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Cyclin D1 gene contains a cryptic promoter that is functional in human cancer cells
A Terrinoni1, R Dell'Arciprete, M Fornaro
1Laboratory of Experimental Oncology, Department of Cell Biology and Oncology, Istituto Mario Negri-Consorzio Mario Negri Sud, Santa Maria Imbaro (Chieti), Italy.
Abstract:
A novel cyclin D1 (CCND1)-TROP2 fusion oncogene has been isolated from human cancer cells. Unexpectedly, the chimeric cDNA was found to express TROP2 in the absence of exogenous promoters. Mutagenesis of the TROP2 and CCND1 sequences and in vitro transcription/translation show that a cryptic promoter is present in the 3' coding region of CCND1. The CCND1 cryptic promoter is functional in luciferase assays, where it augments the basal expression levels by eightfold and efficiently cooperates with an SV-40 enhancer. The transcription start sites of the cryptic promoter map at bases 797 and 935 of CCND1, as determined by RNase protection assays. The cryptic promoter possesses canonical binding sites for ubiquitous transcription factors and W/S, X1, and CAAT/Y boxes that are characteristic of major histocompatibility complex class II gene promoters. Remarkably, the cryptic CCND1 promoter is active in human cancer cells and generates a truncated transcript that contains CCND1 instability sequences. Thus, this novel CCND1 transcription unit may play a role in the regulation of the expression of cyclin D1 and in tumor cell growth.
Insights
Researchers discovered a new cyclin D1 (CCND1) cryptic promoter within human cancer cells. This promoter drives TROP2 expression in a novel CCND1-TROP2 fusion oncogene, potentially regulating tumor cell growth.
Area of Science:
- Molecular Oncology
- Gene Regulation
- Cancer Genomics
Background:
- A novel cyclin D1 (CCND1)-TROP2 fusion oncogene was identified in human cancer cells.
- The fusion unexpectedly expressed TROP2 without external promoters, suggesting an internal regulatory mechanism.
Purpose of the Study:
- To investigate the mechanism behind TROP2 expression in the CCND1-TROP2 fusion.
- To identify and characterize the cryptic promoter within the CCND1 gene.
Main Methods:
- Site-directed mutagenesis of CCND1 and TROP2 sequences.
- In vitro transcription/translation assays.
- Luciferase reporter assays with SV-40 enhancer.
- RNase protection assays to map transcription start sites.
Main Results:
- A cryptic promoter was identified in the 3' coding region of CCND1.
- This CCND1 cryptic promoter enhanced basal expression eightfold in luciferase assays and cooperated with an SV-40 enhancer.
- Transcription start sites were mapped to CCND1 bases 797 and 935.
- The promoter contains binding sites for transcription factors and elements similar to MHC class II gene promoters.
- The promoter is active in cancer cells, producing a truncated transcript with CCND1 instability sequences.
Conclusions:
- A novel, functional CCND1 cryptic promoter exists within the CCND1 gene.
- This promoter is active in human cancer cells and may contribute to oncogene expression and tumor progression.
- The CCND1 cryptic promoter represents a new regulatory element in cancer biology.