q000378:metabolism*; D056950:Period Circadian Proteins; D010641:Phenotype; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D012097:Repressor Proteins / Q000378:metabolism*; D013329:Structure-Activity Relationship; D014158:Transcription, Genetic
D029721:Drosophila Proteins; D050505:Mutant Proteins; D009687:Nuclear Proteins; C539040:PER protein, Drosophila; D056950:Period Circadian Proteins; D012097:Repressor Proteins