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D-loop somatic mutations and ∼5 kb “common” deletion in mitochondrial DNA: important molecular markers to distinguish oral precancer and cancer

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Tumor Biology

Abstract

Apart from genomic DNA, mutations at mitochondrial DNA (mtDNA) have been hypothesized to play vital roles in cancer development. In this study, ∼5 kb deletion and D-loop mutations in mtDNA and alteration in mtDNA content were investigated in buccal smears from 104 healthy controls and 74 leukoplakia and 117 cancer tissue samples using Taqman-based quantitative assay and re-sequencing. The ∼5 kb deletion in mtDNA was significantly less (9.8 and 10.5 folds, P < 0.0001) in cancer tissues compared to control and leukoplakia tissues, respectively. On the other hand, somatic mutations in D-loop, investigated in 54 controls, 50 leukoplakias and 56 cancer patients, were found to be significantly more in cancer tissues, but not in leukoplakia tissues, compared to control (Z-score = 5.4). MtDNA contents were observed to be significantly more in leukoplakia (2.1 folds, P = 0.004) and cancer (1.6 folds, P = 0.03) tissues compared to control tissues. So, D-loop somatic mutations and ∼5 kb deletion patterns could be used as distinguishing markers between precancer and cancer tissues. This observation further suggests that somatic mutations in D-loop may facilitate carcinogenesis and cancer cells with less ∼5 kb deletion, i.e., intact mtDNA, may become resistant to apoptosis.

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Abbreviations

HVS:

Hypervariable sequence

np:

Nucleotide position

mtDNA:

Mitochondrial DNA

D-loop:

Displacement loop

ROS:

Reactive oxygen species

OXPHOS:

Oxidative phosphorylation

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Acknowledgments

Authors are grateful for the cooperation received from all patients and healthy individuals. We acknowledge the technical help from Mr. Badal Dey. This work was supported by Indian Statistical Institute.

Conflict of interest

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Author information

Author notes

  1. Sayantan Datta

    Present address: Department of Otolaryngology, John Hopkins School of Medicine, Baltimore, MD, USA

  2. Mousumi Majumder

    Present address: Schulich School of Medicine and Dentistry, University of Western Ontario Medical School, London, ON, N6A5C1, Canada

  3. Puspita Das Roy

    Present address: Department of Biochemistry, University of Calcutta, 35 Ballygunge Circular Road, Kolkata, 700019, India

Authors and Affiliations

  1. Human Genetics Unit, Indian Statistical Institute, 203 B.T. Road, Kolkata, 700108, India

    Sayantan Datta, Esita Chattopadhyay, Mousumi Majumder, Puspita Das Roy & Bidyut Roy

  2. Department of Oral Pathology, Dr. R Ahmed Dental College and Hospital, 114, AJC Bose Road, Kolkata, 700014, India

    Jay Gopal Ray

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  1. Sayantan Datta
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Corresponding author

Correspondence to Bidyut Roy.

Electronic supplementary material

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Supplementary Figure 1

RNaseP is a single copy nuclear gene that was used as nuclear DNA standard. ND1 is a rarely deleted region of the mitochondrion that was used to estimate mitochondrial DNA content. XDEL, DNA sequences from the flanks of 4977 bp deletion region, was used for to amplify specifically mtDNA having 4977 bp deletion. Though several other deletions are possible in mtDNA (marked by thin colored lines), XDEL is specific to 4977 bp deletion (marked by thick red line). Black () arrow indicates arbitrary positions of forward and reverse primers used for amplification of RNaseP, ND1and XDEL for Taqman assay. (GIF 41 kb)

High Resolution Image (TIFF 3781 kb)

Supplementary Figure 2

∆Ct (XDEL-ND1) values were drawn against ages (yrs) of patients and controls. Blue dots (), red dots () and green dots () were values for cancer, leukoplakia and control samples, respectively. Regression lines (blue, red and green) were drawn to check the R2 and P values. (GIF 38 kb)

High Resolution Image (TIFF 3777 kb)

Supplementary Figure 3

Number of mutations in D-loop of the cancer patients was drawn against the age (yrs) of them. Regression line shows the value of R2 = 0.0047 and p value = 0.78 (GIF 27 kb)

High Resolution Image (TIFF 2782 kb)

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Datta, S., Chattopadhyay, E., Ray, J.G. et al. D-loop somatic mutations and ∼5 kb “common” deletion in mitochondrial DNA: important molecular markers to distinguish oral precancer and cancer. Tumor Biol. 36, 3025–3033 (2015). https://doi.org/10.1007/s13277-014-2937-2

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  • DOI: https://doi.org/10.1007/s13277-014-2937-2

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