ABSTRACT
The three EglN prolyl hydroxylases (EglN1, EglN2, and EglN3) regulate the stability of the HIF transcription factor. We recently showed that loss of EglN2, however, also leads to down-regulation of Cyclin D1 and decreased cell proliferation in a HIF-independent manner. Here we report that EglN2 can hydroxylate FOXO3a on two specific prolyl residues in vitro and in vivo. Hydroxylation of these sites prevents the binding of USP9x deubiquitinase, thereby promoting the proteasomal degradation of FOXO3a. FOXO transcription factors can repress Cyclin D1 transcription. Failure to hydroxylate FOXO3a promotes its accumulation in cells, which in turn suppresses Cyclin D1 expression. These findings provide new insights into post-transcriptional control of FOXO3a and provide a new avenue for pharmacologically altering Cyclin D1 activity.
Subject(s)
Forkhead Transcription Factors/metabolism , Gene Expression Regulation, Neoplastic , Hypoxia-Inducible Factor-Proline Dioxygenases/metabolism , Ubiquitin Thiolesterase/metabolism , Animals , Cell Line , Cells, Cultured , Cyclin D1/genetics , Forkhead Box Protein O3 , Hydroxylation , MCF-7 Cells , Mice , Protein Binding , Protein StabilityABSTRACT
We developed a cell division-activated Cre-lox system for stochastic recombination of loxP-flanked loci in mice. Cre activation by frameshift reversion is modulated by DNA mismatch-repair status and occurs in individual cells surrounded by normal tissue, mimicking spontaneous cancer-causing mutations. This system should be particularly useful for delineating pathways of neoplasia, and determining the developmental and aging consequences of specific gene alterations.
Subject(s)
Adenosine Triphosphatases/genetics , DNA Mismatch Repair , DNA Repair Enzymes/genetics , DNA-Binding Proteins/genetics , Disease Models, Animal , Integrases/genetics , Aging/genetics , Animals , Frameshift Mutation , Genes, ras/genetics , Intestines/enzymology , Mice , Mismatch Repair Endonuclease PMS2 , Neoplasms/etiology , Neoplasms/genetics , Recombination, Genetic , beta-Galactosidase/geneticsABSTRACT
A pretumor progression model predicts many oncogenic cancer mutations may first accumulate in normal appearing colon. Although direct observations of early pretumor mutations are impractical, it may be possible to retrospectively reconstruct tumor histories from contemporary cancer mutations. To infer when and in what order mutations occur during occult pretumor progression, we examined 14 cancers from individuals with heterozygous germline mutations in DNA mismatch repair (MMR) genes or hereditary nonpolyposis colorectal cancer (HNPCC). Somatic inactivation of the normal allele occurs sometime during a lifetime and results in loss of MMR, elevated mutation rates, and subsequent widespread somatic microsatellite mutations in HNPCC cancers. Patient ages at MMR loss can be estimated because intervals between MMR loss and cancer removal can be inferred from numbers of microsatellite tumor mutations. The relative order of MMR loss during pretumor progression may also be inferred from its collective ages of occurrence. Somatic MMR loss preceded cancer removal by an average of 6.1 years, occurred relatively late in life (average of 41.6 versus 47.7 years at cancer removal), and was a surprisingly late (fifth or sixth) step. Calculations indicate five or six oncogenic mutations could accumulate with relatively normal replication fidelity in normal appearing colon. HNPCC pretumor progression essentially begins from birth and ends with MMR loss, implying elevated mutation rates and tumorigenesis may be unnecessary for most progression.
Subject(s)
Colorectal Neoplasms, Hereditary Nonpolyposis/genetics , DNA-Binding Proteins/genetics , Neoplasm Proteins/genetics , Precancerous Conditions/pathology , Proto-Oncogene Proteins/genetics , Adaptor Proteins, Signal Transducing , Adult , Age Factors , Base Pair Mismatch/genetics , Carrier Proteins , Cell Transformation, Neoplastic , DNA Mutational Analysis , DNA Repair/genetics , Disease Progression , Germ-Line Mutation , Humans , Microsatellite Repeats/genetics , Middle Aged , Models, Biological , MutL Protein Homolog 1 , MutS Homolog 2 Protein , Nuclear Proteins , Retrospective Studies , Time FactorsABSTRACT
Diet is an important risk factor for many cancers. High fat/low calcium (HFLC) diets are associated with increased tumorigenesis, whereas caloric restriction (CR) reproducibly increases lifespan and decreases tumors. Mutations are involved in aging and cancer, and different diets may alter mutagenesis. However, a number of repair pathways normally counteract mutations by correcting errors before they can be fixed in the genome. To further understand interactions between diet, aging and cancer, mice deficient in a major repair pathway called DNA mismatch repair (MMR) were fed HFLC, CR or control diets. Mlh1 deficient mice are prone to lymphomas and intestinal adenomas and carcinomas. No significant changes in adenocarcinoma or lymphoma incidence were observed with HFLC or CR diets. Significantly more (2.2-fold) adenomas occurred with HFLC diets although adenoma numbers were unchanged with CR. Only a small increase in lifespan (116% of control) was achieved with CR. In addition, levels of microsatellite mutations in the small and large intestines were unchanged with the different diets. Our studies indicate that MMR deficiency may be epistatic to certain otherwise strong environmental influences on carcinogenesis or aging.
