Molecular Cloning of the Human Platelet-Derived Growth Factor Receptor β (PDGFR-β) Promoter and Drug Targeting of the G-Quadruplex-Forming Region To Repress PDGFR-β Expression

详细信息    查看全文

• 作者：Yong Qin ; Jessica S. Fortin ; Denise Tye ; Mary Gleason-Guzman ; Tracy A. Brooks ; Laurence H. Hurley
• 刊名：Biochemistry
• 出版年：2010
• 出版时间：May 18, 2010
• 年：2010
• 卷：49
• 期：19
• 页码：4208-4219
• 全文大小：1204K
• 年卷期：v.49,no.19(May 18, 2010)
• ISSN：1520-4995

文摘

To understand the mechanisms controlling platelet-derived growth factor receptor β (PDGFR-β) expression in malignancies, we have cloned and characterized the first functional promoter of the human PDGFR-β gene, which has been confirmed by luciferase reporter gene assays. The transcription initiation sites were mapped by primer extension. Promoter deletion experiments demonstrate that the proximal, highly GC-rich region (positions −165 to −139) of the human PDGFR-β promoter is crucial for basal promoter activity. This region is sensitive to S1 nuclease and likely to assume a non-B-form DNA secondary structure within the supercoiled plasmid. The G-rich strand in this region contains a series of runs of three or more guanines that can form multiple different G-quadruplex structures, which have been subsequently assessed by circular dichroism. A Taq polymerase stop assay has shown that three different G-quadruplex-interactive drugs can each selectively stabilize different G-quadruplex structures of the human PDGFR-β promoter. However, in transfection experiments, only telomestatin significantly reduced the human PDGFR-β basal promoter activity relative to the control. Furthermore, the PDGFR-β mRNA level in Daoy cells was significantly decreased after treatment with 1 μM telomestatin for 24 h. Therefore, we propose that ligand-mediated stabilization of specific G-quadruplex structures in the human PDGFR-β promoter can modulate its transcription.