Abstract
The circadian clock regulates daily variations in physiologic processes. CLOCK acts as a regulator in the circadian apparatus controlling the expression of other clock genes, including PER1. Clock genes have been implicated in cancer-related functions; in this work, we investigated CLOCK as a possible target of somatic mutations in microsatellite unstable colorectal cancers. Combining microarray gene expression data and public gene sequence information, we identified CLOCK as 1 of 790 putative novel microsatellite instability (MSI) target genes. A total of 101 MSI colorectal carcinomas (CRC) were sequenced for a coding microsatellite in CLOCK. The effect of restoring CLOCK expression was studied in LS180 cells lacking wild-type CLOCK by stably expressing GST-CLOCK or glutathione S-transferase empty vector and testing the effects of UV-induced apoptosis and radiation by DNA content analysis using flow cytometry. Putative novel CLOCK target genes were searched by using ChIP-seq. CLOCK mutations occurred in 53% of MSI CRCs. Restoring CLOCK expression in cells with biallelic CLOCK inactivation resulted in protection against UV-induced apoptosis and decreased G(2)-M arrest in response to ionizing radiation. Using ChIP-Seq, novel CLOCK-binding elements were identified near DNA damage genes p21, NBR1, BRCA1, and RAD50. CLOCK is shown to be mutated in cancer, and altered response to DNA damage provides one plausible mechanism of tumorigenesis.
Highlights
Circadian rhythms are daily oscillations in physiologic processes
Gene expression profiling Combining microarray gene expression data [15] and public gene sequence information, we identified CLOCK as 1 of 790 genes with reduced expression in microsatellite instability (MSI) CRCs compared with normal colon samples, and as having a mononucleotide microsatellite in the coding region
To examine whether mutations in CLOCK were involved in colorectal tumorigenesis, a total of 101 MSI CRCs were analyzed for a coding microsatellite of nine thymines (T9) in CLOCK exon 8
Summary
Circadian rhythms are daily oscillations in physiologic processes. They regulate various functions in the human body, including sleep, body temperature, hormone production, digestive secretion, and immune activity. The rhythms are controlled by an endogenous clock. At the Authors' Affiliations: 1Genome-Scale Biology Research Program and Department of Medical Genetics, University of Helsinki; 2Department of Clinical Genetics, Helsinki University Central Hospital; 3Genome-Scale Biology Research Program and Institute of Biomedicine, University of Helsinki, and Department of Molecular Medicine, National Public Health Institute; 4Department of Computer Science, University of Helsinki; 5Biomedicum Bioinformatics Unit, University of Helsinki; 6Computational Systems Biology Laboratory, Institute of Biomedicine and Genome-Scale Biology Program, University of Helsinki; 7The Second Department of Surgery, Helsinki University Central Hospital, Helsinki, Finland; 8Departments of Surgery and Pathology, Jyväskylä Central Hospital, Jyväskylä, Finland; 9Department of Clinical Biochemistry, Aarhus University Hospital Skejby, Aarhus, Denmark; and 10Group of Molecular Oncology, Molecular Biology and Biochemistry Research Center (CIBBIM), Nanomedicine Program, Vall d'Hebron Hospital, and CIBER de Bioingeniería, Biomateriales y Nanomedicina, Barcelona, Spain.
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