Cyclin D1 is a direct transcriptional target of GATA3 in neuroblastoma tumor cells.
Almost all neuroblastoma tumors express excess levels of Cyclin D1 (CCND1) compared to normal tissues and other tumor types. Only a small percentage of these neuroblastoma tumors have high-level amplification of the Cyclin D1 gene. The other neuroblastoma tumors have equally high Cyclin D1 expression without amplification. Silencing of Cyclin D1 expression was previously found to trigger differentiation of neuroblastoma cells. Overexpression of Cyclin D1 is therefore one of the most frequent mechanisms with a postulated function in neuroblastoma pathogenesis. The cause for the Cyclin D1 overexpression is unknown. Here we show that Cyclin D1 overexpression results from transcriptional upregulation. To identify upstream regulators, we searched in mRNA profiles of neuroblastoma tumor series for transcription factors with expression patterns correlating to Cyclin D1. GATA3 most consistently correlated to Cyclin D1 in four independent data sets. We identified a highly conserved GATA3 binding site 27 bp upstream of the Cyclin D1 transcriptional start. Chromatin immune precipitation confirmed binding of GATA3 to the Cyclin D1 promoter. Overexpression of GATA3 induced Cyclin D1 promoter activity, which decreased after site-directed mutagenesis of the GATA3 binding site in the Cyclin D1 promoter. Silencing of GATA3 resulted in reduced Cyclin D1 promoter activity and reduced Cyclin D1 mRNA and protein levels. Moreover, GATA3 silencing caused differentiation that was similar to that caused by Cyclin D1 inhibition. These finding implicate GATA3 in Cyclin D1 overexpression in neuroblastoma.
Department of Human Genetics, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands. j.j.molenaar@amc.uva.nl
J J
JJ
Molenaar
M E
ME
Ebus
J
J
Koster
E
E
Santo
D
D
Geerts
R
R
Versteeg
H N
HN
Caron
D001665:Binding Sites; D019938:Cyclin D1 / Q000235:genetics*; D050990:GATA3 Transcription Factor / Q000502:physiology*; D020869:Gene Expression Profiling; D006801:Humans; D009447:Neuroblastoma / Q000209:etiology / Q000378:metabolism / Q000473:pathology*; D011401:Promoter Regions, Genetic; D014158:Transcription, Genetic
C530104:CCND1 protein, human; D050990:GATA3 Transcription Factor; C494749:GATA3 protein, human; D019938:Cyclin D1