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NPAS2: An Analog of Clock Operative in the Mammalian Forebrain

Reick, Martin, Garcia, Joseph A., Dudley, Carol, McKnight, Steven L.·2001·Science

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Abstract

Neuronal PAS domain protein 2 (NPAS2) is a transcription factor expressed primarily in the mammalian forebrain. NPAS2 is highly related in primary amino acid sequence to Clock, a transcription factor expressed in the suprachiasmatic nucleus that heterodimerizes with BMAL1 and regulates circadian rhythm. To investigate the biological role of NPAS2, we prepared a neuroblastoma cell line capable of conditional induction of the NPAS2:BMAL1 heterodimer and identified putative target genes by representational difference analysis, DNA microarrays, and Northern blotting. Coinduction of NPAS2 and BMAL1 activated transcription of the endogenous Per1, Per2, and Cry1 genes, which encode negatively activating components of the circadian regulatory apparatus, and repressed transcription of the endogenous BMAL1 gene. Analysis of the frontal cortex of wild-type mice kept in a 24-hour light-dark cycle revealed that Per1, Per2, and Cry1 mRNA levels were elevated during darkness and reduced during light, whereas BMAL1 mRNA displayed the opposite pattern. In situ hybridization assays of mice kept in constant darkness revealed that Per2 mRNA abundance did not oscillate as a function of the circadian cycle in NPAS2-deficient mice. Thus, NPAS2 likely functions as part of a molecular clock operative in the mammalian forebrain.

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Mouse genomic sequences containing the 5′ untranslated sequences of BMAL1 mRNA were identified by BLAST (www.ncbi.nlm.nih.gov/BLAST) search. Then 2.8 kb of 5′-flanking sequences of the BMAL 5′ UTR were amplified by PCR (with the primers 5′-GCGGCGGTACCAATGAGGTTCCACCCACAC-3′ and 5′-GCGGCTCGAGGACGGACGCCCGAGACGG-3′) and cloned into pGEMT and subsequently into the Xho I site of the pGL3-basic vector.
Supplementary data are available at Science Online (www.sciencemag.org/cgi/content/full/1060699/DC1).

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Authors: Reick, M., Garcia, J. A., Dudley, C., & McKnight, S. L.
Affiliations: The University of Texas Southwestern Medical Center (US)
Department of Biochemistry,
Corresponding author: McKnight, Steven L.
ORCID: Reick, Martin
Garcia, Joseph A.
McKnight, Steven L.
Venue: Science
Publisher: American Association for the Advancement of Science (AAAS)
Published: 2001-07-20
Volume: 293
Issue: 5529
Pages: 506-509
ISSN: 0036-8075 (ISSN-L)
1095-9203
DOI: 10.1126/science.1060699
OpenAlex: W1965359283
Cited by: 572
References: 26
Citations by year: 2026: 10 · 2025: 20 · 2024: 12 · 2023: 12 · 2022: 17 · 2021: 15 · 2020: 14 · 2019: 13 · 2018: 18 · 2017: 15 · 2016: 20 · 2015: 29 · 2014: 25 · 2013: 28 · 2012: 24
FWCI: 9.74
Type: journal article
Language: en
Open access: closed
Locations: Science (publishedVersion)
PubMed (publishedVersion)
Funders: National Institute of Mental Health (R01MH059388, 1RO1MH59388)
Topics: Life Sciences › Neuroscience › Endocrine and Autonomic Systems › Circadian rhythm and melatonin
Life Sciences › Neuroscience › Cognitive Neuroscience › Sleep and Wakefulness Research
Life Sciences › Agricultural and Biological Sciences › Plant Science › Plant Molecular Biology Research
Keywords: NPAS2, clocks, Bmal1, circadian rhythm, transcription factor heterodimer, Per1, Per2, CRY1, clock genes, light-dark cycle, constant darkness, frontal cortex, gene expression oscillations
MeSH terms: Amino Acid Sequence, Animals, Biological Clocks, Cell Line, Circadian Rhythm, Cloning, Molecular, Darkness, Ecdysterone, Eye Proteins, Flavoproteins, Gene Expression Regulation, Humans, Light, Mice, Inbred Strains, Molecular Sequence Data, Nerve Tissue Proteins, Nuclear Proteins, Transcription Factors, Transfection, Tumor Cells, Cultured, Blotting, Northern, Trans-Activators, Prosencephalon, In Situ Hybridization, Photoreceptor Cells, Invertebrate, Cell Cycle Proteins, Dimerization, Oligonucleotide Array Sequence Analysis, Drosophila Proteins, Receptors, G-Protein-Coupled, Mice, Basic Helix-Loop-Helix Transcription Factors, Basic Helix-Loop-Helix Proteins, CLOCK Proteins, ARNTL Transcription Factors, Cryptochromes, Period Circadian Proteins

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NPAS2: An Analog of Clock Operative in the Mammalian Forebrain

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