Specific sequences outside the E-box are required for proper per expression and behavioral rescue.
A 69 bp circadian regulatory sequence (CRS) upstream of the per gene is sufficient to drive circadian transcription, mediate proper spatial expression, and rescue behavioral rhythmicity in per01 flies. Within the CRS, an E-box is required for transcriptional activation by two basic-helix-loop-helix (bHLH) PERARNT-SIM (PAS) transcription factors, dCLOCK (dCLK) and CYCLE (CYC). To define sequences within the CRS that are required for spatial expression, circadian expression, and behavioral rhythmicity, a series of mutants that alter blocks of 3 to 12 nucleotides across the entire CRS were used to drive lacZ or per expression in vivo. As expected, the E-box within the CRS is necessary for high-level expression and behavioral rhythmicity, but sequences outside the E-box are also required for transcriptional activation, proper spatial expression, and behavioral rhythmicity. These results indicate that the dCLK-CYC target site extends beyond the E-box and that factors other than dCLK and CYC modulate per transcription.
Lyons
L C
LC
Department of Biology and Biochemistry, University of Houston, TX, USA.
Darlington
T K
TK
Hao
H
H
Houl
J
J
Kay
S A
SA
Hardin
P E
PE
D056930:ARNTL Transcription Factors; D000818:Animals; D030801:Animals, Genetically Modified; D001483:Base Sequence; D051792:Basic Helix-Loop-Helix Proteins; D056926:CLOCK Proteins; D002940:Circadian Rhythm / Q000235:genetics / Q000502:physiology*; D004330:Drosophila / Q000235:genetics / Q000502:physiology*; D029721:Drosophila Proteins; D005786:Gene Expression Regulation / Q000502:physiology*; D017930:Genes, Reporter; D018257:Helix-Loop-Helix Motifs; D019476:Insect Proteins / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009687:Nuclear Proteins / Q000235:genetics*; D056950:Period Circadian Proteins; D012045:Regulatory Sequences, Nucleic Acid; D015534:Trans-Activators / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism; D015533:Transcriptional Activation; D001616:beta-Galactosidase / Q000235:genetics
D056930:ARNTL Transcription Factors; D051792:Basic Helix-Loop-Helix Proteins; D056926:CLOCK Proteins; D029721:Drosophila Proteins; D019476:Insect Proteins; D009687:Nuclear Proteins; D056950:Period Circadian Proteins; D015534:Trans-Activators; D014157:Transcription Factors; D001616:beta-Galactosidase; C120526:Clk protein, Drosophila; C539040:PER protein, Drosophila; C112805:cyc protein, Drosophila