Meeting report: mitochondrial DNA and cancer epidemiology.
Explore the source record for details and available documents.
DNA, Mitochondrial↗
Biomedical subjects
Keshav K Singh
Publications and source records attributed to Keshav K Singh.
Proteomic analysis of mitochondria-to-nucleus retrograde response in human cancer.
All tumors examined to date contain mutations in mitochondrial DNA (mtDNA). In addition, depletion of mtDNA is reported in a variety of tumors. Mitochondrial dysfunction resulting from changes in mtDNA invokes mitochondria-to-nucleus retrograde response in human cells. To identify proteins involved in retrograde response and their potential role in tumorigenesis, we carried out a comparative proteomic analysis using a cell line in which the mitochondrial genome was completely depleted (rho(0) cells lacking all mtDNA-encoded protein subunits), a cybrid cell line in which mtDNA was restored, and the parental cell line. Our comparative proteomic approach revealed marked changes in the cellular proteome and led us to identify quantitative changes in expression of several proteins. We found that subunits of complex I and complex III, molecular chaperones, and a protein involved in cell cycle control were downregulated and Inosine 5'-monophosphate dehydrogenase type 2 (IMPDH2) involved in nucleotide biosynthesis was upregulated in rho(0) cells. Our findings demonstrate that the expression of proteins is restored to wild type level by transfer of wild type mitochondria to rho(0) cells, suggesting that these proteins play key roles in retrograde response. To determine a potential role for identified retrograde responsive proteins in tumorigenesis, we analyzed the expression of UQCRC1 gene (encoding ubiquinol cytochrome-c reductase core protein I) in breast and ovarian tumors. We found that (1) UQCRC1 was highly expressed in breast (74%) and ovarian tumors (34%) and (2) the expression positively correlated with cytochrome c-oxidase (COXII) encoded by mtDNA. Our study opens an avenue for identification of retrograde proteins as potential tumor suppressors or oncogenes involved in carcinogenesis.
Adenocarcinoma↗
Mitochondrial aconitase and citrate metabolism in malignant and nonmalignant human prostate tissues.
BACKGROUND: In prostate cancer, normal citrate-producing glandular secretory epithelial cells undergo a metabolic transformation to malignant citrate-oxidizing cells. m-Aconitase is the critical step involved in this altered citrate metabolism that is essential to prostate malignancy. The limiting m-aconitase activity in prostate epithelial cells could be the result of a decreased level of m-aconitase enzyme and/or the inhibition of existing m-aconitase. Earlier studies identified zinc as an inhibitor of m-aconitase activity in prostate cells; and that the depletion of zinc in malignant cells is an important factor in this metabolic transformation. However, a possibility remains that an altered expression and level of m-aconitase enzyme might also be involved in this metabolic transformation. To address this issue, the in situ level of m-aconitase enzyme was determined by immunohistochemical analysis of prostate cancer tissue sections and malignant prostate cell lines. RESULTS: The immunocytochemical procedure successfully identified the presence of m-aconitase localized in the mitochondrial compartment in PC-3, LNCaP, and DU-145 malignant prostate cell lines. The examination of prostate tissue sections from prostate cancer subjects demonstrated that m-aconitase enzyme is present in the glandular epithelium of normal glands, hyperplastic glands, adenocrcinomatous glands, and prostatic intraepithelial neoplastic foci. Quantitative analysis of the relative level of m-aconitase in the glandular epithelium of citrate-producing adenomatous glands versus the citrate-oxidizing adenocarcinomatous glands revealed no significant difference in m-aconitase enzyme levels. This is in contrast to the down-regulation of ZIP1 zinc transporter in the malignant glands versus hyperplastic glands that exists in the same tissue samples. CONCLUSION: The results demonstrate the existence of m-aconitase enzyme in the citrate-producing glandular epithelial cells; so that deficient m-aconitase enzyme is not associated with the limiting m-aconitase activity that prevents citrate oxidation in these cells. The level of m-aconitase is maintained in the malignant cells; so that an altered enzyme level is not associated with the increased m-aconitase activity. Consequently, the elevated zinc level that inhibits m-aconitase enzyme is responsible for the impaired citrate oxidation in normal and hyperplastic prostate glandular epithelial cells. Moreover, the down-regulation of ZIP1 zinc transporter and corresponding depletion of zinc results in the increase in the activity of the existing m-aconitase activity in the malignant prostate cells. The studies now define the mechanism for the metabolic transformation that characterizes the essential transition of normal citrate-producing epithelial cells to malignant citrate-oxidizing cells.
Aconitate Hydratase↗
Saccharomyces cerevisiae polymerase zeta functions in mitochondria.
The MtArg8 reversion assay, which measures point mutation in mtDNA, indicates that in budding yeast Saccharomyces cerevisiae, DNA polymerase zeta and Rev1 proteins participate in the mitochondrial DNA mutagenesis. Supporting this evidence, both polymerase zeta and Rev1p were found to be localized in the mitochondria. This is the first report demonstrating that the DNA polymerase zeta and Rev1 proteins function in the mitochondria.
DNA, Mitochondrial↗
Mitochondria damage checkpoint, aging, and cancer.
There is growing evidence supporting the progressing decline in mitochondrial function with age. Mitochondria are the major site of reactive oxygen species (ROS) production in the cell; therefore it is likely that progressive decline in mitochondrial function is due to the accumulation of oxidative damage with age. Despite this notion, a role for mitochondria in cellular senescence has been largely ignored. Our studies using mitochondrial gene knockout cells (rho(0)) from a variety of tissue types demonstrate that loss of mitochondrial function leads to cell cycle arrest, cellular senescence, and tumorigenic phenotype. In light of these and earlier studies we hypothesize the existence of a mitochondria damage checkpoint (mitocheckpoint) in human cells. Mitocheckpoint permits cells to arrest in the cell cycle in order to repair/restore mitochondrial function to the normal level. Upon overwhelming, persistent, or severe damage to mitochondria, mitocheckpoint machinery may allow cells to undergo senescence. Thus cellular senescence may function as another checkpoint before cells decide to initiate programmed cell death resulting in aging of tissues and organs. Alternatively, mutations occur in the mitochondrial and/or nuclear DNA, resulting in tumorigenesis.
Aged↗
Cross talk between mitochondria and superoxide generating NADPH oxidase in breast and ovarian tumors.
Reactive oxygen species (ROS) signal cascades involved in cell growth, cell death, mitogenesis, angiogenesis and carcinogenesis. ROS are produced as a byproduct of oxidative phosphorylation (OXPHOS) in the mitochondria. It is estimated that 2-4% of the oxygen consumed during OXPHOS is converted to ROS. Besides mitochondria, NADPH-oxidase 1 (Nox1) also generates a significant amount of ROS in the cell. In this paper, we tested the hypothesis that mitochondria control Nox 1 redox signaling and the loss of control of this signaling contributes to tumorigenesis. We analyzed Nox1 expression in a mitochondrial gene knockout (rho(0)) cell line and in the isogenic cybrid cell line in which mitochondrial genes were restored by transfer of wild type mitochondria into rho(0) cells. Our study revealed, for the first time, that the inactivation of mitochondrial genes leads to down-regulation of Nox1 and that the transfer of wild type mitochondrial genes restored the Nox1 expression to a level comparable to that in the parental cell line. Consistent with Nox1 down-regulation, we found that rho(0) cells contained low levels of superoxide anion and that superoxide levels reversed to parental levels in cybrid cells when Nox1 expression was restored by transfer of wild type mitochondria. Increasing mitochondrial superoxide levels also increased the expression of Nox1 in parental cells. Confocal microscopy studies revealed that Nox1 localizes in the mitochondria. Nox1 was highly expressed in breast (86%) and ovarian (71%) tumors and that its expression positively correlated with expression of cytochrome C oxidase encoded by mtDNA. Our study, described in this paper demonstrates the existence of cross talk between the mitochondria and NADPH oxidase. Furthermore, our studies suggest that mitochondria control Nox1 redox signaling and the loss of control of this signaling contributes to breast and ovarian tumorigenesis.
Breast Neoplasms↗
hZIP1 zinc uptake transporter down regulation and zinc depletion in prostate cancer.
BACKGROUND: The genetic and molecular mechanisms responsible for and associated with the development and progression of prostate malignancy are largely unidentified. The peripheral zone is the major region of the human prostate gland where malignancy develops. The normal peripheral zone glandular epithelium has the unique function of accumulating high levels of zinc. In contrast, the ability to accumulate zinc is lost in the malignant cells. The lost ability of the neoplastic epithelial cells to accumulate zinc is a consistent factor in their development of malignancy. Recent studies identified ZIP1 (SLC39A1) as an important zinc transporter involved in zinc accumulation in prostate cells. Therefore, we investigated the possibility that down-regulation of hZIP1 gene expression might be involved in the inability of malignant prostate cells to accumulate zinc. To address this issue, the expression of hZIP1 and the depletion of zinc in malignant versus non-malignant prostate glands of prostate cancer tissue sections were analyzed. hZIP1 expression was also determined in malignant prostate cell lines. RESULTS: hZIP1 gene expression, ZIP1 transporter protein, and cellular zinc were prominent in normal peripheral zone glandular epithelium and in benign hyperplastic glands (also zinc accumulating glands). In contrast, hZIP1 gene expression and transporter protein were markedly down-regulated and zinc was depleted in adenocarcinomatous glands and in prostate intra-epithelial neoplastic foci (PIN). These changes occur early in malignancy and are sustained during its progression in the peripheral zone. hZIP1 is also expressed in the malignant cell lines LNCaP, PC-3, DU-145; and in the nonmalignant cell lines HPr-1 and BPH-1. CONCLUSION: The studies clearly establish that hZIP1 gene expression is down regulated and zinc is depleted in adenocarcinomatous glands. The fact that all the malignant cell lines express hZIP1 indicates that the down-regulation in adenocarcinomatous glands is likely due to in situ gene silencing. These observations, coupled with the numerous and consistent reports of loss of zinc accumulation in malignant cells in prostate cancer, lead to the plausible proposal that down regulation of hZIP1 is a critical early event in the development prostate cancer.
Cation Transport Proteins↗
Inter-genomic cross talk between mitochondria and the nucleus plays an important role in tumorigenesis.
Mitochondrial dysfunction is a hallmark of cancer cells. Consistent with this phenotype mutations in mitochondrial genome have been reported in all cancers examined to date. However, it is not clear whether mitochondrial genomic status in human cells affects nuclear genome stability and whether proteins involved in inter-genomic cross talk are involved in tumorigenesis. Using cell culture model and cybrid cell technology, we provide evidence that mitochondrial genetic status impacts nuclear genome stability in human cells. In particular our studies demonstrate 1) that depletion of mitochondrial genome (rho0) leads to chromosomal instability (CIN) reported to be present in variety of human tumors and 2) rho0 cells show transformed phenotype. Our study also demonstrates that mitochondrial genetic status plays a key role in regulation of a multifunctional protein APE1 (also known as Ref1 or HAP1) involved in transcription and DNA repair in the nucleus and the mitochondria. Interestingly we found that altered expression of APE1 in rho0 cells and tumorigenic phenotype can be reversed by exogenous transfer of wild type mitochondria in rho0 cells. Furthermore, we demonstrate that APE1 expression is altered in variety of primary tumors. Taken together, these studies suggest that inter-genomic cross talk between mitochondria and the nucleus plays an important role in tumorigenesis and that APE1 mediates this process.
Blotting, Western↗
Polymorphisms in genes related to oxidative stress (MPO, MnSOD, CAT) and survival after treatment for breast cancer.
The proximate cause of cancer cell death by radiation therapy and a number of therapeutic agents is through generation of reactive oxygen species, resulting in DNA damage as well as mitochondrial membrane disruption, triggering the apoptotic cascade. Because mitochondrial manganese superoxide dismutase catalyzes conversion of superoxide radicals to H(2)O(2), with catalase neutralizing H(2)O(2) and myeloperoxidase converting H(2)O(2) to highly reactive hypochlorous acid, we hypothesized that gene variants could impact the efficacy of treatment for breast cancer and improve survival. Women who were treated with radiation and/or chemotherapy for incident breast cancer at the Arkansas Cancer Research Center from 1985 to 1996 were identified. DNA was extracted from paraffin-embedded normal tissue (n = 279), and MnSOD, CAT, and MPO genotypes were determined using mass spectrometry. Cox proportional hazards models were adjusted for age, race, stage with node status, and estrogen receptor and progesterone receptor status. Women who were homozygous for MPO G alleles, associated with increased transcription, had better survival (hazard ratio, 0.60; 95% confidence interval, 0.38-0.95; P = 0.03) than those with common alleles. Both CAT TT and MnSOD CC genotypes were associated with nonsignificant reduced hazard of death. When we combined genotypes associated with higher levels of reactive oxygen species for MnSOD and MPO, women with MnSOD CC and MPO GG genotypes had a 3-fold decrease in hazard of death (hazard ratio, 0.33; 95% confidence interval, 0.13-0.80; P = 0.01). These data indicate that gene variants that impact oxidative stress modify prognosis after treatment for breast cancer.
Breast Neoplasms↗
Mitochondrial correlation microscopy and nanolaser spectroscopy - new tools for biophotonic detection of cancer in single cells.
Currently, pathologists rely on labor-intensive microscopic examination of tumor cells using century-old staining methods that can give false readings. Emerging BioMicroNano-technologies have the potential to provide accurate, realtime, high-throughput screening of tumor cells without the need for time-consuming sample preparation. These rapid, nano-optical techniques may play an important role in advancing early detection, diagnosis, and treatment of disease. In this report, we show that laser scanning confocal microscopy can be used to identify a previously unknown property of certain cancer cells that distinguishes them, with single-cell resolution, from closely related normal cells. This property is the correlation of light scattering and the spatial organization of mitochondria. In normal liver cells, mitochondria are highly organized within the cytoplasm and highly scattering, yielding a highly correlated signal. In cancer cells, mitochondria are more chaotically organized and poorly scattering. These differences correlate with important bioenergetic disturbances that are hallmarks of many types of cancer. In addition, we review recent work that exploits the new technology of nanolaser spectroscopy using the biocavity laser to characterize the unique spectral signatures of normal and transformed cells. These optical methods represent powerful new tools that hold promise for detecting cancer at an early stage and may help to limit delays in diagnosis and treatment.
Animals↗
Cellular aging of mitochondrial DNA-depleted cells.
We have reported that mitochondrial DNA-depleted rho(0) cells are resistant to cell death. Because aged cells have frequent mitochondrial DNA mutations, the resistance of rho(0) cells against cell death might be related to the apoptosis resistance of aged cells and frequent development of cancers in aged individuals. We studied if rho(0) cells have features simulating aged cells. SK-Hep1 hepatoma rho(0) cells showed typical morphology associated with aging such as increased size and elongated appearance. They had increased senescence-associated beta-Gal activity, lipofuscin pigment, and plasminogen activator inhibitor-1 expression. Consistent with their decreased proliferation, the expression of mitotic cyclins was decreased and that of cdk inhibitors was increased. Rb hypophosphorylation and decreased telomerase activity were also noted. Features simulating aged cells were also observed in MDA-MB-435 rho(0) cells. These results support the mitochondrial theory of aging, and suggest that rho(0) cells could serve as an in vitro model for aged cells.
Apoptosis↗
Mitochondrial inhibition of uracil-DNA glycosylase is not mutagenic.
BACKGROUND: Uracil DNA glycosylase (UDG) plays a major role in repair of uracil formed due to deamination of cytosine. UDG in human cells is present in both the nucleus and mitochondrial compartments. Although, UDG's role in the nucleus is well established its role in mitochondria is less clear. RESULTS: In order to identify UDG's role in the mitochondria we expressed UGI (uracil glycosylase inhibitor) a natural inhibitor of UDG in the mitochondria. Our studies suggest that inhibition of UDG by UGI in the mitochondria does not lead to either spontaneous or induced mutations in mtDNA. Our studies also suggest that UGI expression has no affect on cellular growth or cytochrome c-oxidase activity. CONCLUSIONS: These results suggest that human cell mitochondria contain alternatives glycosylase (s) that may function as back up DNA repair protein (s) that repair uracil in the mitochondria.
Breast↗
Mitochondrial dysfunction in cancer.
Nearly a century of scientific research has revealed a number of notable differences in the structure and function of mitochondria between normal and cancer cells, including differences in metabolic activity, molecular composition, and mtDNA sequence. This article reviews several of these differences and discusses their clinical implications, especially with regard to the use of mitochondria as biomarkers for early detection of cancer, or as unique cellular targets for novel and selective anti-cancer agents.
Journal Article↗
Genome-wide analysis of signal transducers and regulators of mitochondrial dysfunction in Saccharomyces cerevisiae.
Mitochondrial dysfunction is a hallmark of cancer cells. However, genetic response to mitochondrial dysfunction during carcinogenesis is unknown. To elucidate genetic response to mitochondrial dysfunction we used Saccharomyces cerevisiae as a model system. We analyzed genome-wide expression of nuclear genes involved in signal transduction and transcriptional regulation in a wild-type yeast and a yeast strain lacking the mitochondrial genome (rho(0)). Our analysis revealed that the gene encoding cAMP-dependent protein kinase subunit 3 (PKA3) was upregulated. However, the gene encoding cAMP-dependent protein kinase subunit 2 (PKA2) and the VTC1, PTK2, TFS1, CMK1, and CMK2 genes, involved in signal transduction, were downregulated. Among the known transcriptional factors, OPI1, MIG2, INO2, and ROX1 belonged to the upregulated genes, whereas MSN4, MBR1, ZMS1, ZAP1, TFC3, GAT1, ADR1, CAT8, and YAP4 including RFA1 were downregulated. RFA1 regulates DNA repair genes at the transcriptional level. RFA is also involved directly in DNA recombination, DNA replication, and DNA base excision repair. Downregulation of RFA1 in rho(0) cells is consistent with our finding that mitochondrial dysfunction leads to instability of the nuclear genome. Together, our data suggest that gene(s) involved in mitochondria-to-nucleus communication play a role in mutagenesis and may be implicated in carcinogenesis.
Cell Line↗
Mitochondria damage checkpoint in apoptosis and genome stability.
Mitochondria perform multiple cellular functions including energy production, cell proliferation and apoptosis. Studies described in this paper suggest a role for mitochondria in maintaining genomic stability. Genomic stability appears to be dependent on mitochondrial functions involved in maintenance of proper intracellular redox status, ATP-dependent transcription, DNA replication, DNA repair and DNA recombination. To further elucidate the role of mitochondria in genomic stability, I propose a mitochondria damage checkpoint (mitocheckpoint) that monitors and responds to damaged mitochondria. Mitocheckpoint can coordinate and maintain proper balance between apoptotic and anti-apoptotic signals. When mitochondria are damaged, mitocheckpoint can be activated to help cells repair damaged mitochondria, to restore normal mitochondrial function and avoid production of mitochondria-defective cells. If mitochondria are severely damaged, mitocheckpoint may not be able to repair the damage and protect cells. Such an event triggers apoptosis. If damage to mitochondria is continuous or persistent such as damage to mitochondrial DNA resulting in mutations, mitocheckpoint may fail which can lead to genomic instability and increased cell survival in yeast. In human it can cause cancer. In support of this proposal we provide evidence that mitochondrial genetic defects in both yeast and mammalian systems lead to impaired DNA repair, increased genomic instability and increased cell survival. This study reveals molecular genetic mechanisms underlying a role for mitochondria in carcinogenesis in humans.
Adenosine Triphosphate↗
Loss of p53 function in colon cancer cells results in increased phosphocholine and total choline.
Mutations in the p53 gene are the most frequently observed genetic lesions in human cancers. Human cancers that contain a p53 mutation are more aggressive, more apt to metastasize, and more often fatal. p53 controls numerous downstream targets that can influence various outcomes such as apoptosis, growth arrest, and DNA repair. Based on previous observations using (1)H magnetic resonance spectroscopy (MRS), we have identified choline phospholipid metabolite intensities typical of increased malignancy. Here we have used (1)H MRS to characterize the choline phospholipid metabolite levels of p53(+/ +) and p53(-/-) cells, and demonstrated that loss of p53 function results in increased phosphocholine and total choline. These data suggest that the increased malignancy of cancer cells resulting from loss of p53 may be mediated, in part, through the choline phospholipid pathway.
Cell Line, Tumor↗
Mitochondrial dysfunction is a common phenotype in aging and cancer.
An interesting clue with regard to molecular mechanisms underlying age-associated cancers is the apparent defect in mitochondrial function. Recent studies demonstrate a progressive decline in mitochondrial function during aging. Studies have established that the decline in mitochondrial function is due to the accumulation of mutations in mitochondrial DNA. These observations suggest that the mitochondrial dysfunction that accompanies aging may exert a major influence on carcinogenesis.
Aging↗
Mitochondria-mediated nuclear mutator phenotype in Saccharomyces cerevisiae.
Using Saccharomyces cerevisiae as a model organism, we analyzed the consequences of disrupting mitochondrial function on mutagenesis of the nuclear genome. We measured the frequency of canavanine-resistant colonies as a measure of nuclear mutator phenotype. Our data suggest that mitochondrial dysfunction leads to a nuclear mutator phenotype (i) when oxidative phosphorylation is blocked in wild-type yeast at mitochondrial complex III by antimycin A and (ii) in mutant strains lacking the entire mitochondrial genome (rho(0)) or those with deleted mitochondrial DNA (rho(-)). The nuclear mutation frequencies obtained for antimycin A-treated cells as well as for rho(-) and rho(0) cells were approximately 2- to 3-fold higher compared to untreated control and wild-type cells, respectively. Blockage of oxidative phosphorylation by antimycin A treatment led to increased intracellular levels of reactive oxygen species (ROS). In contrast, inactivation of mitochondrial activity (rho(-) and rho(0)) led to decreased intracellular levels of ROS. We also demonstrate that in rho(0) cells the REV1, REV3 and REV7 gene products, all implicated in error-prone translesion DNA synthesis (TLS), mediate mutagenesis in the nuclear genome. However, TLS was not involved in nuclear DNA mutagenesis caused by inhibition of mitochondrial function by antimycin A. Together, our data suggest that mitochondrial dysfunction is mutagenic and multiple pathways are involved in this nuclear mutator phenotype.
Antimycin A↗