Genomic response to Wnt signalling is highly context-dependent--evidence from DNA microarray and chromatin immunoprecipitation screens of Wnt/TCF targets.
Wnt proteins are important regulators of embryonic development, and dysregulated Wnt signalling is involved in the oncogenesis of several human cancers. Our knowledge of the downstream target genes is limited, however. We used a chromatin immunoprecipitation-based assay to isolate and characterize the actual gene segments through which Wnt-activatable transcription factors, TCFs, regulate transcription and an Affymetrix microarray analysis to study the global transcriptional response to the Wnt3a ligand. The anti-beta-catenin immunoprecipitation of DNA-protein complexes from mouse NIH3T3 fibroblasts expressing a fusion protein of beta-catenin and TCF7 resulted in the identification of 92 genes as putative TCF targets. GeneChip assays of gene expression performed on NIH3T3 cells and the rat pheochromocytoma cell line PC12 revealed 355 genes in NIH3T3 and 129 genes in the PC12 cells with marked changes in expression after Wnt3a stimulus. Only 2 Wnt-regulated genes were shared by both cell lines. Surprisingly, Disabled-2 was the only gene identified by the chromatin immunoprecipitation approach that displayed a marked change in expression in the GeneChip assay. Taken together, our approaches give an insight into the complex context-dependent nature of Wnt pathway transcriptional responses and identify Disabled-2 as a potential new direct target for Wnt signalling.
Railo
Antti
A
Oulu Centre for Cell Matrix Research, Biocenter Oulu, Laboratory of Developmental Biology and Department of Medical Biochemistry and Molecular Biology, FIN-90014, University of Oulu, P. O. Box 5000, Finland.
Pajunen
Antti
A
Itäranta
Petri
P
Naillat
Florence
F
Vuoristo
Jussi
J
Kilpeläinen
Pekka
P
Vainio
Seppo
S
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport / Q000235:genetics / Q000378:metabolism; D000818:Animals; D051017:Apoptosis Regulatory Proteins; D047369:Chromatin Immunoprecipitation; D004622:Embryo, Mammalian / Q000033:anatomy & histology / Q000502:physiology; D018507:Gene Expression Regulation, Developmental; D051538:Hepatocyte Nuclear Factor 1-alpha; D006801:Humans; D017403:In Situ Hybridization; D051379:Mice; D008969:Molecular Sequence Data; D041681:NIH 3T3 Cells; D020411:Oligonucleotide Array Sequence Analysis; D016716:PC12 Cells; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology*; D051822:T Cell Transcription Factor 1 / Q000235:genetics / Q000378:metabolism*; D051153:Wnt Proteins / Q000235:genetics / Q000378:metabolism*; D060508:Wnt3 Protein; D060509:Wnt3A Protein; D051176:beta Catenin / Q000235:genetics / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; D051017:Apoptosis Regulatory Proteins; C478773:Dab2 protein, mouse; D051538:Hepatocyte Nuclear Factor 1-alpha; C496488:Hnf1a protein, mouse; D011993:Recombinant Fusion Proteins; D051822:T Cell Transcription Factor 1; C555510:WNT3A protein, human; D051153:Wnt Proteins; D060508:Wnt3 Protein; D060509:Wnt3A Protein; C555511:Wnt3a protein, mouse; D051176:beta Catenin