A chemical biology approach reveals period shortening of the mammalian circadian clock by specific inhibition of GSK-3beta.
The circadian clock controls daily oscillations of gene expression at the cellular level. We report the development of a high-throughput circadian functional assay system that consists of luminescent reporter cells, screening automation, and a data analysis pipeline. We applied this system to further dissect the molecular mechanisms underlying the mammalian circadian clock using a chemical biology approach. We analyzed the effect of 1,280 pharmacologically active compounds with diverse structures on the circadian period length that is indicative of the core clock mechanism. Our screening paradigm identified many compounds previously known to change the circadian period or phase, demonstrating the validity of the assay system. Furthermore, we found that small molecule inhibitors of glycogen synthase kinase 3 (GSK-3) consistently caused a strong short period phenotype in contrast to the well-known period lengthening by lithium, another presumed GSK-3 inhibitor. siRNA-mediated knockdown of GSK-3beta also caused a short period, confirming the phenotype obtained with the small molecule inhibitors. These results clarify the role of GSK-3beta in the period regulation of the mammalian clockworks and highlight the effectiveness of chemical biology in exploring unidentified mechanisms of the circadian clock.
Hirota
Tsuyoshi
T
Cell and Developmental Biology, Division of Biological Sciences, University of California San Diego, 9500 Gilman Drive, La Jolla, CA 92093, USA.
Lewis
Warren G
WG
Liu
Andrew C
AC
Lee
Jae Wook
JW
Schultz
Peter G
PG
Kay
Steve A
SA
D001683:Biological Clocks / Q000187:drug effects* / Q000235:genetics; D045744:Cell Line, Tumor; D002940:Circadian Rhythm / Q000187:drug effects* / Q000235:genetics; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D005786:Gene Expression Regulation / Q000187:drug effects / Q000235:genetics; D055785:Gene Knockdown Techniques; D038362:Glycogen Synthase Kinase 3 / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism*; D000071679:Glycogen Synthase Kinase 3 beta; D006801:Humans; D008094:Lithium / Q000494:pharmacology; D034741:RNA, Small Interfering / Q000235:genetics
D004791:Enzyme Inhibitors; D034741:RNA, Small Interfering; D008094:Lithium; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; D038362:Glycogen Synthase Kinase 3