Changqing Zeng

Integrating common and rare genetic variation in diverse human populations.

Despite great progress in identifying genetic variants that influence human disease, most inherited risk remains unexplained. A more complete understanding requires genome-wide studies that fully examine less common alleles in populations with a wide range of ancestry. To inform the design and interpretation of such studies, we genotyped 1.6 million common single nucleotide polymorphisms (SNPs) in 1,184 reference individuals from 11 global populations, and sequenced ten 100-kilobase regions in 692 of these individuals. This integrated data set of common and rare alleles, called ‘HapMap 3’, includes both SNPs and copy number polymorphisms (CNPs). We characterized population-specific differences among low-frequency variants, measured the improvement in imputation accuracy afforded by the larger reference panel, especially in imputing SNPs with a minor allele frequency of ≤5%, and demonstrated the feasibility of imputing newly discovered CNPs and SNPs. This expanded public resource of genome variants in global populations supports deeper interrogation of genomic variation and its role in human disease, and serves as a step towards a high-resolution map of the landscape of human genetic variation.

Natural selection on EPAS1 (HIF2α) associated with low hemoglobin concentration in Tibetan highlanders

By impairing both function and survival, the severe reduction in oxygen availability associated with high-altitude environments is likely to act as an agent of natural selection. We used genomic and candidate gene approaches to search for evidence of such genetic selection. First, a genome-wide allelic differentiation scan (GWADS) comparing indigenous highlanders of the Tibetan Plateau (3,200–3,500 m) with closely related lowland Han revealed a genome-wide significant divergence across eight SNPs located near EPAS1. This gene encodes the transcription factor HIF2α, which stimulates production of red blood cells and thus increases the concentration of hemoglobin in blood. Second, in a separate cohort of Tibetans residing at 4,200 m, we identified 31 EPAS1 SNPs in high linkage disequilibrium that correlated significantly with hemoglobin concentration. The sex-adjusted hemoglobin concentration was, on average, 0.8 g/dL lower in the major allele homozygotes compared with the heterozygotes. These findings were replicated in a third cohort of Tibetans residing at 4,300 m. The alleles associating with lower hemoglobin concentrations were correlated with the signal from the GWADS study and were observed at greatly elevated frequencies in the Tibetan cohorts compared with the Han. High hemoglobin concentrations are a cardinal feature of chronic mountain sickness offering one plausible mechanism for selection. Alternatively, as EPAS1 is pleiotropic in its effects, selection may have operated on some other aspect of the phenotype. Whichever of these explanations is correct, the evidence for genetic selection at the EPAS1 locus from the GWADS study is supported by the replicated studies associating function with the allelic variants.

A six-nucleotide insertion-deletion polymorphism in the CASP8 promoter is associated with susceptibility to multiple cancers

Caspases are important in the life and death of immune cells and therefore influence immune surveillance of malignancies. We tested whether genetic variants in CASP8, CASP10 and CFLAR, three genes important for death receptor–induced cell killing residing in tandem order on chromosome 2q33, are associated with cancer susceptibility. Using a haplotype-tagging SNP approach, we identified a six-nucleotide deletion (−652 6N del) variant in the CASP8 promoter associated with decreased risk of lung cancer. The deletion destroys a stimulatory protein 1 binding site and decreases CASP8 transcription. Biochemical analyses showed that T lymphocytes with the deletion variant had lower caspase-8 activity and activation-induced cell death upon stimulation with cancer cell antigens. Case-control analyses of 4,995 individuals with cancer and 4,972 controls in a Chinese population showed that this genetic variant is associated with reduced susceptibility to multiple cancers, including lung, esophageal, gastric, colorectal, cervical and breast cancers, acting in an allele dose–dependent manner. These results support the hypothesis that genetic variants influencing immune status modify cancer susceptibility.

Rapid growth of a hepatocellular carcinoma and the driving mutations revealed by cell-population genetic analysis of whole-genome data

We present the analysis of the evolution of tumors in a case of hepatocellular carcinoma. This case is particularly informative about cancer growth dynamics and the underlying driving mutations. We sampled nine different sections from three tumors and seven more sections from the adjacent nontumor tissues. Selected sections were subjected to exon as well as whole-genome sequencing. Putative somatic mutations were then individually validated across all 9 tumor and 7 nontumor sections. Among the mutations validated, 24 were amino acid changes; in addition, 22 large indels/copy number variants (>1 Mb) were detected. These somatic mutations define four evolutionary lineages among tumor cells. Separate evolution and expansion of these lineages were recent and rapid, each apparently having only one lineage-specific protein-coding mutation. Hence, by using a cell-population genetic definition, this approach identified three coding changes (CCNG1, P62, and an indel/fusion gene) as tumor driver mutations. These three mutations, affecting cell cycle control and apoptosis, are functionally distinct from mutations that accumulated earlier, many of which are involved in inflammation/immunity or cell anchoring. These distinct functions of mutations at different stages may reflect the genetic interactions underlying tumor growth.

Assessment of Immunoreactive Synthetic Peptides from the Structural Proteins of Severe Acute Respiratory Syndrome Coronavirus

Abstract Background: The widespread threat of severe acute respiratory syndrome (SARS) to human life has spawned challenges to develop fast and accurate analytical methods for its early diagnosis and to create a safe antiviral vaccine for preventive use. Consequently, we thoroughly investigated the immunoreactivities with patient sera of a series of synthesized peptides from SARS-coronavirus structural proteins. Methods: We synthesized 41 peptides ranging in size from 16 to 25 amino acid residues of relatively high hydrophilicity. The immunoreactivities of the peptides with SARS patient sera were determined by ELISA. Results: Four epitopic sites, S599, M137, N66, and N371-404, located in the SARS-coronavirus S, M, and N proteins, respectively, were detected by screening synthesized peptides. Notably, N371 and N385, located at the COOH terminus of the N protein, inhibited binding of antibodies to SARS-coronavirus lysate and bound to antibodies in >94% of samples from SARS study patients. N385 had the highest affinity for forming peptide-antibody complexes with SARS serum. Conclusions: Five peptides from SARS structural proteins, especially two from the COOH terminus of the N protein, appear to be highly immunogenic and may be useful for serologic assays. The identification of these antigenic peptides contributes to the understanding of the immunogenicity and persistence of SARS coronavirus.

Community Engagement and Informed Consent in the International HapMap Project

The International HapMap Consortium has developed the HapMap, a resource that describes the common patterns of human genetic variation (haplotypes). Processes of community/public consultation and individual informed consent were implemented in each locality where samples were collected to understand and attempt to address both individual and group concerns. Perceptions about the research varied, but we detected no critical opposition to the research. Incorporating community input and responding to concerns raised was challenging. However, the experience suggests that approaching genetic variation research in a spirit of openness can help investigators better appreciate the views of the communities whose samples they seek to study and help communities become more engaged in the science.

Haplotypes in Matrix Metalloproteinase Gene Cluster on Chromosome 11q22 Contribute to the Risk of Lung Cancer Development and Progression

Purpose: Matrix metalloproteinases (MMP) play important roles in cancer development and single nucleotide polymorphisms (SNP) in some MMP genes were shown to confer susceptibility to certain cancers. This study examined the association between genotypes and haplotypes in the MMP1-MMP3-MMP12 gene cluster and risk of lung cancer development and metastasis. Experimental Design: A two-stage investigation was conducted. First, 35 SNPs covering these genes were selected and validated in 190 patients and 190 controls. Twenty-two validated SNPs were then analyzed in an entire case-control panel consisting of 711 patients and 716 controls. Associations with the risk of lung cancer were estimated by logistic regression. Results: The investigated MMP gene region could be partitioned into two major haplotype blocks. One common haplotype in the block composed of major part of MMP1 transcription region was significantly associated with increased risk for the development [odds ratio (OR), 1.35; 95% confidence interval (95% CI), 1.11-1.63; P = 0.01; permutated P = 0.134] and distant metastasis of lung cancer (ORs for stage IV versus stages I-III, 1.67; 95% CI, 1.12-2.50; P = 0.009; permutated P = 0.048) and the other showed a protective effect against metastasis (ORs for stage IV versus stages I-III, 0.22; 95% CI, 0.07-0.62; P = 0.001; permutated P = 0.011). Another common haplotype in the block across MMP3 was significantly associated with decreased risk for developing lung cancer (OR, 0.71; 95% CI, 0.59-0.86; P = 0.003; permutated P = 0.027). Conclusions: The observed multiple cancer-associated genetic variants suggested that the MMP1-MMP3-MMP12 gene cluster plays important roles in lung cancer development and progression.

SNP@Evolution: a hierarchical database of positive selection on the human genome.

BackgroundPositive selection is a driving force that has shaped the modern human. Recent developments in high throughput technologies and corresponding statistics tools have made it possible to conduct whole genome surveys at a population scale, and a variety of measurements, such as heterozygosity (HET), FST, and Tajimas D, have been applied to multiple datasets to identify signals of positive selection. However, great effort has been required to combine various types of data from individual sources, and incompatibility among datasets has been a common problem. SNP@Evolution, a new database which integrates multiple datasets, will greatly assist future work in this area.DescriptionAs part of our research scanning for evolutionary signals in HapMap Phase II and Phase III datasets, we built SNP@Evolution as a multi-aspect database focused on positive selection. Among its many features, SNP@Evolution provides computed FSTand HET of all HapMap SNPs, 5+ HapMap SNPs per qualified gene, and all autosome regions detected from whole genome window scanning. In an attempt to capture multiple selection signals across the genome, selection-signal enrichment strength (ES) values of HET, FST, and P-values of iHS of most annotated genes have been calculated and integrated within one frame for users to search for outliers. Genes with significant ES or P-values (with thresholds of 0.95 and 0.05, respectively) have been highlighted in color. Low diversity chromosome regions have been detected by sliding a 100 kb window in a 10 kb step. To allow this information to be easily disseminated, a graphical user interface (GBrowser) was constructed with the Generic Model Organism Database toolkit.ConclusionAvailable at http://bighapmap.big.ac.cn, SNP@Evolution is a hierarchical database focused on positive selection of the human genome. Based on HapMap Phase II and III data, SNP@Evolution includes 3,619,226/1,389,498 SNPs with their computed HET and FST, as well as qualified genes of 21,859/21,099 with ES values of HET and FST. In at least one HapMap population group, window scanning for selection signals has resulted in 1,606/10,138 large low HET regions. Among Phase II and III geographical groups, 660 and 464 regions show strong differentiation.

An unusual haplotype structure on human chromosome 8p23 derived from the inversion polymorphism

Chromosomal inversion is an important type of genomic variations involved in both evolution and disease pathogenesis. Here, we describe the refined genetic structure of a 3.8‐Mb inversion polymorphism at chromosome 8p23. Using HapMap data of 1,073 SNPs generated from 209 unrelated samples from CEPH—Utah residents with ancestry from northern and western Europe (CEU); Yoruba in Ibadan, Nigeria (YRI); and Asian (ASN) samples, which were comprised of Han Chinese from Beijing, China (CHB) and Japanese from Tokyo, Japan (JPT)—we successfully deduced the inversion orientations of all their 418 haplotypes. In particular, distinct haplotype subgroups were identified based on principal component analysis (PCA). Such genetic substructures were consistent with clustering patterns based on neighbor‐joining tree reconstruction, which revealed a total of four haplotype clades across all samples. Metaphase fluorescence in situ hybridization (FISH) in a subset of 10 HapMap samples verified their inversion orientations predicted by PCA or phylogenetic tree reconstruction. Positioning of the outgroup haplotype within one of YRI clades suggested that Human NCBI Build 36‐inverted order is most likely the ancestral orientation. Furthermore, the population differentiation test and the relative extended haplotype homozygosity (REHH) analysis in this region discovered multiple selection signals, also in a population‐specific manner. A positive selection signal was detected at XKR6 in the ASN population. These results revealed the correlation of inversion polymorphisms to population‐specific genetic structures, and various selection patterns as possible mechanisms for the maintenance of a large chromosomal rearrangement at 8p23 region during evolution. In addition, our study also showed that haplotype‐based clustering methods, such as PCA, can be applied in scanning for cryptic inversion polymorphisms at a genome‐wide scale. Hum Mutat 0, 1–8, 2008.

Discovery and validation of prognostic markers in gastric cancer by genome-wide expression profiling

AIM To develop a prognostic gene set that can predict patient overall survival status based on the whole genome expression analysis. METHODS Using Illumina HumanWG-6 BeadChip followed by semi-supervised analysis, we analyzed the expression of 47,296 transcripts in two batches of gastric cancer patients who underwent surgical resection. Thirty-nine samples in the first batch were used as the training set to discover candidate markers correlated to overall survival, and thirty-three samples in the second batch were used for validation. RESULTS A panel of ten genes were identified as prognostic marker in the first batch samples and classified patients into a low- and a high-risk group with significantly different survival times (P = 0.000047). This prognostic marker was then verified in an independent validation sample batch (P = 0.0009). By comparing with the traditional Tumor-node-metastasis (TNM) staging system, this ten-gene prognostic marker showed consistent prognosis results. It was the only independent prognostic value by multivariate Cox regression analysis (P = 0.007). Interestingly, six of these ten genes are ribosomal proteins, suggesting a possible association between the deregulation of ribosome related gene expression and the poor prognosis. CONCLUSION A ten-gene marker correlated with overall prognosis, including 6 ribosomal proteins, was identified and verified, which may complement the predictive value of TNM staging system.