Inter-genomic cross talk between mitochondria and the nucleus plays an important role in tumorigenesis

Abstract

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.

Introduction

Mitochondria perform multiple essential cellular functions (Modica-Napolitano and Singh, 2002, Modica-Naplotinano and Singh, 2004). Although the mitochondrial and nuclear genomes are physically distinct, there is a high degree of functional interdependence between the two genomes. Of the 82 structural subunits that make up the oxidative phosphorylation system in the mitochondria, the mitochondrial genome encodes 13 subunits (see below) and the rest of the subunits are encoded by the nuclear genome. Correct mitochondrial functions depend on an orchestrated cross talk between the nuclear and mitochondrial genomes.

The mitochondrial genome is a small 16.6 unit molecule that encodes 13 subunits of the respiratory chain complexes, 22 tRNAs and 2 ribosomal RNAs. Mammalian cells typically contain 10³–10⁴ copies of mitochondrial DNA (mtDNA). Unlike nuclear DNA, mammalian mtDNA contains no introns, has no protective histones and is exposed to deleterious reactive oxygen species generated by oxidative phosphorylation. In addition, replication of mtDNA might be error prone. The accumulation of mutations in mtDNA is approximately tenfold greater than that in nuclear DNA (Grossman and Shoubridge, 1996, Johns, 1995, Penta et al., 2001). Mutations in mtDNA have been reported in mitochondrial diseases and in a variety of cancers, including ovarian, thyroid, salivary, kidney, liver, lung, colon, gastric, brain, bladder, head and neck, and breast cancers, and leukemia (reviewed in Penta et al., 2001, Modica-Napolitano and Singh, 2002, Modica-Naplotinano and Singh, 2004). Deletions, point mutations, insertions and duplications have been detected throughout the genome, and certain mutations in mtDNA are associated with specific cancers. For example, a 40 bp insertion localized in the COX I gene appears to be specific for renal cell oncocytomas (Welter et al., 1989), and a deletion mutation resulting in the loss of mtDNA within NADH dehydrogenase subunit III is a phenotype associated with renal cell carcinoma (Selvanayagam and Rajaraman, 1996). The D-loop region appears particularly susceptible to DNA mutations. Both hepatocellular carcinoma (Nomoto et al., 2002) and breast cancer (Parrella et al., 2001) are associated with certain deletion/insertion mutations in the C-tract, a hotspot and a potential replication start site within the D-loop of the mitochondrial genome. These studies suggest that mutations in mtDNA is a common feature of cancer cells. Unfortunately, to date it is not clear whether mutations in the mitochondrial genome affect nuclear genome stability and whether inter-genomic cross talk is involved in tumorigenesis.

Our studies conducted in yeast Saccharomyces cerevisiae model system suggest that mutations in the mitochondrial genome cause nuclear genomic instability (Rasmussen et al., 2003). We also identified that nuclear genome stability was mediated by Rev1p dependent error prone repair pathway (Rasmussen et al., 2003). Using a human cell culture model our recent study provided evidence that mitochondrial genomic dysfunction leads to impaired oxidative DNA repair in the nucleus (Delsite et al., 2003). Our study also revealed that mitochondrial dysfunction leads to elevated expression of MnSOD which causes resistance to apoptosis (Park et al. 2004). In the present paper, we analyzed the importance of the mitochondrial genome in chromosomal instability (CIN) and its role in tumorigenesis. This study reveal that depletion of mitochondrial genome (rho⁰) leads to 1) chromosomal instability 2) altered expression of APE1, a DNA repair gene and 3) observed instability in nuclear genome plays a critical role in tumorigenesis. Furthermore, our study for the first time suggest that the tumorigenesis phenotype and the altered expression of APE1 can be reversed by transfer of exogenous wild type mitochondrial genome into rho⁰ cells. These studies suggest that inter-genomic cross talk between mitochondria and the nucleus plays an important role in tumorigenesis.