Pericentromeric tandem DNA transcription in malignant cells and tumour microenvironment in mice NSLC model

In higher primates, only about 1-5% of DNA is made up of protein-coding genes; the other 99% is noncoding. Most of the genome consists of repetitive DNA elements, dispersed and tandem repeats. Tandem repeats (TR, satellite DNA) are located mainly in the centromere and pericentromeric regions of the chromosomes and are the DNA component of constitutive heterochromatin. In some tumours, the TR DNA decondenses and goes into a transcriptionally active state. However, it is not clear yet, what are the cells of the tumour (the stroma, malignant cells) where transcription occurs. The aim of the work was to study the transcription of TR DNA in K-rasG12D –induced carcinogenesis in mouse lung. Methods. Cancer cells and cancer-associated macrophages and fibroblasts were obtained from non-small-cell lung carcinoma (NSLC) of K-rasG12D mice. Transcription of mouse pericentromeric TR DNA (major satellite, MaSat) was analysed by qPCR, RNA-DNA FISH, immunohistochemistry methods. Results. The level of long non-coding MaSat (lncRNA) RNA in a tumour was lower than in the adjacent tissue and normal lung tissue of a healthy mouse. In the culture of NSLC epithelial tumour cells, the content of MaSat lncRNA was lower than in cancer-associated mouse fibroblasts (CAF). An increase in MaSat transcription in NSLC tumour cells occured only when apoptosis was induced by a cytotoxic drugs cisplatin, or after heat exposure. In the tumour microenvironment, the transcription was increased only in CAF but not in cancer-associated macrophages. The increase of MaSat transcription was shown both in histological specimens and in cell cultures, where CAF phenotype was induced either by co-cultivation with cancer cells or by TGFb treatment. The MaSat transcription was acoompanied with the appearance of markers of a CAF phenotype, including markers of cellular senescence. Transcripts were detected only in the nucleus, near chromocenters. No signals were detected in cytoplasm. Pre-treatment with RNAase resulted in the disappearance of signals. Conclusion. Thus, the transcription of pericentromeric satellite DNA observed in NSLC in mice occured mainly in CAF but not associated macrophages or malignant cells. Treatment with cytotoxic drugs induced appearance of CAF markers in fibroblasts . Thus, treatment with cytostatics does not induce death of CAF but promote fibroblasts transition into CAF. Aknowledgements. The work was supported by grants from the Grants’ Council of the President of RF (MK-6706.