Helena Nord

A segmental maximum a posteriori approach to genome-wide copy number profiling

MOTIVATION Copy number profiling methods aim at assigning DNA copy numbers to chromosomal regions using measurements from microarray-based comparative genomic hybridizations. Among the proposed methods to this end, Hidden Markov Model (HMM)-based approaches seem promising since DNA copy number transitions are naturally captured in the model. Current discrete-index HMM-based approaches do not, however, take into account heterogeneous information regarding the genomic overlap between clones. Moreover, the majority of existing methods are restricted to chromosome-wise analysis. RESULTS We introduce a novel Segmental Maximum A Posteriori approach, SMAP, for DNA copy number profiling. Our method is based on discrete-index Hidden Markov Modeling and incorporates genomic distance and overlap between clones. We exploit a priori information through user-controllable parameterization that enables the identification of copy number deviations of various lengths and amplitudes. The model parameters may be inferred at a genome-wide scale to avoid overfitting of model parameters often resulting from chromosome-wise model inference. We report superior performances of SMAP on synthetic data when compared with two recent methods. When applied on our new experimental data, SMAP readily recognizes already known genetic aberrations including both large-scale regions with aberrant DNA copy number and changes affecting only single features on the array. We highlight the differences between the prediction of SMAP and the compared methods and show that SMAP accurately determines copy number changes and benefits from overlap consideration.

Recurrent genomic alterations in benign and malignant pheochromocytomas and paragangliomas revealed by whole-genome array comparative genomic hybridization analysis

Pheochromocytomas and abdominal paragangliomas are adrenal and extra-adrenal catecholamine-producing tumours. They arise due to heritable cancer syndromes, or more frequently occur sporadically due to an unknown genetic cause. The majority of cases are benign, but malignant tumours are observed. Previous comparative genomic hybridization (CGH) and loss of heterozygosity studies have shown frequent deletions of chromosome arms 1p, 3q and 22q in pheochromocytomas. We applied high-resolution whole-genome array CGH on 53 benign and malignant pheochromocytomas and paragangliomas to narrow down candidate regions as well as to identify chromosomal alterations more specific to malignant tumours. Minimal overlapping regions (MORs) were identified on 16 chromosomes, with the most frequent MORs of deletion (> or = 32%) occurring on chromosome arms 1p, 3q, 11p/q, 17p and 22q, while the chromosome arms 1q, 7p, 12q and 19p harboured the most common MORs of gain (> or = 14%). The most frequent MORs (61-75%) in the pheochromocytomas were identified at 1p, and the four regions of common losses encompassed 1p36, 1p32-31, 1p22-21 and 1p13. Tumours that did not show 1p loss generally demonstrated aberrations on chromosome 11. Gain of chromosomal material was significantly more frequent among the malignant cases. Moreover, gain at 19q, trisomy 12 and loss at 11q were positively associated with malignant pheochromocytomas, while 1q gain was commonly observed in the malignant paragangliomas. Our study revealed novel and narrow recurrent chromosomal regions of loss and gain at several autosomes, a prerequisite for identifying candidate tumour suppressor genes and oncogenes involved in the development of adrenal and extra-adrenal catecholamine-producing tumours.

Characterization of novel and complex genomic aberrations in glioblastoma using a 32K BAC array.

Glioblastomas (GBs) are malignant CNS tumors often associated with devastating symptoms. Patients with GB have a very poor prognosis, and despite treatment, most of them die within 12 months from diagnosis. Several pathways, such as the RAS, tumor protein 53 (TP53), and phosphoinositide kinase 3 (PIK3) pathways, as well as the cell cycle control pathway, have been identified to be disrupted in this tumor. However, emerging data suggest that these aberrations represent only a fraction of the genetic changes involved in gliomagenesis. In this study, we have applied a 32K clone-based genomic array, covering 99% of the current assembly of the human genome, to the detailed genetic profiling of a set of 78 GBs. Complex patterns of aberrations, including high and narrow copy number amplicons, as well as a number of homozygously deleted loci, were identified. Amplicons that varied both in number (three on average) and in size (1.4 Mb on average) were frequently detected (81% of the samples). The loci encompassed not only previously reported oncogenes (EGFR, PDGFRA, MDM2, and CDK4) but also numerous novel oncogenes as GRB10, MKLN1, PPARGC1A, HGF, NAV3, CNTN1, SYT1, and ADAMTSL3. BNC2, PTPLAD2, and PTPRE, on the other hand, represent novel candidate tumor suppressor genes encompassed within homozygously deleted loci. Many of these genes are already linked to several forms of cancer; others represent new candidate genes that may serve as prognostic markers or even as therapeutic targets in the future. The large individual variation observed between the samples demonstrates the underlying complexity of the disease and strengthens the demand for an individualized therapy based on the genetic profile of the patient.

PATZ1 down-regulates FADS1 by binding to rs174557 and is opposed by SP1/SREBP1c

Abstract The FADS1 and FADS2 genes in the FADS cluster encode the rate-limiting enzymes in the synthesis of long-chain polyunsaturated fatty acids (LC-PUFAs). Genetic variation in this region has been associated with a large number of diseases and traits many of them correlated to differences in metabolism of PUFAs. However, the causative variants leading to these associations have not been identified. Here we find that the multiallelic rs174557 located in an AluYe5 element in intron 1 of FADS1 is functional and lies within a PATZ1 binding site. The derived allele of rs174557, which is the common variant in most populations, diminishes binding of PATZ1, a transcription factor conferring allele-specific downregulation of FADS1. The PATZ1 binding site overlaps with a SP1 site. The competitive binding between the suppressive PATZ1 and the activating complex of SP1 and SREBP1c determines the enhancer activity of this region, which regulates expression of FADS1.

lobChIP: from cells to sequencing ready ChIP libraries in a single day

BackgroundChIP-seq is the method of choice for genome-wide studies of protein–DNA interactions. We describe a new method for ChIP-seq sample preparation, termed lobChIP, where the library reactions are performed on cross-linked ChIP fragments captured on beads.ResultsThe lobChIP method was found both to reduce time and cost and to simplify the processing of many samples in parallel. lobChIP has an early incorporation of barcoded sequencing adaptors that minimizes the risk of sample cross-contamination and can lead to reduced amount of adaptor dimers in the sequencing libraries, while allowing for direct decross-linking and amplification of the sample.ConclusionsWith results for histone modifications and transcription factors, we show that lobChIP performs equal to or better than standard protocols and that it makes it possible to go from cells to sequencing ready libraries within a single day.

Allele-specific transcription factor binding in liver and cervix cells unveils many likely drivers of GWAS signals

Genome-wide association studies (GWAS) point to regions with associated genetic variants but rarely to a specific gene and therefore detailed knowledge regarding the genes contributing to complex traits and diseases remains elusive. The functional role of GWAS-SNPs is also affected by linkage disequilibrium with many variants on the same haplotype and sometimes in the same regulatory element almost equally likely to mediate the effect. Using ChIP-seq data on many transcription factors, we pinpointed genetic variants in HepG2 and HeLa-S3 cell lines which show a genome-wide significant difference in binding between alleles. We identified a collection of 3713 candidate functional regulatory variants many of which are likely drivers of GWAS signals or genetic difference in expression. A recent study investigated many variants before finding the functional ones at the GALNT2 locus, which we found in our genome-wide screen in HepG2. This illustrates the efficiency of our approach.

Looking beyond GWAS: allele-specific transcription factor binding drives the association of GALNT2 to HDL-C plasma levels.

BackgroundPlasma levels of high-density lipoprotein cholesterol (HDL-C) have been associated to cardiovascular disease. The high heritability of HDL-C plasma levels has been an incentive for several genome wide association studies (GWASs) which identified, among others, variants in the first intron of the GALNT2 gene strongly associated to HDL-C levels. However, the lead GWAS SNP associated to HDL-C levels in this genomic region, rs4846914, is located outside of transcription factor (TF) binding sites defined by chromatin immunoprecipitation followed by DNA sequencing (ChIP-seq) experiments in the ENCODE project and is therefore unlikely to be functional. In this study we apply a bioinformatics approach which rely on the premise that ChIP-seq reads can identify allele specific binding of a TF at cell specific regulatory elements harboring allele specific SNPs (AS-SNPs). EMSA and luciferase assays were used to validate the allele specific binding and to test the enhancer activity of the regulatory element harboring the AS-SNP rs4846913 as well as the neighboring rs2144300 which are in high LD with rs4846914.FindingsUsing luciferase assays we found that rs4846913 and the neighboring rs2144300 displayed allele specific enhancer activity. We propose that an inhibitor binds preferentially to the rs4846913-C allele with an inhibitory boost from the synergistic binding of other TFs at the neighboring SNP rs2144300. These events influence the transcription level of GALNT2.ConclusionsThe results suggest that rs4846913 and rs2144300 drive the association to HDL-C plasma levels through an inhibitory regulation of GALNT2 rather than the reported lead GWAS SNP rs4846914.

Genetic prevention of hepatitis C virus‐induced liver fibrosis by allele‐specific downregulation of MERTK

Infection by hepatitis C virus (HCV) can result in the development of liver fibrosis and may eventually progress into cirrhosis and hepatocellular carcinoma. However, the molecular mechanisms for this process are not fully known. Several genome‐wide association studies have been carried out to pinpoint causative variants in HCV‐infected patient cohorts, but these variants are usually not the functional ones. The aim of this study was to identify the regulatory single nucleotide polymorphism associated with the risk of HCV‐induced liver fibrosis and elucidate its molecular mechanism.

Genetic prevention of HCV induced liver fibrosis by allele-specific down-regulation of MERTK†

Infection by hepatitis C virus (HCV) can result in the development of liver fibrosis and may eventually progress into cirrhosis and hepatocellular carcinoma. However, the molecular mechanisms for this process are not fully known. Several genome‐wide association studies have been carried out to pinpoint causative variants in HCV‐infected patient cohorts, but these variants are usually not the functional ones. The aim of this study was to identify the regulatory single nucleotide polymorphism associated with the risk of HCV‐induced liver fibrosis and elucidate its molecular mechanism.

Genetic prevention of hepatitis C virus-induced liver fibrosis by allele-specific downregulation of MERTK: Novel regulatory SNP linked to liver fibrosis

Infection by hepatitis C virus (HCV) can result in the development of liver fibrosis and may eventually progress into cirrhosis and hepatocellular carcinoma. However, the molecular mechanisms for this process are not fully known. Several genome‐wide association studies have been carried out to pinpoint causative variants in HCV‐infected patient cohorts, but these variants are usually not the functional ones. The aim of this study was to identify the regulatory single nucleotide polymorphism associated with the risk of HCV‐induced liver fibrosis and elucidate its molecular mechanism.