A systematic approach identifies p53-DREAM target genes associated with blood or brain abnormalities
Abstract
p53 is mainly known as a tumor suppressor, but mouse models revealed that increased p53 activity may cause bone marrow failure, through mechanisms that likely include gene repression mediated by the p53-DREAM pathway. Here we designed a systematic approach to identify p53-DREAM targets whose repression might contribute to abnormal hematopoiesis. We used gene ontology to analyze transcriptomic changes associated with bone marrow cell differentiation and p53 activation, then ChIP-seq data to find promoters bound by the DREAM complex. We next created positional frequency matrices to identify evolutionary conserved sequence elements potentially bound by DREAM. The same approach was developed to find p53-DREAM targets associated with brain abnormalities, also observed in mice with increased p53 activity. Putative DREAM binding sites were found for 151 candidate p53-DREAM target genes, of which 106 are mutated in a blood or brain genetic disorder. Twenty-one DREAM binding sites were tested and found to impact on gene expression in luciferase reporter assays, notably regulating genes mutated in dyskeratosis congenita (Rtel1), Fanconi anemia (Fanca), Diamond-Blackfan anemia (Tsr2), primary microcephaly (Casc5,Ncaph,Wdr62) or pontocerebellar hypoplasia (Toe1). These results provide clues on the role of the p53-DREAM pathway in regulating hematopoiesis and brain development, with implications for tumorigenesis.
Author Summary
The capacity of p53 to activate the transcription of genes important for cell cycle arrest, apoptosis or cellular metabolism has been recognized for decades. By contrast, the potential importance of p53-dependent transcriptional repression emerged more recently. Although p53 frequently appears to repress genes indirectly via the DREAM repressor complex, only a few studies attempted to define p53-DREAM target gene repertoires, often by analyzing cell cycle regulation in fibroblasts. Here we aimed to gain a better appreciation of the clinical relevance of the p53-DREAM pathway by designing a systematic approach for the identification of p53-DREAM targets. Because mouse models with increased p53 activity suffer from bone marrow failure or brain hypoplasia, we relied on transcriptomic changes associated with bone marrow cell differentiation, blood- and brain-related gene ontology terms and RNAseq data from hematopoietic or neural progenitor cells to identify p53-DREAM targets, then created positional frequency matrices to find putative DREAM binding sites. Our study provides a resource of predicted DREAM binding sites for 151 genes associated with blood and/or brain abnormalities, many of which were not previously known to be DREAM targets. Furthermore, our analysis suggests that p53-DREAM alterations may contribute to phenotypic variations in glioblastoma cells.
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