H. Jin1,2, M. Roy1, A. Dammalapati1, A. Harrispn1, A. Ma1, R. Jaskula-Sztul1,2, H. Chen1,2 1University Of Alabama,Surgery,Birmingham, AL, USA 2University Of Alabama,Surgery,Birmingham, Alabama, USA 3University Of Wisconsin,Surgery,Madison, WI, USA
Introduction: ~~It is known that Notch signaling is minimally active in neuroendocrine (NE) cancer cells and the induction of Notch isoforms alter the malignant neuroendocrine phenotype. Although the induc¬tion of Notch1 by Histone Deacetylase Inhibitors (HDACi) appeared to be the result of increased Notch1 expression at the transcriptional level, the effects of HDACi on the Notch1 promoter regulation have not been determined thus far. The aim of our study is to investigate the molecular mechanism of HDACi activation on the Notch1 pathway.
Methods: ~~We functionally characterized the Notch1 promoter using deletion mapping. The mapping started with the truncated genomic DNA fragment fused with a luciferase reporter, transfected into BON cell, a carcinoid cell line, screened for luciferase activity. Protein-DNA binding was then performed by electrophoretic mobility shift assay (EMSA).
Results:~~Two HDACi, Valproid Acid and Tailandepsin-A, were shown to induce luciferase activity controlled by a small distal region of Notch1 promoter, from -80 to +1 of the start codon ATG. Further, we identified a functional DNA motif that is responsible for HDACi induction located at -75 to -55 of the Notch1 promoter region. Thus, an in vitro assay, EMSA revealed the transcription factor-DNA complex firmed in the flanking sequence.
Conclusion:~~We have identified the DNA motif located in the Notch1 promoter region that is responsive to HDACi. This understanding of how HDACi act on the Notch1 promoter may lead to the development of future novel therapies for neuroendocrine cancers.
