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Molecular and Cellular Biology, October 2007, p. 7220-7235, Vol. 27, No. 20
0270-7306/07/$08.00+0     doi:10.1128/MCB.00274-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Functional Analysis of the Transcription Repressor PLU-1/JARID1B ^,

Breast Cancer Biology Group, King's College London School of Medicine, 3rd Floor, Thomas Guy House, Guy's Hospital, London, United Kingdom,¹ Cancer Research UK, Medical Oncology Unit, Barts and the Royal London School of Medicine and Dentistry, London, United Kingdom,² Division of Cellular and Molecular Medicine, St. Georges Hospital, London, United Kingdom³

Received 15 February 2007/ Returned for modification 9 April 2007/ Accepted 2 August 2007

The PLU-1/JARID1B nuclear protein, which is upregulated in breast cancers, belongs to the ARID family of DNA binding proteins and has strong transcriptional repression activity. To identify the target genes regulated by PLU-1/JARID1B, we overexpressed or silenced the human PLU-1/JARID1B gene in human mammary epithelial cells by using adenovirus and RNA interference systems, respectively, and then applied microarray analysis to identify candidate genes. A total of 100 genes showed inversely correlated differential expression in the two systems. Most of the candidate genes were downregulated by the overexpression of PLU-1/JARID1B, including the MT genes, the tumor suppressor gene BRCA1, and genes involved in the regulation of the M phase of the mitotic cell cycle. Chromatin immunoprecipitation assays confirmed that the metallothionein 1H (MT1H), -1F, and -1X genes are direct transcriptional targets of PLU-1/JARID1B in vivo. Furthermore, the level of trimethyl H3K4 of the MT1H promoter was increased following silencing of PLU-1/JARID1B. Both the PLU-1/JARID1B protein and the ARID domain selectively bound CG-rich DNA. The GCACA/C motif, which is abundant in metallothionein promoters, was identified as a consensus binding sequence of the PLU-1/JARID1B ARID domain. As expected from the microarray data, cells overexpressing PLU-1/JARID1B have an impaired G₂/M checkpoint. Our study provides insight into the molecular function of the breast cancer-associated transcriptional repressor PLU-1/JARID1B.

Published ahead of print on 20 August 2007.

Supplemental material for this article may be found at http://mcb.asm.org/.

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