Molecular Genetics

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    Mitochondrial dysfunction and DNA methylation in type 2 diabetes and cognitive impairment
    (2018-03-14) Phillips, Nicole; Barber, Robert C.; Sun, Jie; O'Bryant, Sid E.; Johnson, Leigh A.; Silzer, Talisa K.
    Background. Mexican American populations are disproportionately affected by type 2 diabetes (T2D) and Alzheimer’s disease (AD). Although commonly characterized by the accumulation of amyloid plaques and tau tangles, the role of mitochondrial dysfunction (changes in mitochondrial dynamics, apoptosis, and/or oxidative stress signaling) in AD pathophysiology has become further elucidated over recent years. Methylation of key nuclear-encoded genes may be involved in regulation of critical mitochondrial processes. Evidence suggests that differential methylation and expression of mitochondrial related genes (e.g. POLG) may correlate with mitochondrial DNA copy number (mtDNACN). Hypothesis. We tested the following hypotheses: (1) methylation of mitochondrial-related genes is negatively correlated with their respective gene expression, (2) methylation of mitochondrial biogenesis genes (e.g., TFAM, POLG) negatively correlates with mtDNACN per cell, and (3) methylation of genes related to oxidative stress response, mitophagy, endosomal/exosomal trafficking and apoptosis correlates with cell-free mtDNA (mtDNACF) levels. Methods. DNA from 14 female Mexican American subjects enrolled in HABLE, the Healthy Aging Brains of Latino Elders cohort, was used for this study. Subjects were grouped based on T2D diagnosis, and were matched across groups based on age and cognitive status. For mitochondrial-related gene expression, cDNA was synthesized from blood buffy coat RNA and tested using the RT2 Profiler™ Human Mitochondria Array. For methylation analysis, nDNA from the blood buffy coat extract was bisulfite-converted and methylation levels were determined using the MethylationEPIC™ beadchip. Data was analyzed using Genome Studio. Buffy coat mtDNACN and plasma mtDNACF were quantified using TaqMan®-based qPCR. Methylation levels of CpG regions around candidate genes were then correlated with (1) gene expression, (2) mtDNACN, and (3) mtDNACF. Results. Preliminary results indicate that hypomethylation of some mitochondrial-related genes corresponds with increased expression (e.g., COX10); methylation of two sites associated with a known CpG island for POLG are negatively correlated with mtDNA copy number per cell. Conclusions. Significant correlations between mitochondrial phenotypes and candidate gene epigenetic loci may point to novel regulatory mechanisms of mitochondrial function. Future studies will include exploratory analysis at the genome-wide level using a larger cohort.
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    DNA repair polymorphisms and age-related diseases - Alzheimer’s and Cancer: Insights from SNP-set analysis and gene expression association
    (2018-03-14) Phillips, Nicole R.; Pathak, Gita A.
    Purpose: DNA repair response is a common thread for age-related diseases. Genomic stability is the result of an elaborate machinery consisting of damage response, repair, cell-cycle checkpoints, and apoptosis. A compromised DNA damage-repair response either due to time-dependent accumulation of damage or an individual’s reduced DNA repair capacity has been known to derail the genomic defenses, resulting in disease. Recent research findings and epidemiological studies speculate an inverse association between Alzheimer’s and cancer. Since impaired DNA repair is known to accelerate age-related disease, our goal is to evaluate DNA damage/repair genes and identify the role of DNA repair polymorphisms in Alzheimer’s, Breast and Prostate Cancer in individuals. Methods: The raw genotype and phenotype data were obtained via authorized access application for Alzheimer’s Disease Neuroimaging Initiative and Breast and Prostate Cancer Cohort Consortium; genotype data were generated using the Illumina Human Quad610™ Beadchip. Controls with positive family history were removed; all subjects used were [greater than] 50 years. Data were processed with in-house codes for QC, mapping SNPs to genes and extracting SNP sets based on 274 candidate genes. SNPs within each set were tested (permutation protocol, mperm=5000) and interpreted for biological relevance after correcting for multiple set-tests. Association analyses accounted for key covariates such as age and sex. Results with genomic inflation of more than 1.03 were adjusted using first three eigenvectors as covariates. Gene expression was imputed for candidate genes. Analyses were performed in Plink(v1.9), EIGENSOFT-6.4, Rstudio 3.4, Bioconductor 3.6, VEGA2, Python 3, PrediXcan and MAGMA. Results: After two-level QC filtering, the datasets – ADNI, Breast and Prostate cancer – had 677, 578 and 3857 individuals, respectively. Gene-sets of ~167 genes were created for each dataset. Preliminary results point to cancer-specific variants in key DNA repair genes, some of which have not previously been reported. Structured sets of DNA repair pathways and gene expression imputation are in the analysis phase. Conclusion: This study investigates DNA repair genes in both cancer and Alzheimer’s using SNP-set analysis to improve detection of association that sometimes get lost in whole-genome associations. Our results provide a detailed overview of various DNA repair genes and their association with complex phenotypes of age-associated diseases.