RESUMO
Genome-wide association studies (GWASs) have identified approximately 100 colorectal cancer (CRC) risk loci. However, the causal genes in these loci have not been systematically interrogated. We conducted a high-throughput RNA-interference functional screen to identify the genes essential for proliferation in the CRC risk loci of Asian populations. We found that ATF1, located in the 12q13.12 region, functions as an oncogene that facilitates cell proliferation; ATF1 has the most significant effect of the identified genes and promotes CRC xenograft growth by affecting cell apoptosis. Next, by integrating a fine-mapping analysis, a two-stage affected-control study consisting of 6,213 affected individuals and 10,388 controls, and multipronged experiments, we elucidated that two risk variants, dbSNP: rs61926301 and dbSNP: rs7959129, that located in the ATF1 promoter and first intron, respectively, facilitate a promoter-enhancer interaction, mediated by the synergy of SP1 and GATA3, to upregulate ATF1 expression, thus synergistically predisposing to CRC risk (OR = 1.77, 95% CI = 1.42-2.21, p = 3.16 × 10-7; Pmultiplicative-interaction = 1.20 × 10-22; Padditive-interaction = 6.50 × 10-3). Finally, we performed RNA-seq and ChIP-seq assays in CRC cells treated with ATF1 overexpression in order to dissect the target programs of ATF1. Results showed that ATF1 activates a subset of genes, including BRAF, NRAS, MYC, BIRC2, DAAM1, MAML2, STAT1, ID1, and NKD2, related to apoptosis, Wnt, TGF-ß, and MAPK pathways, and these effects could cooperatively increase the risk of CRC. These findings reveal the clinical potential of ATF1 in CRC development and illuminate a promoter-enhancer interaction module between the ATF1 regulatory elements dbSNP: rs61926301 and dbSNP: rs7959129, and they bring us closer to understanding the molecular drivers of cancer.
Assuntos
Fator 1 Ativador da Transcrição/metabolismo , Neoplasias Colorretais/patologia , Elementos Facilitadores Genéticos , Regulação Neoplásica da Expressão Gênica , Estudo de Associação Genômica Ampla , Polimorfismo de Nucleotídeo Único , Regiões Promotoras Genéticas , Fator 1 Ativador da Transcrição/antagonistas & inibidores , Fator 1 Ativador da Transcrição/genética , Animais , Apoptose , Sistemas CRISPR-Cas , Estudos de Casos e Controles , Proliferação de Células , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Feminino , Edição de Genes , Predisposição Genética para Doença , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Locos de Características Quantitativas , Interferência de RNA , Fatores de Risco , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
N6-Methyladenosine (m6A) is the most prevalent modification of RNA in eukaryotes, and is associated with many cellular processes and even the development of cancers. We hypothesized that single-nucleotide polymorphisms (SNPs) in m6A modification genes, including its "writers", "erasers" and "readers", might affect the m6A functions and associate with the susceptibility to pancreatic ductal adenocarcinoma (PDAC). We first conducted a two-stage case-control study in Chinese population to interrogate all SNPs in 22 m6A modification genes. In the discovery stage, a total of 2735 SNPs were genotyped in 980 patients and 1991 controls. Then, the promising SNP was replicated in another independent population consisting of 858 cases and 2084 controls. As a result, we found the rs7495 in 3'UTR of hnRNPC was significantly associated with increased risk of PDAC in both stages (combined odds ratio = 1.22, 95% confidence interval = 1.12-1.32, P = 2.39 × 10-6). To further reveal the biological function of rs7495 and hnRNPC, we performed a series of biochemical experiments. Luciferase reporter assays indicated that rs7495G allele promoted hnRNPC expression through disrupting a putative binding site for has-miR-183-3p. Cell viability assay demonstrated that knockdown of hnRNPC suppressed the proliferation of PDAC cells. RNA-seq analysis suggested that as an m6A "reader", hnRNPC played an important role in RNA biological processes. In conclusion, our findings elucidated that rs7495G could confer higher risk of PDAC via promoting the expression of hnRNPC through a miRNA-mediated manner. These results provided a novel insight into the critical role of m6A modification in tumorigenesis.
Assuntos
Adenosina/análogos & derivados , Carcinoma Ductal Pancreático/genética , Ribonucleoproteínas Nucleares Heterogêneas Grupo C/genética , Neoplasias Pancreáticas/genética , Regiões 3' não Traduzidas/genética , Adenosina/genética , Povo Asiático/genética , Estudos de Casos e Controles , Linhagem Celular Tumoral , Proliferação de Células/genética , Sobrevivência Celular/genética , Técnicas de Silenciamento de Genes , Variação Genética , Genótipo , Humanos , MicroRNAs/genética , Polimorfismo de Nucleotídeo ÚnicoRESUMO
Protein post-translational modifications (PTMs), including phosphorylation, ubiquitination, methylation, acetylation, glycosylation et al, are very important biological processes. PTM changes in some critical genes, which may be induced by base-pair substitution, are shown to affect the risk of diseases. Recently, large-scale exome-wide association studies found that missense single nucleotide polymorphisms (SNPs) play an important role in the susceptibility for complex diseases or traits. One of the functional mechanisms of missense SNPs is that they may affect PTMs and leads to a protein dysfunction and its downstream signaling pathway disorder. Here, we constructed a database named AWESOME (A Website Exhibits SNP On Modification Event, http://www.awesome-hust.com), which is an interactive web-based analysis tool that systematically evaluates the role of SNPs on nearly all kinds of PTMs based on 20 available tools. We also provided a well-designed scoring system to compare the performance of different PTM prediction tools and help users to get a better interpretation of results. Users can search SNPs, genes or position of interest, filter with specific modifications or prediction methods, to get a comprehensive PTM change induced by SNPs. In summary, our database provides a convenient way to detect PTM-related SNPs, which may potentially be pathogenic factors or therapeutic targets.
Assuntos
Bases de Dados de Proteínas , Polimorfismo de Nucleotídeo Único , Processamento de Proteína Pós-Traducional/genética , Genoma Humano , HumanosRESUMO
Alternative splicing (AS) is a widespread process that increases structural transcript variation and proteome diversity. Aberrant splicing patterns are frequently observed in cancer initiation, progress, prognosis and therapy. Increasing evidence has demonstrated that AS events could undergo modulation by genetic variants. The identification of splicing quantitative trait loci (sQTLs), genetic variants that affect AS events, might represent an important step toward fully understanding the contribution of genetic variants in disease development. However, no database has yet been developed to systematically analyze sQTLs across multiple cancer types. Using genotype data from The Cancer Genome Atlas and corresponding AS values calculated by TCGASpliceSeq, we developed a computational pipeline to identify sQTLs from 9 026 tumor samples in 33 cancer types. We totally identified 4 599 598 sQTLs across all cancer types. We further performed survival analyses and identified 17 072 sQTLs associated with patient overall survival times. Furthermore, using genome-wide association study (GWAS) catalog data, we identified 1 180 132 sQTLs overlapping with known GWAS linkage disequilibrium regions. Finally, we constructed a user-friendly database, CancerSplicingQTL (http://www.cancersplicingqtl-hust.com/) for users to conveniently browse, search and download data of interest. This database provides an informative sQTL resource for further characterizing the potential functional roles of SNPs that control transcript isoforms in human cancer.
Assuntos
Processamento Alternativo , Bases de Dados de Ácidos Nucleicos , Neoplasias/genética , Polimorfismo de Nucleotídeo Único , Locos de Características Quantitativas , Estudo de Associação Genômica Ampla , Humanos , Desequilíbrio de LigaçãoRESUMO
OBJECTIVE: Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal cancers worldwide. Thus far, most drugs have failed to significantly improve patient survival. N6-methyladenosine (m6A) plays an important role in the progression of PDAC, but its aberrant regulation driven by germline variants in human diseases remains unclear. DESIGN: We first performed an exome-wide association analysis in 518 PDAC patients with overall survival and replicated in an independent population containing 552 PDAC patients. Then, a series of biochemical experiments in vitro and in vivo were conducted to investigate potential mechanisms of the candidate variant and its target gene PIK3CB underlying the PDAC progression. Moreover, the PIK3CB-selective inhibitor KIN-193 was used to block PDAC tumour growth. RESULTS: We identified a missense variant rs142933486 in PIK3CB that is significantly associated with the overall survival of PDAC by reducing the PIK3CB m6A level, which facilitated its mRNA and protein expression levels mediated by the m6A 'writer' complex (METTL13/METTL14/WTAP) and the m6A 'reader' YTHDF2. The upregulation of PIK3CB is widely found in PDAC tumour tissues and significantly correlated with the poor prognosis of PDAC, especially in PTEN-deficient patients. We further demonstrated that PIK3CB overexpression substantially enhanced the proliferation and migration abilities of PTEN-deficient PDAC cells and activated AKT signalling pathway. Remarkably, KIN-193, a PIK3CB-selective inhibitor, is shown to serve as an effective anticancer agent for blocking PTEN-deficient PDAC. CONCLUSIONS: These findings demonstrate aberrant m6A homoeostasis as an oncogenic mechanism in PDAC and highlight the potential of PIK3CB as a therapeutic target for this disease.
Assuntos
Carcinoma Ductal Pancreático/patologia , Classe I de Fosfatidilinositol 3-Quinases/genética , Neoplasias Pancreáticas/patologia , Adenosina/análogos & derivados , Adenosina/genética , Animais , Carcinoma Ductal Pancreático/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Progressão da Doença , Estudo de Associação Genômica Ampla , Humanos , Metilação , Camundongos Nus , Pessoa de Meia-Idade , Mutação de Sentido Incorreto , PTEN Fosfo-Hidrolase/antagonistas & inibidores , PTEN Fosfo-Hidrolase/deficiência , Neoplasias Pancreáticas/genética , Proteínas Proto-Oncogênicas c-akt , Pirimidinonas/farmacologia , RNA Mensageiro/metabolismo , Transdução de Sinais , Regulação para Cima , ortoaminobenzoatos/farmacologiaRESUMO
N 6-methyladenosine (m6A) is an abundant modification in RNAs that affects RNA metabolism, and it is reported to be closely related to cancer occurrence and metastasis. In this study, we focused on evaluating the associations between genetic variants in m6A modification genes and the risk of esophageal squamous-cell carcinoma (ESCC). By integrating data of our previous genome-wide association studies and the predictions of several annotation tools, we identified a single nucleotide polymorphism, rs2416282 in the promoter of YTHDC2, that was significantly associated with the susceptibility of ESCC (odds ratio = 0.84, 95% CI: 0.77-0.92, P = 2.81 × 10-4). Through further functional experiments in vitro, we demonstrated that rs2416282 regulated YTHDC2 expression. Knockdown of YTHDC2 substantially promoted the proliferation rate of ESCC cells by affecting several cancer-related signaling pathways. Our results suggested that rs2416282 contributed to ESCC risk by regulating YTHDC2 expression. This study provided us a valuable insight into the roles of genetic variants in m6A modification genes for ESCC susceptibility and may contribute to the prevention of this disease in the future.
Assuntos
Adenosina/análogos & derivados , Biomarcadores Tumorais/genética , Neoplasias Esofágicas/patologia , Carcinoma de Células Escamosas do Esôfago/patologia , Polimorfismo de Nucleotídeo Único , RNA Helicases/genética , Processamento Pós-Transcricional do RNA , Adenosina/química , Apoptose , Povo Asiático/genética , Estudos de Casos e Controles , Proliferação de Células , China/epidemiologia , Neoplasias Esofágicas/epidemiologia , Neoplasias Esofágicas/genética , Carcinoma de Células Escamosas do Esôfago/epidemiologia , Carcinoma de Células Escamosas do Esôfago/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Prognóstico , RNA Helicases/química , Células Tumorais CultivadasRESUMO
As promising biomarkers and therapy targets, microRNAs (miRNAs) are involved in various physiological and tumorigenic processes. Genetic variants in miRNA-binding sites can lead to dysfunction of miRNAs and contribute to disease. However, systematic investigation of the miRNA-related single nucleotide polymorphisms (SNPs) for pancreatic cancer (PC) risk remains elusive. We performed integrative bioinformatics analyses to select 31 SNPs located in miRNA-target binding sites using the miRNASNP v2.0, a solid database providing miRNA-related SNPs for genetic research, and investigated their associations with risk of PC in two large case-control studies totally including 1847 cases and 5713 controls. We observed that the SNP rs3802266 is significantly associated with increased risk of PC (odds ratio (OR) = 1.21, 95% confidence intervals (CI) = 1.11-1.31, P = 1.29E-05). Following luciferase reporter gene assays show that rs3802266-G creates a stronger binding site for miR-181a-2-3p in 3' untranslated region (3'UTR) of the gene ZHX2. Expression quantitative trait loci (eQTL) analysis suggests that ZHX2 expression is lower in individuals carrying rs3802266-G with increased PC risk. In conclusion, our findings highlight the involvement of miRNA-binding SNPs in PC susceptibility and provide new clues for PC carcinogenesis.
Assuntos
Povo Asiático/genética , Sítios de Ligação/genética , Predisposição Genética para Doença/genética , MicroRNAs/genética , Neoplasias Pancreáticas/genética , Polimorfismo de Nucleotídeo Único/genética , Regiões 3' não Traduzidas/genética , Estudos de Casos e Controles , Linhagem Celular Tumoral , Feminino , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
The N6 -Methyladenosine (m6 A) modification plays an important role in many biological processes, especially tumor development. However, little is still known about how it affects colorectal cancer (CRC) carcinogenesis. Here, we first systematically investigate the association of variants related to m6 A modification with the CRC risk in 1,062 CRC cases and 2,184 controls by using our exome-wide association data and followed by two replication sets including 7,341 CRC cases and 7,902 controls. The variant rs8100241 located in ANKLE1 was significantly associated with CRC risk (odds ratio = 0.88, 95% confidence interval = 0.84-0.92, p = 4.85 × 10-8 ) in 8,403 cases and 10,086 controls. This variant was previously identified to be associated with the susceptibility of breast cancer with BRCA1 mutation triple negative breast cancer. Further functional analysis indicated that overexpression of the rs8100241[A] allele significantly increased the ANKLE1 m6 A level and facilitated the ANKLE1 protein expression compared to that of rs8100241[G] allele. We further found the ANKLE1 m6 A modification was catalyzed by the "writer" complex (METTL3, METTL14, or WTAP) and recognized by the "reader" YTHDF1. Mechanistically, we found that the ANKLE1 functions as a potential tumor suppressor that inhibits cell proliferation and facilitates the genomic stability. An elevated frequency of micronucleated cells, increased cell proliferation, and colony formation ability were observed when ANKLE1 knockdown. Our study illustrated that the germline missense variant can increase CRC risk by influencing ANKLE1 m6 A level, highlighting a clinical potential of variants-associated m6 A modification as a risk marker for CRC prevention.
Assuntos
Adenosina/análogos & derivados , Neoplasias Colorretais/genética , Endonucleases/genética , Predisposição Genética para Doença , Instabilidade Genômica , Adenosina/metabolismo , Idoso , Carcinogênese/genética , Estudos de Casos e Controles , Proliferação de Células/genética , Metilação de DNA , Epigênese Genética , Feminino , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Células HCT116 , Humanos , Masculino , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo ÚnicoRESUMO
Our previous study identified a tag single-nucleotide polymorphism (SNP) rs204900 in TNXB associated with risk of esophageal squamous-cell carcinoma (ESCC) in the Chinese population. However, the functional role of TNXB and causal variants had not been interrogated in that study. In the present study, we explored the effects of TNXB expression in the development of ESCC and searched for functional variants in this gene. We found TNXB was downregulated in ESCC tumors. Using small interfering RNAs and CRISPR-Cas9 methods, we identified that both knockdown and knockout of TNXB significantly promoted ESCC cell growth in vitro, suggesting a tumor suppressor role of this gene in ESCC. Through further fine-mapping analysis, we identified that a noncoding variant in the promoter of TNXB, rs411337, predisposed to ESCC risk (odds ratio = 1.36, 95% confidence interval: 1.22-1.51, P = 9.10 × 10-9 ). These findings revealed the functional mechanism of TNXB in the development of ESCC and may contribute to the prevention and treatment of this disease in the future.
Assuntos
Carcinoma de Células Escamosas/genética , Neoplasias Esofágicas/genética , Predisposição Genética para Doença/genética , Polimorfismo de Nucleotídeo Único , Regiões Promotoras Genéticas/genética , Tenascina/genética , Sistemas CRISPR-Cas , Carcinoma de Células Escamosas/patologia , Linhagem Celular Tumoral , Proliferação de Células/genética , Neoplasias Esofágicas/patologia , Perfilação da Expressão Gênica/métodos , Regulação Neoplásica da Expressão Gênica , Redes Reguladoras de Genes , Humanos , Interferência de RNA , Fatores de RiscoRESUMO
Metabolic reprogramming has been regarded as one of the core hallmarks of cancer and increased de novo fatty acid synthesis has been documented in multiple tumors including esophageal squamous cell carcinoma (ESCC). Our previous exome-wide analyses found a Val1937Ile variant (rs17848945) in the 34th exon of fatty acid synthase (FASN) that showed a strong association with the risk of ESCC. In this study, we performed a series of functional assays to investigate the biological functions underlying this variant in the development of ESCC. We demonstrated that FASN was upregulated in ESCC and both knockdown and knockout of FASN significantly inhibited ESCC cell proliferation, suggesting a tumor promoter role for this gene in ESCC. Furthermore, the results showed that overexpression of FASN[I] in the ESCC cells substantially enhanced cell proliferation, compared with overexpression of FASN[V], or the control vector. Intriguingly, we found that the FASN[I] variant can enhance the enzyme activity of FASN, and, thus, increase the amount of the FASN end-product, palmitate in the ESCC cells. We also observed elevated palmitate levels in the plasma of the FASN[I] genotype carriers among a total of 632 healthy Chinese adults. In conclusion, our results suggested that the FASN V1937I variant influenced ESCC cell proliferation and susceptibility by altering the catabolic activity of FASN on palmitate. These findings may highlight an important role of palmitate metabolism in the development of ESCC and may contribute to the personalized medicine of this disease.
Assuntos
Biomarcadores Tumorais/metabolismo , Metabolismo Energético , Neoplasias Esofágicas/enzimologia , Carcinoma de Células Escamosas do Esôfago/enzimologia , Ácido Graxo Sintase Tipo I/metabolismo , Biomarcadores Tumorais/genética , Linhagem Celular Tumoral , Proliferação de Células , Bases de Dados Genéticas , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/patologia , Carcinoma de Células Escamosas do Esôfago/genética , Carcinoma de Células Escamosas do Esôfago/patologia , Ácido Graxo Sintase Tipo I/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Palmitatos/metabolismo , Polimorfismo de Nucleotídeo Único , Transdução de SinaisRESUMO
GWAS-identified 10q22.3 loci with lead SNP rs704017 are significantly associated with CRC risk in both Asian and European populations. However, the functional mechanism of this region is unclear. In this study, we performed a fine-mapping analysis to identify the causal SNPs. To identify potential functional SNPs in linkage disequilibrium with the lead SNP, we searched for the potential target genes using a Hi-C database and an RNA interfering-based on-chip approach. The results indicated that rs12263636 (r2 = 0.41) showed the highest potential to be functional. It resided in a region with enhancer markers and a topologically associating domain. We found that RPS24 was the only gene that significantly promoted the proliferation rate of CRC cells and might have promoter-enhancer interaction with rs12263636. Dual-luciferase reporter assays confirmed that the risk alleles of two variants (rs3740253 and rs7071351) in RPS24 promoter could increase the expression of luciferase. Case control study consisting of 1134 cases and 2039 health controls confirmed that both the two variants were associated with risk of CRC (rs3740253: P = 0.0079, OR = 1.15, 95% CI 1.04-1.28; rs7071351: P = 0.0085, OR = 1.15, 95% CI 1.04-1.28). And plasmid containing mutant haplotypes containing all the three mutations (rs12263636 or rs3740253 and rs7071351) could most significantly increase luciferase expression, compared with any haplotype of the three mutations. The study explained the functional mechanism for the 10q22.3 loci and provided new insights into the prevention and treatment of CRC.
Assuntos
Neoplasias Colorretais/genética , Polimorfismo de Nucleotídeo Único , Proteínas Ribossômicas/genética , Povo Asiático/genética , Estudos de Casos e Controles , Elementos Facilitadores Genéticos , Regulação Neoplásica da Expressão Gênica , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Humanos , Regiões Promotoras GenéticasRESUMO
As the proper binding of CCCTC-binding factor (CTCF) in the boundaries of topological association domains (TADs) was important for chromatin structures and gene regulation, we hypothesized that single nucleotide polymorphisms (SNPs) affecting CTCF binding in TAD boundaries might contribute to pancreatic cancer (PC) susceptibility. We first genome widely screened out potential SNPs via bioinformatics analysis on Hi-C data, ChIP-seq data, and CTCF binding motif, then tested their associations with PC risk in a previous genome-wide association studies (GWASs) data set (981 cases and 1,991 controls), followed by another independent replication set (1,208 cases and 1,465 controls). Electrophoretic mobility shift assays (EMSAs), expression Quantitative Trait Loci (eQTL) analyses and cell proliferation experiments were performed to uncover the biological mechanisms. The positive SNP rs2001389 was found significantly associated with PC risk with odds ratio (OR) being 1.166 (95% confidence interval (CI) = 1.075-1.264, P = 2.143E-04) in the combined study. The allele G of rs2001389 weakened the binding activity with CTCF, and it was related to the lower expression of a putative antioncogene MFSD13A whose knockdown promoted proliferation of PC cells. By integrating analysis on multiomics data, association studies and functional assays, we proposed that the common variant rs2001389 and the gene MFSD13A might be genetic modifiers of PC tumorigenesis.
Assuntos
Fator de Ligação a CCCTC/genética , Carcinogênese/genética , Proteínas de Membrana/genética , Neoplasias Pancreáticas/genética , Adulto , Idoso , Sítios de Ligação/genética , Proliferação de Células/genética , Cromatina/genética , Proteínas de Ligação a DNA/genética , Regulação Neoplásica da Expressão Gênica , Estudo de Associação Genômica Ampla , Humanos , Masculino , Pessoa de Meia-Idade , Neoplasias Pancreáticas/patologia , Polimorfismo de Nucleotídeo Único/genética , Domínios Proteicos/genética , Locos de Características Quantitativas/genética , Fatores de RiscoRESUMO
BACKGROUND & AIMS: Genome-wide association studies have identified multiple genetic signals associated with the risk of persistent hepatitis B virus (HBV) infection and HBV-related hepatocellular carcinoma. However, the majority of the associated variants may only be markers of functional variants and the underlying biological mechanisms remain elusive. We hypothesized that the functional variants with modulating transcription factor (TF) binding affinity in genome-wide association studies-identified loci may influence the risk of persistent HBV infection in Chinese people. METHODS: A systematic bioinformatics approach was implemented to prioritize potential functional variants that may influence TF binding. A two-stage case-control study, including 1595 HBV-persistent carriers and 1590 subjects with HBV natural clearance, was conducted to examine the associations between candidate variants and susceptibility to persistent HBV infection. Biological assays were carried out to elucidate the underlying mechanism of the associated genetic variants. RESULTS: Twelve candidate variants were identified, and rs2523454 G > A increased the risk of persistent HBV infection (dominant model: ORcombined = 1.37, 95% CI = 1.19-1.58, P = 1.610 × 10-5 ). Functional assays indicated that the rs2523454 A allele significantly decreased transcriptional activity compared to the G allele by influencing TF-binding affinity. In addition, expression quantitative trait loci analyses revealed that the A allele was associated with the reduced expression of MICA (P < 0.01). CONCLUSIONS: Our findings suggest that the germline G > A variation at rs2523454 may influence TF-DNA interaction, downregulate the expression of MICA and play an important role in the development of persistent HBV infection in the Chinese population.
Assuntos
Hepatite B/genética , Antígenos de Histocompatibilidade Classe I/genética , Polimorfismo de Nucleotídeo Único , Adulto , Alelos , Estudos de Casos e Controles , China , Feminino , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Vírus da Hepatite B , Heterozigoto , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
Despite the successes of genome-wide association study (GWAS) in identifying breast cancer (BC) risk-associated variants, only a small fraction of the heritability can be explained. The greatest challenge in the post-GWAS is to identify causal variants and underlying mechanisms responsible for BC susceptibility. In this study, we integrated functional genomic data from ENCODE ChIP-seq, ANNOVAR, and the TRANSFAC matrix to identify potentially regulatory variants with modulating FOXA1-binding affinity across the whole genome, and then conducted a two-stage case-control study including 2164 cases and 2382 controls to investigate the associations between candidate SNPs and BC susceptibility. We identified a BC susceptibility SNP, rs6506689 G>T, with an odds ratio (OR) of 1.23 (95% confidence interval = 1.07-1.40, P = 0.003) under a dominant model in the combined study. Biological assays indicated that the germline G>T variation at rs6506689 creates a FOXA1-binding site and up-regulates the expression of RAB31, thus playing an important role in the development of BC. Our results highlight the importance of regulatory genetic variants in the development of BC by influencing TF-DNA interaction and provide critical insights to pinpoint causal genetic variants.
Assuntos
Neoplasias da Mama/genética , Perfilação da Expressão Gênica/métodos , Estudos de Associação Genética/métodos , Mutação em Linhagem Germinativa , Polimorfismo de Nucleotídeo Único , Proteínas rab de Ligação ao GTP/genética , Sítios de Ligação , Estudos de Casos e Controles , Linhagem Celular Tumoral , Feminino , Predisposição Genética para Doença , Fator 3-alfa Nuclear de Hepatócito/metabolismo , Humanos , Células MCF-7 , Razão de Chances , Análise de Sequência de RNA/métodos , Proteínas rab de Ligação ao GTP/química , Proteínas rab de Ligação ao GTP/metabolismoRESUMO
The gender disparity observed in the incidence of hepatocellular carcinoma (HCC) suggests an important role of estrogens in HCC pathogenesis. In this study, we conducted a case-control study to investigate whether breast cancer risk-associated single nucleotide polymorphisms (SNPs) located at estrogens loci identified by genome-wide association studies (GWASs) also predispose to HCC in a Chinese population. Three candidate SNPs at 6q25.1 were genotyped in 2025 HCC cases and 2032 healthy controls. Differential expression analyses and expression quantitative trait loci (eQTL) analyses were conducted to further explore the potential function of significant SNPs and genes they reside in. Two of the three candidate SNPs (rs9383951 and rs9485372) were observed to be significantly associated with HCC risk. Under a dominant model, the odds ratios (OR) for rs9383951 and rs9485372 were 1.28 (95% CI: 1.10-1.49, P = 0.002) and 1.34 (95% CI: 1.17-1.53, P = 2.75 × 10-5), respectively. We also found a significant accumulative effect of these two SNPs and there was a gradual increase in OR with a greater number of hazard genotypes. Moreover, the association between rs9383951 and HCC risk was specific in males. Lower ESR1 and TAB2 expressions were investigated in hepatic tumor tissues than adjacent normal tissues. We found a significant association between rs9383951 and ESR1 expression (P = 0.047). Besides, ESR1 expression was significantly correlated with the expression of TAB2. Taken together, our study identified two genetic variants at 6q25.1 newly associated with HCC risk, suggesting ESR1 and estrogen signaling may play a role in mediating susceptibility to HCC in Chinese population.
Assuntos
Povo Asiático/genética , Neoplasias da Mama/genética , Carcinoma Hepatocelular/genética , Cromossomos Humanos Par 6/genética , Predisposição Genética para Doença/genética , Neoplasias Hepáticas/genética , Polimorfismo de Nucleotídeo Único/genética , Proteínas Adaptadoras de Transdução de Sinal/genética , Idoso , Estudos de Casos e Controles , Receptor alfa de Estrogênio/genética , Feminino , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Fatores de RiscoRESUMO
The TGF-ß pathway plays an essential role in regulating cell proliferation and differentiation. GWASs and candidate approaches have identified a battery of genetic variants in the TGF-ß pathway contributing to colorectal cancer (CRC). However, most of the significant variants are common variants and their functions remain ambiguous. To identify causal variants with low-frequency in the TGF-ß pathway contributing to CRC susceptibility in Chinese population, we performed targeted sequencing of 12 key genes in TGF-ß signaling in CRC patients followed by a two-stage case-control study with a total of 5109 cases and 5169 controls. Bioinformatic annotations and biochemical experiments were applied to reveal the potential functions of significant variants. Seven low-frequency genetic variants were captured through targeted sequencing. The two stage association studies showed that missense variant rs3764482 (c. 83C>T; p. S28F) in the gene SMAD7 was consistently and significantly associated with CRC risk. Compared with the wild type, the ORs for variant allele were 1.37 (95%CI: 1.10-1.70, P = 0.005), 1.55 (95%CI: 1.30-1.86, P = 1.15 × 106 ), and 1.48 (1.29-1.70, P = 2.44 × 10;8 ) in stage 1, stage 2, and the combined analyses, respectively. Functional annotations revealed that the minor allele T of rs3764482 was more effective than the major allele C in blocking the TGF-ß signaling and inhibiting the phosphorylation of receptor-regulated SMADs (R-SMADs). In conclusion, low-frequency coding variant rs3764482 in SMAD7 is associated with CRC risk in Chinese population. The rs3764482 variant may block the TGF-ß signaling via impeding the activation of downstream genes, leading to cancer cell proliferation, thus contributing to CRC pathogenesis.
Assuntos
Neoplasias Colorretais/genética , Regulação Neoplásica da Expressão Gênica , Polimorfismo de Nucleotídeo Único/genética , Proteína Smad7/genética , Fator de Crescimento Transformador beta/metabolismo , Adulto , Idoso , Biomarcadores Tumorais , Estudos de Casos e Controles , China/epidemiologia , Neoplasias Colorretais/epidemiologia , Feminino , Seguimentos , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Fosforilação , Prognóstico , Fatores de Risco , Transdução de Sinais , Fator de Crescimento Transformador beta/genéticaRESUMO
BACKGROUND: Pancreatic adenocarcinoma (PAAD) is an extremely lethal malignancy. Identification of the functional genes and genetic variants related to PAAD prognosis is important and challenging. Previously identified prognostic genes from several expression profile analyses were inconsistent. The regulatory genetic variants that affect PAAD prognosis were largely unknown. METHODS: Firstly, a meta-analysis was performed with seven published datasets to systematically explore the candidate prognostic genes for PAAD. Next, to identify the regulatory variants for those candidate genes, expression quantitative trait loci analysis was implemented with PAAD data resources from The Cancer Genome Atlas. Then, a two-stage association study in a total of 893 PAAD patients was conducted to interrogate the regulatory variants and find the prognostic locus. Finally, a series of biochemical experiments and phenotype assays were carried out to demonstrate the biological function of variation and genes in PAAD progression process. RESULTS: A total of 128 genes were identified associated with the PAAD prognosis in the meta-analysis. Fourteen regulatory loci in 12 of the 128 genes were discovered, among which, only rs4887783, the functional variant in the promoter of Ring Finger and WD Repeat Domain 3 ( RFWD3 ), presented significant association with PAAD prognosis in both stages of the population study. Dual-luciferase reporter and electrophoretic mobility shift assays demonstrated that rs4887783-G allele, which predicts the worse prognosis, enhanced the binding of transcript factor REST, thus elevating RFWD3 expression. Further phenotypic assays revealed that excess expression of RFWD3 promoted tumor cell migration without affecting their proliferation rate. RFWD3 was highly expressed in PAAD and might orchestrate the genes in the DNA repair process. CONCLUSIONS: RFWD3 and its regulatory variant are novel genetic factors for PAAD prognosis.
Assuntos
Adenocarcinoma , Neoplasias Pancreáticas , Adenocarcinoma/genética , Biomarcadores Tumorais/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Pancreáticas/patologia , Prognóstico , Locos de Características Quantitativas/genética , Ubiquitina-Proteína Ligases/genética , Repetições WD40 , Neoplasias PancreáticasRESUMO
AIM: This study aimed to identify the functional genes and genetic variants associated with the prognosis of pancreatic ductal adenocarcinoma (PDAC) and reveal the mechanism underlying their prognostic roles. METHODS: First, we implement a two-stage exome-wide association study in a total of 1070 patients to identify the genetic variant correlated with PDAC prognosis. Then we performed fine mapping through bioinformatics analysis and dual-luciferase reporter assays to reveal the causal functional variant and prognostic gene. Next, we established the gene knockdown, knockout, and overexpression cell lines with small interfering RNA, CRISPR/Cas9, and lentivirus, respectively, and investigated the gene function on cell proliferation and migration in vivo and in vitro. Finally, we performed the RNA-seq to elucidate downstream genes and mechanisms altering PDAC prognosis. RESULTS: We identified the CAV1-CAV2 locus tagged by rs8940 was significantly associated with PDAC prognosis, and rs10249656 in the 3'untranslated region of CAV2 was the real functional variant, which upregulated CAV2 expression through abolishing miR-548s binding. We observed upregulated CAV2 in PDAC and the higher expression correlated with worse prognosis. Transient knockdown of CAV2 inhibited PDAC migration without affecting proliferation rate. Knockout of CAV2 suppressed PDAC progression and metastasis, whereas stable overexpression of CAV2 promoted. Overexpressed CAV2 promoted the PDAC progression and metastasis via perturbing genes in the focal adhesion (CCND1, IGTA1, and ZYX) and extracellular matrix organisation (PLOD2, CAST, and ITGA1) pathways mechanically. CONCLUSION: These findings shed light on an important role of CAV2 on PDAC progression and the prognostic impact of its genetic variation.
Assuntos
Carcinoma Ductal Pancreático/metabolismo , Caveolina 2/metabolismo , Movimento Celular , Neoplasias Pulmonares/metabolismo , Neoplasias Pancreáticas/metabolismo , Polimorfismo de Nucleotídeo Único , Animais , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/secundário , Caveolina 2/genética , Linhagem Celular Tumoral , Progressão da Doença , Matriz Extracelular/genética , Matriz Extracelular/metabolismo , Matriz Extracelular/patologia , Adesões Focais/genética , Adesões Focais/metabolismo , Adesões Focais/patologia , Regulação Neoplásica da Expressão Gênica , Estudos de Associação Genética , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/secundário , Camundongos Endogâmicos BALB C , Camundongos Nus , MicroRNAs/genética , MicroRNAs/metabolismo , Invasividade Neoplásica , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , RNA-Seq , Transdução de Sinais , Sequenciamento do ExomaRESUMO
The activating transcription factor 1 (ATF1) has been identified as a vital pathogenic factor in the progression of colorectal cancer (CRC), whiles, the precise regulatory mechanisms remain elusive. Here, we comprehensively characterized the ATF1 cistrome by RNA-seq and ChIP-seq assays in CRC cell lines. As the results, we identified 358 genes differentially regulated and 15,029 ATF1 binding sites and demonstrated that ATF1 was widely involved in major signaling pathways in CRC, such as Wnt, TNF, Jak-STAT. Subsequently, by the expression quantitative trait loci (eQTL) analyses, we found that rs7017386 was associated with the expression of CCAT1 and PVT1 in the Wnt pathway. By a two-stage population study with 6,131 CRC cases and 10,022 healthy controls, we identified the variant was associated with CRC risk. Mechanistically, we found rs7017386 allele-specifically enhanced the binding affinity of ATF1 and promoted the expressions of PVT1 and CCAT1, via forming a long-range chromatin loop. Moreover, those two lncRNAs could synergistically facilitate c-Myc expression to activate the Wnt pathway in CRC progression. Our findings not only demonstrated the transcriptomic profiling of ATF1 in CRC, but also provided important clues for the etiology of CRC.
Assuntos
Fator 1 Ativador da Transcrição/genética , Cromatina/genética , Neoplasias Colorretais/genética , Variação Genética/genética , Oncogenes/genética , RNA Longo não Codificante/genética , Carcinogênese/genética , Linhagem Celular Tumoral , Perfilação da Expressão Gênica/métodos , Células HCT116 , Humanos , Transdução de Sinais/genética , Via de Sinalização Wnt/genéticaRESUMO
Although genome-wide association studies (GWAS) have identified more than 100 colorectal cancer risk loci, most of the biological mechanisms associated with these loci remain unclear. Here we first performed a comprehensive expression quantitative trait loci analysis in colorectal cancer tissues adjusted for multiple confounders to test the determinants of germline variants in established GWAS susceptibility loci on mRNA and long noncoding RNA (lncRNA) expression. Combining integrative functional genomic/epigenomic analyses and a large-scale population study consisting of 6,024 cases and 10,022 controls, we then prioritized rs174575 with a C>G change as a potential causal candidate for colorectal cancer at 11q12.2, as its G allele was associated with an increased risk of colorectal cancer (OR = 1.26; 95% confidence interval = 1.17-1.36; P = 2.57 × 10-9). rs174575 acted as an allele-specific enhancer to distally facilitate expression of both FADS2 and lncRNA AP002754.2 via long-range enhancer-promoter interaction loops, which were mediated by E2F1. AP002754.2 further activated a transcriptional activator that upregulated FADS2 expression. FADS2, in turn, was overexpressed in colorectal cancer tumor tissues and functioned as a potential oncogene that facilitated colorectal cancer cell proliferation and xenograft growth in vitro and in vivo by increasing the metabolism of PGE2, an oncogenic molecule involved in colorectal cancer tumorigenesis. Our findings represent a novel mechanism by which a noncoding variant can facilitate long-range genome interactions to modulate the expression of multiple genes including not only mRNA, but also lncRNA, which provides new insights into the understanding of colorectal cancer etiology. SIGNIFICANCE: This study provides an oncogenic regulatory circuit among several oncogenes including E2F1, FADS2, and AP002754.2 underlying the association of rs174575 with colorectal cancer risk, which is driven by long-range enhancer-promoter interaction loops. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/80/9/1804/F1.large.jpg.