Jung Kyoon Choi

Differential coexpression analysis using microarray data and its application to human cancer

MOTIVATION Microarrays have been used to identify differential expression of individual genes or cluster genes that are coexpressed over various conditions. However, alteration in coexpression relationships has not been studied. Here we introduce a model for finding differential coexpression from microarrays and test its biological validity with respect to cancer. RESULTS We collected 10 published gene expression datasets from cancers of 13 different tissues and constructed 2 distinct coexpression networks: a tumor network and normal network. Comparison of the two networks showed that cancer affected many coexpression relationships. Functional changes such as alteration in energy metabolism, promotion of cell growth and enhanced immune activity were accompanied with coexpression changes. Coregulation of collagen genes that may control invasion and metastatic spread of tumor cells was also found. Cluster analysis in the tumor network identified groups of highly interconnected genes related to ribosomal protein synthesis, the cell cycle and antigen presentation. Metallothionein expression was also found to be clustered, which may play a role in apoptosis control in tumor cells. Our results show that this model would serve as a novel method for analyzing microarrays beyond the specific implications for cancer.

Intrinsic variability of gene expression encoded in nucleosome positioning sequences

Variation in gene expression is an essential material for biological diversity among single cells, individuals and populations or species. Here we show that expression variability is an intrinsic property that persists at those different levels. Each promoter seems to have a unique capacity to respond to external signals that can be environmental, genetic or even stochastic. Our investigation into nucleosome organization of variably responding promoters revealed a commonly positioned nucleosome at a critical regulatory region where most transcription start sites and TATA elements are located, a deviation from typical nucleosome-free status. The nucleotide sequences in this region of variable promoters showed a high propensity for DNA bending and a periodic distribution of particular dinucleotides, encoding preferences for DNA–nucleosome interaction. Variable expression is likely to occur during removal of this nucleosome for gene activation. This is a unique example of how promoter sequences intrinsically encode regulatory flexibility, which is vital for biological processes such as adaptation, development and evolution.

Epigenetic regulation and the variability of gene expression

We characterized the genetic variability of gene expression in terms of trans and cis variability for each yeast transcript. Genes that are highly regulated by nucleosomes showed a high degree of trans variability. From the expression profiles of mutants for various chromatin modifiers, we found that trans-variable genes are distinctly regulated at the chromatin level. The effect of chromatin regulators was highly significant, even when compared with that of transcription factors. The DNA-binding activities of transcription factors had a low influence on trans variability. In the case of the basal transcription factor TBP and TBP-associated factor TAF1, expression variability was coupled with the histone acetyltransferase activities of TAF1 and other factors, rather than with the binding of TBP to DNA. Additionally, we found that the correlation of TATA-box presence and expression variability could be explained in terms of chromatin regulation. The lack of activating histone modifications may subject TATA-containing promoters to chromatin regulation processes. Our results propose that epigenetic regulation has a central role in the variation and evolution of gene expression.

Contrasting chromatin organization of CpG islands and exons in the human genome

BackgroundCpG islands and nucleosome-free regions are both found in promoters. However, their association has never been studied. On the other hand, DNA methylation is absent in promoters but is enriched in gene bodies. Intragenic nucleosomes and their modifications have been recently associated with RNA splicing. Because the function of intragenic DNA methylation remains unclear, I explored the possibility of its involvement in splicing regulation.ResultsHere I show that CpG islands were associated not only with methylation-free promoters but also with nucleosome-free promoters. Nucleosome-free regions were observed only in promoters containing a CpG island. However, the DNA sequences of CpG islands predicted the opposite pattern, implying a limitation of sequence programs for the determination of nucleosome occupancy. In contrast to the methylation-and nucleosome-free states of CpG-island promoters, exons were densely methylated at CpGs and packaged into nucleosomes. Exon-enrichment of DNA methylation was specifically found in spliced exons and in exons with weak splice sites. The enrichment patterns were less pronounced in initial exons and in non-coding exons, potentially reflecting a lower need for their splicing. I also found that nucleosomes, DNA methylation, and H3K36me3 marked the exons of transcripts with low, medium, and high gene expression levels, respectively.ConclusionsHuman promoters containing a CpG island tend to remain nucleosome-free as well as methylation-free. In contrast, exons demonstrate a high degree of methylation and nucleosome occupancy. Exonic DNA methylation seems to function together with exonic nucleosomes and H3K36me3 for the proper splicing of transcripts with different expression levels.

Integrative analysis of multiple gene expression profiles applied to liver cancer study

A statistical method for combining multiple microarray studies has been previously developed by the authors. Here, we present the application of the method to our hepatocellular carcinoma (HCC) data and report new findings on gene expression changes accompanying HCC. From the cross‐verification result of our studies and that of published studies, we found that single microarray analysis might lead to false findings. To avoid those pitfalls of single‐set analyses, we employed our effect size method to integrate multiple datasets. Of 9982 genes analyzed, 477 significant genes were identified with a false discovery rate of 10%. Gene ontology (GO) terms associated with these genes were explored to validate our method in the biological context with respect to HCC. Furthermore, it was demonstrated that the data integration process increases the sensitivity of analysis and allows small but consistent expression changes to be detected. These integration‐driven discoveries contained meaningful and interesting genes not reported in previous expression profiling studies, such as growth hormone receptor, erythropoietin receptor, tissue factor pathway inhibitor‐2, etc. Our findings support the use of meta‐analysis for a variety of microarray data beyond the scope of this specific application.

Nucleosome deposition and DNA methylation at coding region boundaries

BackgroundNucleosome deposition downstream of transcription initiation and DNA methylation in the gene body suggest that control of transcription elongation is a key aspect of epigenetic regulation.ResultsHere we report a genome-wide observation of distinct peaks of nucleosomes and methylation at both ends of a protein coding unit. Elongating polymerases tend to pause near both coding ends immediately upstream of the epigenetic peaks, causing a significant reduction in elongation efficiency. Conserved features in underlying protein coding sequences seem to dictate their evolutionary conservation across multiple species. The nucleosomal and methylation marks are commonly associated with high sequence-encoded DNA-bending propensity but differentially with CpG density. As the gene grows longer, the epigenetic codes seem to be shifted from variable inner sequences toward boundary regions, rendering the peaks more prominent in higher organisms.ConclusionsRecent studies suggest that epigenetic inhibition of transcription elongation facilitates the inclusion of constitutive exons during RNA splicing. The epigenetic marks we identified here seem to secure the first and last coding exons from exon skipping as they are indispensable for accurate translation.

Genetic and Metabolic Characterization of Insomnia

Insomnia is reported to chronically affect 10∼15% of the adult population. However, very little is known about the genetics and metabolism of insomnia. Here we surveyed 10,038 Korean subjects whose genotypes have been previously profiled on a genome-wide scale. About 16.5% reported insomnia and displayed distinct metabolic changes reflecting an increase in insulin secretion, a higher risk of diabetes, and disrupted calcium signaling. Insomnia-associated genotypic differences were highly concentrated within genes involved in neural function. The most significant SNPs resided in ROR1 and PLCB1, genes known to be involved in bipolar disorder and schizophrenia, respectively. Putative enhancers, as indicated by the histone mark H3K4me1, were discovered within both genes near the significant SNPs. In neuronal cells, the enhancers were bound by PAX6, a neural transcription factor that is essential for central nervous system development. Open chromatin signatures were found on the enhancers in human pancreas, a tissue where PAX6 is known to play a role in insulin secretion. In PLCB1, CTCF was found to bind downstream of the enhancer and interact with PAX6, suggesting that it can probably inhibit gene activation by PAX6. PLCB4, a circadian gene that is closely located downstream of PLCB1, was identified as a candidate target gene. Hence, dysregulation of ROR1, PLCB1, or PLCB4 by PAX6 and CTCF may be one mechanism that links neural and pancreatic dysfunction not only in insomnia but also in the relevant psychiatric disorders that are accompanied with circadian rhythm disruption and metabolic syndrome.

Environmental Effects on Gene Expression Phenotype Have Regional Biases in the Human Genome

Phenotypic discordance between monozygotic twins, such as a difference in disease susceptibility, implicates the role of the environment in determining phenotype. To assess genomewide environmental effects on “gene expression phenotype,” we employed a published microarray data set for twins. We found that variations in expression phenotypes between monozygotic twins have biases in their chromosomal locations. They also showed a strong inverse correlation with gene density. Genomic regions of low gene density were environmentally sensitive, containing genes involved in response to external signals, cell differentiation, and development, etc. Genetic factors were found to make no contribution to the observed regional biases, stressing the role of epigenetics. We propose that epigenetic modifications might occur more frequently in heterochromatic, gene-poor regions in response to environmental signals while gene-rich regions tend to remain in an active chromatin configuration for the constitutive expression of underlying genes.

HBx induces hypomethylation of distal intragenic CpG islands required for active expression of developmental regulators

Significance Epigenetic dysregulation by oncoviral protein plays a key role in tumor development. DNA methylome analysis of hepatitis B virus X (HBx)-induced hepatocellular carcinoma (HCC) revealed drastic changes in host epigenome, but in an unconventional way: intragenic CpG islands (CGIs) were dramatically demethylated. We showed methylated intragenic CGIs as previously unidentified regulatory elements associated with active expression. The methylated CGIs are marked with distinct epigenetic signatures and require DNA methyltransferase (DNMT) 3L complex for their high methylation levels. By directly suppressing Dnmt3L and Dnmt3a promoters, HBx induces hypomethylation of the intragenic CGIs and downregulation of the associated developmental regulators. We provide previously unreported functional identification of intragenic CGIs that may enhance our understanding of epigenetic regulation and a new epigenetic role for HBx in promoting HCC development. Epigenetic alterations caused by viral oncoproteins are strong initiation factors for cancer development, but their mechanisms are largely unknown. To identify the epigenetic effects of viral hepatitis B virus X (HBx) that lead to hepatocellular carcinoma (HCC), we profiled the DNA methylomes of normal and HBx transgenic mouse liver. Intriguingly, severe hypomethylation of intragenic CpG islands (CGIs) was observed in HBx liver before the full development of HCC. Normally, these CGIs were highly methylated (mCGIs) by the DNMT3L complex and marked with epigenetic signatures associated with active expression, such as H3K36me3. Hypomethylation of mCGI was caused by the downregulation of Dnmt3L and Dnmt3a due to HBx bound to their promoters, along with HDAC1. These events lead to the downregulation of many developmental regulators that could facilitate tumorigenesis. Here we provide an intriguing epigenetic regulation mediated by mCGI that is required for cell differentiation and describe a previously unidentified epigenetic role for HBx in promoting HCC development.

The characteristics of genome-wide DNA methylation in naïve CD4+ T cells of patients with psoriasis or atopic dermatitis

Psoriasis and atopic dermatitis (AD) are skin diseases that are characterized by polarized CD4+ T cell responses. During the polarization of naïve CD4+ T cells, DNA methylation plays an important role in the regulation of gene transcription. In this study, we profiled the genome-wide DNA methylation status of naïve CD4+ T cells in patients with psoriasis or AD and healthy controls using a ChIP-seq method. Only psoriasis patient T cells, not those of AD patients, showed distinct hypomethylation (>4-fold) compared to healthy control T cells in twenty-six regions of the genome ranging in size from 10 to 70 kb. These regions were mostly pericentromeric on 10 different chromosomes and incidentally coincided with various strong epigenomic signals, such as histone modifications and transcription factor binding sites, that had been observed in the ENCODE project implying the potential epigenetic regulation in psoriasis development. The gene-centric analysis indicated that the promoter regions of 121 genes on the X chromosome had dramatically elevated methylation levels in psoriasis patient T-cells compared to those from healthy controls (>4-fold). Moreover, immune-related genes on the X chromosome had higher hypermethylation than other genes (P=0.046). No such patterns were observed with AD patient T cells. These findings imply that methylation changes in naïve CD4+ T cells may affect CD4+ T cell polarization, especially in the pathogenesis of psoriasis.