The circadian clock-containing photoreceptor cells in Xenopus laevis express several isoforms of casein kinase I.
The frog (Xenopus laevis) retina has been an important model for the analysis of retinal circadian rhythms. In this paper, several isoforms of X. laevis casein kinase I (CKI) were analyzed to address whether they are involved in the phosphorylation and degradation of period protein (PER), as they are in the circadian oscillators of other species. cDNAs encoding two splice variants of CKI(delta) (a full-length form and deletion isoform, which is missing an exon that encodes a putative nuclear localization signal and two evolutionarily conserved protein kinase domains) were isolated and analyzed, together with a previously isolated CKI(epsilon) isoform. Both CKI(delta) and CKI(epsilon) were shown to be constitutively expressed in the photoreceptors of the retina, where a circadian clock has been localized. Both the full-length CKI(delta) and CKI(epsilon) were shown to have kinase activity in vitro, and the full-length CKI(delta) phosphorylated and degraded Drosophila PER when expressed in Drosophila S2 cells. The expression and biochemical characteristics of these CKIs are consistent with an evolutionarily conserved role for CKI in the Xenopus retinal clock. The CKI(delta) deletion isoform did not exhibit kinase activity and did not trigger degradation of PER. Subcellular localization of both CKI(delta) isoforms was cytoplasmic in several cell culture lines, but the full-length CKI(delta) , and not the deletion CKI(delta) isoform, was localized to both the nucleus and the cytoplasm in Drosophila S2 cells. These results indicate that the sequences missing in the deletion CKI(delta) isoform are important for the nuclear localization and kinase activity of the full-length isoform and that one or both of these features are necessary for degradation of Drosophila PER.
Constance
Cara M
CM
The Department of Biology, Center for Biological Timing, University of Virginia, Charlottesville, VA 22904, USA.
Fan
Jin-Yuan
JY
Preuss
Fabian
F
Green
Carla B
CB
Price
Jeffrey L
JL
D000818:Animals; D001345:Autoradiography / Q000379:methods; D015152:Blotting, Northern / Q000379:methods; D015153:Blotting, Western / Q000379:methods; D048149:Casein Kinase 1 epsilon / Q000235:genetics / Q000378:metabolism*; D048148:Casein Kinase Idelta / Q000235:genetics / Q000378:metabolism*; D002452:Cell Count; D002460:Cell Line; D002940:Circadian Rhythm / Q000502:physiology*; D003001:Cloning, Molecular / Q000379:methods; D006224:Cricetinae; D004330:Drosophila; D015870:Gene Expression / Q000502:physiology / Q000528:radiation effects; D015723:Gene Library; D014644:Genetic Variation / Q000502:physiology*; D006801:Humans; D017403:In Situ Hybridization / Q000379:methods; D016296:Mutagenesis / Q000502:physiology; D010786:Photoreceptor Cells / Q000378:metabolism*; D020033:Protein Isoforms / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D012160:Retina / Q000166:cytology* / Q000502:physiology; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D013347:Subcellular Fractions / Q000378:metabolism; D014162:Transfection / Q000379:methods; D014982:Xenopus laevis
D020033:Protein Isoforms; D012333:RNA, Messenger; D048149:Casein Kinase 1 epsilon; D048148:Casein Kinase Idelta