Abstract
Different phenotypes of normal cells might influence genetic profiles, epigenetic profiles, and tumorigenicities of their transformed derivatives. In this study, we investigate whether the whole mitochondrial genome of immortalized cells can be attributed to the different phenotypes (stem vs. non-stem) of their normal epithelial cell originators. To accurately determine mutations, we employed Duplex Sequencing, which exhibits the lowest error rates among currently-available DNA sequencing methods. Our results indicate that the vast majority of the observed mutations of the whole mitochondrial DNA occur at low-frequency (rare mutations). The most prevalent rare mutation types are C→T/G→A and A→G/T→C transitions. Frequencies and spectra of homoplasmic point mutations are virtually identical between stem cell-derived immortalized (SV1) cells and non-stem cell-derived immortalized (SV22) cells, verifying that both cell types were derived from the same woman. However, frequencies of rare point mutations are significantly lower in SV1 cells (5.79 × 10−5) than in SV22 cells (1.16 × 10−4). The significantly lower frequencies of rare mutations are aligned with a finding of longer average distances to adjacent mutations in SV1 cells than in SV22 cells. Additionally, the predicted pathogenicity for rare mutations in the mitochondrial tRNA genes tends to be lower (by 2.5-fold) in SV1 cells than in SV22 cells. While four known/confirmed pathogenic mt-tRNA mutations (m.5650 G>A, m.5521 G>A, m.5690 A>G, m.1630 A>G) were identified in SV22 cells, no such mutations were observed in SV1 cells. Our findings suggest that the immortalization of normal cells with stem cell features leads to decreased mitochondrial mutagenesis, particularly in RNA gene regions. The mutation spectra and mutations specific to stem cell-derived immortalized cells (vs. non-stem cell derived) have implications in characterizing the heterogeneity of tumors and understanding the role of mitochondrial mutations in the immortalization and transformation of human cells.
Highlights
Evidence exists that distinct phenotypes of normal cell [1,2,3,4,5] or precancerous cell [6] originators for tumor derivatives lead to tumor heterogeneities
Non-stem cell-derived immortalized human breast epithelial cells (HBECs) will be referred to as SV22, and stem cell-derived immortalized HBECs will be referred to as SV1 cells. Both SV22 and SV1 cells were cultured under the same conditions, DNA was extracted, and DNA libraries were prepared for Duplex Sequencing as we have described previously [16]
Gordon et al (2014) demonstrated that immortalization of human fetal lung fibroblasts increased DNA methylation at gene promoters and caused large-scale changes in gene expression. While these studies [4,37] have examined tumorigenicity, histopathology, and metastatic behavior of human breast transformed cells or DNA methylation and gene expression of immortalized human fibroblasts, no study has examined the subclonal mutations of the whole mitochondrial genome for immortalized derivatives of phenotypically different normal cells
Summary
Evidence exists that distinct phenotypes of normal cell [1,2,3,4,5] or precancerous cell [6] originators for tumor derivatives lead to tumor heterogeneities. To study consequences of normal cell origins on genetic changes in their immortalized derivatives, we examined two different immortalized (pre-neoplastic) breast epithelial cell types that were derived from normal human breast epithelial cells (HBECs) with different phenotypes (Types I and II). Both Type-I and Type-II normal HBECs were isolated from breast tissue of the same woman; they exhibited different phenotypes. Type-II HBECs exhibit opposite phenotypes in the described features above (i.e., do not express stem cell markers; do not express ER-α; express basal epithelial markers; express GJIC proteins; higher expression of maspin)
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
