Revisiting a selection of target genes for the hematopoietic transcription factor c-Myb using chromatin immunoprecipitation and c-Myb knockdown.
The transcription factor c-Myb is an important regulator of hematopoiesis required for proper development of most blood cell lineages in vertebrates. An increasing number of target genes for c-Myb are being published, although with little or no overlap between the lists of genes reported. This raises the question of which criteria a bona fide c-Myb-target gene should satisfy. In the present paper, we have analyzed a set of previously reported target genes using chromatin immunoprecipitation (ChIP) and siRNA-mediated knockdown. Among the seven well-studied c-Myb target genes that we analyzed by ChIP, only ADA, c-MYC and MAT2A seemed to be occupied by c-Myb under our experimental settings in the Myb-positive cell lines Jurkat and HL60. After siRNA-mediated knockdown of c-Myb expression, the expression levels of two out of three ChIP positive Myb target genes, ADA and c-MYC, were strongly affected. These results clearly demonstrate the importance of combining different methods for target gene validation and suggest that a combination of ChIP and c-Myb knockdown may represent a powerful approach to identify a core collection of c-Myb target genes.
Berge
Tone
T
Department of Molecular Biosciences, University of Oslo, N-0316 Oslo, Norway.
Matre
Vilborg
V
Brendeford
Elen M
EM
Saether
Thomas
T
Lüscher
Bernhard
B
Gabrielsen
Odd Stokke
OS
D000243:Adenosine Deaminase / Q000235:genetics / Q000378:metabolism; D045744:Cell Line, Tumor; D047369:Chromatin Immunoprecipitation; D005786:Gene Expression Regulation; D020868:Gene Silencing; D006410:Hematopoiesis / Q000235:genetics*; D006801:Humans; D019901:Locus Control Region; D008716:Methionine Adenosyltransferase / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D020598:Proto-Oncogene Proteins c-myb / Q000235:genetics / Q000378:metabolism*; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics / Q000378:metabolism
D020598:Proto-Oncogene Proteins c-myb; D016271:Proto-Oncogene Proteins c-myc; D008716:Methionine Adenosyltransferase; D000243:Adenosine Deaminase