Identification of human STAT5-dependent gene regulatory elements based on interspecies homology.
STAT5 is a transcription factor essential for hematopoietic physiology. STAT5 functions to transduce signals from cytokines to the nucleus where it regulates gene expression. Although several important transcriptional targets of STAT5 are known, most remain unidentified. To identify novel STAT5 targets, we searched chromosomes 21 and 22 for clusters of STAT5 binding sites contained within regions of interspecies homology. We identified four such regions, including one with tandem STAT5 binding sites in the first intron of the NCAM2 gene. Unlike known STAT5 binding sites, this site is found within a very large intron and resides approximately 200 kb from the first coding exon of NCAM2. We demonstrate that this region confers STAT5-dependent transcriptional activity. We show that STAT5 binds in vivo to the NCAM2 intron in the NKL natural killer cell line and that this binding is induced by cytokines that activate STAT5. Neither STAT1 nor STAT3 bind to this region, despite sharing a consensus binding sequence with STAT5. Activation of STAT4 and STAT5 causes the accumulation of both of these STATs to the NCAM2 regulatory region. Therefore, using an informatics based approach to identify STAT5 targets, we have identified NCAM2 as both a STAT4- and STAT5-regulated gene, and we show that its expression is regulated by cytokines essential for natural killer cell survival and differentiation. This strategy may be an effective way to identify functional binding regions for transcription factors with known cognate binding sites anywhere in the genome.
Nelson
Erik A
EA
Department of Medical Oncology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts 02115, USA.
Walker
Sarah R
SR
Li
Wei
W
Liu
X Shirley
XS
Frank
David A
DA
D001483:Base Sequence; D001665:Binding Sites; D002460:Cell Line; D002891:Chromosomes, Human, Pair 21; D002892:Chromosomes, Human, Pair 22; D005786:Gene Expression Regulation; D006801:Humans; D016898:Interferon-alpha / Q000378:metabolism; D007376:Interleukin-2 / Q000378:metabolism; D007438:Introns; D007694:Killer Cells, Natural / Q000166:cytology / Q000378:metabolism; D008969:Molecular Sequence Data; D000072340:NIMA-Interacting Peptidylprolyl Isomerase; D039842:Neural Cell Adhesion Molecule L1 / Q000235:genetics / Q000378:metabolism; D019006:Neural Cell Adhesion Molecules; D019696:Peptidylprolyl Isomerase; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D012319:RNA Polymerase II / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid; D050794:STAT1 Transcription Factor / Q000235:genetics / Q000378:metabolism; D050796:STAT3 Transcription Factor / Q000235:genetics / Q000378:metabolism; D050799:STAT5 Transcription Factor / Q000235:genetics / Q000378:metabolism*; D012689:Sequence Homology, Nucleic Acid; D014158:Transcription, Genetic; D050880:p300-CBP Transcription Factors / Q000378:metabolism
D016898:Interferon-alpha; D007376:Interleukin-2; C108033:NCAM2 protein, human; D000072340:NIMA-Interacting Peptidylprolyl Isomerase; D039842:Neural Cell Adhesion Molecule L1; D019006:Neural Cell Adhesion Molecules; D020033:Protein Isoforms; D050794:STAT1 Transcription Factor; D050796:STAT3 Transcription Factor; D050799:STAT5 Transcription Factor; D050880:p300-CBP Transcription Factors; D012319:RNA Polymerase II; C000606846:PIN1 protein, human; D019696:Peptidylprolyl Isomerase