Expression of the gene for Dec2, a basic helix-loop-helix transcription factor, is regulated by a molecular clock system.
Dec2, a member of the basic helix-loop-helix superfamily, is a recently confirmed regulatory protein for the clockwork system. Transcripts of Dec2, as well as those of its related gene Dec1, exhibit a striking circadian oscillation in the suprachiasmatic nucleus, and Dec2 inhibits transcription from the Per1 promoter induced by Clock/Bmal1 [Honma, Kawamoto, Takagi, Fujimoto, Sato, Noshiro, Kato and Honma (2002) Nature (London) 419, 841-844]. It is known that mammalian circadian rhythms are controlled by molecular clockwork systems based on negative-feedback loop(s), but the molecular mechanisms for the circadian regulation of Dec2 gene expression have not been clarified. We show here that transcription of the Dec2 gene is regulated by several clock molecules and a negative-feedback loop. Luciferase and gel retardation assays showed that expression of Dec2 was negatively regulated by binding of Dec2 or Dec1 to two CACGTG E-boxes in the Dec2 promoter. Forced expression of Clock/Bmal1 and Clock/Bmal2 markedly increased Dec2 mRNA levels, and up-regulated the transcription of the Dec2 gene through the CACGTG E-boxes. Like Dec, Cry and Per also suppressed Clock/Bmal-induced transcription from the Dec2 promoter. Moreover, the circadian expression of Dec2 transcripts was abolished in the kidney of Clock/Clock mutant mice. These findings suggest that the Clock/Bmal heterodimer enhances Dec2 transcription via the CACGTG E-boxes, whereas the induced transcription is suppressed by Dec2, which therefore must contribute to its own rhythmic expression. In addition, Cry and Per may also modulate Dec2 transcription.
Hamaguchi
Hidenori
H
Department of Dental and Medical Biochemistry, Graduate School of Biomedical Sciences, Hiroshima University, 1-2-3 Kasumi, Minami-ku, Hiroshima 734-8553, Japan.
Fujimoto
Katsumi
K
Kawamoto
Takeshi
T
Noshiro
Mitsuhide
M
Maemura
Koji
K
Takeda
Norihiko
N
Nagai
Ryozo
R
Furukawa
Masae
M
Honma
Sato
S
Honma
Ken-ichi
K
Kurihara
Hidemi
H
Kato
Yukio
Y
D024506:5' Flanking Region / Q000235:genetics; D056930:ARNTL Transcription Factors; D000256:Adenoviridae; D000595:Amino Acid Sequence / Q000235:genetics; D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins; D001683:Biological Clocks / Q000502:physiology*; D056926:CLOCK Proteins; D002460:Cell Line; D056931:Cryptochromes; D024721:E-Box Elements / Q000235:genetics; D042783:Endothelial Cells / Q000737:chemistry / Q000378:metabolism / Q000821:virology; D025461:Feedback, Physiological / Q000235:genetics; D005420:Flavoproteins / Q000235:genetics; D005786:Gene Expression Regulation / Q000235:genetics / Q000502:physiology*; D018257:Helix-Loop-Helix Motifs / Q000235:genetics*; D018398:Homeodomain Proteins / Q000378:metabolism; D006801:Humans; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D041681:NIH 3T3 Cells / Q000737:chemistry / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000502:physiology; D012333:RNA, Messenger / Q000235:genetics / Q000502:physiology; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017422:Sequence Analysis, DNA / Q000379:methods; D015534:Trans-Activators / Q000172:deficiency / Q000235:genetics / Q000502:physiology; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism / Q000502:physiology; D014471:Umbilical Veins / Q000166:cytology
D056930:ARNTL Transcription Factors; D051792:Basic Helix-Loop-Helix Proteins; D056926:CLOCK Proteins; D056931:Cryptochromes; D005420:Flavoproteins; D018398:Homeodomain Proteins; D012333:RNA, Messenger; D011994:Recombinant Proteins; D015534:Trans-Activators; D014157:Transcription Factors; C538960:BMAL1 protein, human; C538961:Bmal1 protein, mouse; C470968:Bhlhb3 protein, mouse; C491539:Bhlhe40 protein, mouse; C538943:CLOCK protein, human; C538944:Clock protein, mouse