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Processing of three different types of DNA damage in cell lines of a cutaneous squamous cell carcinoma progression
1Department of Dermatology, University of Würzburg, Germany.
Abstract:
In order to study the role of DNA damage processing in the development of cutaneous squamous cell carcinoma (SCC), we assessed the ability of six keratinocyte cell lines from a multistage-tumor progression model to repair three types of DNA damage: pyrimidine dimers, oxidative DNA lesions and DNA double strand breaks (DSB). The model comprised the spontaneously immortalized, non-tumorigenic human keratinocyte cell line HaCaT, four different c-Ha-ras transfectants of HaCaT (non-, benign- and two malignant-tumorigenic) and a SCC-derived cell line. Host cell reactivation assays with UVB-treated plasmid vectors pRSVcat showed no significantly altered repair of UVB-induced pyrimidine dimers in the tumorigenic cell lines, compared with the non-tumorigenic lines. Using the singlet oxygen-treated plasmids pRSVcat the Ha-ras-HaCaT-clones and the SCC-cells, exerted a DNA repair efficiency that was not significantly different from HaCaT cells. In order to assess the ability of the cells to ligate free DNA ends (repair of DSB), we used a plasmid shuttle vector assay with linearized plasmid pZ189. We found a significant increase of DNA end joining ability in the non-tumorigenic, the benign and in one of the malignant HaCaT-clones II-4. The malignant HaCaT-clone II-3, however, exerted a significantly lower rate of rejoining the linearized plasmid. This cell line also showed a highly and significantly elevated rate of micronuclei, which reflects a pronounced chromosomal instability. The SCC-cells exhibited a more efficient repair of DNA DSB than the HaCaT cells. We conclude that in the examined model, progression of human keratinocytes from the non-tumorigenic to the highly tumorigenic phenotype, is not accompanied by a decrease in the cell's capacity to repair UVB- and singlet oxygen-induced DNA lesions. However, an acquired deficiency in repairing DNA double strand breaks can be one mechanism promoting progression towards malignancy, possibly through impairing chromosomal stability.
Insights
DNA repair capacity for pyrimidine dimers and oxidative lesions remains stable during skin cancer progression. However, impaired DNA double-strand break repair may drive malignancy by increasing chromosomal instability.
Area of Science:
- Molecular Biology
- Dermatology
- Cancer Research
Background:
- Cutaneous squamous cell carcinoma (SCC) development involves genetic alterations.
- DNA damage processing is crucial for maintaining genomic integrity.
- Understanding DNA repair mechanisms in keratinocytes is vital for SCC research.
Purpose of the Study:
- To investigate the role of DNA damage repair in the multistage development of cutaneous SCC.
- To compare the repair efficiency of different DNA lesions in a human keratinocyte model.
- To correlate DNA repair capacity with tumor progression and chromosomal instability.
Main Methods:
- Utilized a multistage SCC progression model using HaCaT keratinocyte cell lines and SCC-derived cells.
- Assessed repair of pyrimidine dimers (UVB-induced) and oxidative DNA lesions using host cell reactivation assays.
- Evaluated DNA double-strand break (DSB) repair capacity via a plasmid shuttle vector assay measuring DNA end joining.
Main Results:
- No significant difference in pyrimidine dimer or oxidative lesion repair between non-tumorigenic and tumorigenic keratinocytes.
- Variable DNA double-strand break (DSB) repair efficiency observed; one malignant clone showed decreased repair and increased micronuclei (chromosomal instability).
- SCC-derived cells demonstrated more efficient DSB repair compared to the parental HaCaT cell line.
Conclusions:
- Keratinocyte progression to tumorigenicity is not linked to reduced repair of UVB-induced or oxidative DNA damage.
- Deficiency in DNA double-strand break (DSB) repair can be a mechanism promoting SCC malignancy, potentially via chromosomal instability.
- Specific defects in DSB repair pathways may contribute to the development of cutaneous squamous cell carcinoma.