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Extracellular proteases of haloarchaea are adapted to function at high concentrations of NaCl, and could find useful applications in industrial or biotechnological processes where hypersaline conditions are desired. The diversity of extracellular proteases produced by haloarchaea is largely unknown, although the genomes of many species have been sequenced and are publicly available. In this study, a gene encoding the extracellular protease Hly176B from the haloarchaeon Haloarchaeobius sp. FL176 was cloned and expressed in Escherichia coli. The catalytic triad Asp-His-Ser was confirmed via site-directed mutagenesis, indicating that Hly176B belongs to the class of serine proteases (halolysin). Unlike previously reported extracellular proteases from haloarchaea, Hly176B remained active in almost salt-free solution for a relatively long time. In addition, Hly176B displayed good tolerance to some metal ions, surfactants and organic solvents, and exerts its highest enzyme activities at 40 °C, pH 8.0 and 0.5 M NaCl. Therefore, this study enriches our knowledge of extracellular proteases, and also expands their applications in various industrial uses.

Genome-wide association studies (GWAS) of colorectal cancer have identified several common susceptible variants in gene regulatory regions. However, low-frequency or rare coding risk variants have not been systematically investigated in patients with colorectal cancer from Chinese populations. In this study, we performed an exome-wide association analysis with 1,062 patients with colorectal cancer and 2,184 controls from a Chinese population. Promising associations were further replicated in two replication sets: replication stage I with 2,478 cases and 3,880 controls, and replication stage II with 3,761 cases and 4,058 controls. We identified two variants significantly associated with colorectal cancer risk: a novel rare missense variant in TCF7L2 [rs138649767, OR = 2.08, 95% confidence interval (CI): 1.69–2.57, P = 5.66 × 10−12] and a previous European GWAS-identified 3′-UTR variant in ATF1 (rs11169571, OR = 1.18, 95% CI: 1.13–1.24, P = 1.65 × 10−12). We found a significant interaction between the TCF7L2 missense variant rs138649767 and a previous GWAS-identified regulatory variant rs6983267 in the MYC enhancer (Pinteraction = 0.0002). Functional analysis of this variant revealed that TCF7L2 with rs138649767-A allele harbored the ability to activate the MYC enhancer with rs6983267-G allele and enhance colorectal cancer cell proliferation. In addition, the ATF1 rs11169571 variant significantly correlated with ATF1 expression by affecting hsa-miR-1283 and hsa-miR-520d-5p binding. Further ChIP-seq and gene coexpression analyses showed that oncogenes NRAS and BRAF were activated by ATF1 in colorectal cancer. These results widen our understanding of the molecular basis of colorectal cancer risk and provide insight into pathways that might be targeted to prevent colorectal cancer.Significance: Exome-wide association analysis identifies a rare missense variant in TCF7L2 and a common regulatory variant in ATF1 as susceptibility factors of colorectal cancer.Graphical Abstract: http://cancerres.aacrjournals.org/content/canres/78/17/5164/F1.large.jpg. Cancer Res; 78(17); 5164–72. ©2018 AACR.

Background: Genome-wide association studies (GWAS) have identified dozens of loci associated with colon and rectal adenocarcinoma risk. As tissue-specific super-enhancers (SE) play important roles in tumorigenesis, we systematically investigate SEs and inner variants in established GWAS loci to decipher the underlying biological mechanisms. Methods: Through a comprehensive bioinformatics analysis on multi-omics data, we screen potential single-nucleotide polymorphisms (SNP) in cancer-specific SEs, and then subject them to a two-stage case–control study containing 4,929 cases and 7,083 controls from the Chinese population. A series of functional assays, including reporter gene assays, electrophoretic mobility shift assays (EMSA), CRISPR–Cas9 genome editing, chromosome conformation capture (3C) assays, and cell proliferation experiments, are performed to characterize the variant's molecular consequence and target genes. Results: The SNP rs11064124 in 12p13.31 is found significantly associated with the risk of colon and rectal adenocarcinoma with an odds ratio (OR) of 0.87 [95% confidence interval (CI), 0.82–0.92, P = 8.67E-06]. The protective rs11064124-G weakens the binding affinity with vitamin D receptor (VDR) and increases the enhancer's activity and interactions with two target genes' promoters, thus coactivating the transcription of CD9 and PLEKHG6, which are both putative tumor suppressor genes for colon and rectal adenocarcinoma. Conclusions: Our integrative study highlights an SE polymorphism rs11064124 and two susceptibility genes CD9 and PLEKHG6 in 12p13.31 for colon and rectal adenocarcinoma. Impact: These findings suggest a novel insight for genetic pathogenesis of colon and rectal adenocarcinoma, involving transcriptional coactivation of diverse susceptibility genes via the SE element as a gene regulation hub.

The high and low-temperature performance of five hydraulic bitumen binders was evaluated using the dynamic shear rheometer (DSR) test, infrared spectrum test and direct tensile (DT) test. These hydraulic bitumen binders were respectively applied for several pumped storage power stations (PSPS) projects that were constructed or under construction. In order to relate the bitumen performance to the mixture performance, the slope flow test, three-point bending test and thermal stress restrained specimen test were carried out on hydraulic asphalt mixtures. The test results indicated the DSR rheological master curves can well distinguish the difference of each bitumen binder as well as the effect of polymer modification. Phase angle master curves, black diagrams and infrared spectra all indicated that several penetration-grade hydraulic bitumen binders were not virgin bitumen binders but were modified with relatively lower SBS polymer content when compared with traditional SBS-modified bitumen. When selecting the commonly used Karamay SG70 hydraulic bitumen as a reference, the normal SBS-modified bitumen was superior to other bitumen in terms of low- and high-temperature performance. Several slightly SBS-modified bitumen binders did not always show consistent results, which indicated that slightly modified bitumen may not really have the desired performance as expected. Therefore, SBS-modified bitumen will be more promising when dealing with extremely low or high temperatures. Bitumen performance was well compared with the mixture performance by using the bitumen creep, relaxation and tensile failure strain corresponding to the asphalt concrete slope flow, the maximum bending strain and the failure temperature, respectively. Compared with the traditional penetration, softening point and ductility test, it indicated that the DSR rheological test, creep test, direct tensile test and stress relaxation test can be used as more powerful tools for the characterization and optimization of hydraulic bitumen binders.

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.

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.

Interpreting disease-causing variants, especially in noncoding regions by genome-wide association studies (GWAS), has become one of the most challenging and demanding tasks. We hypothesized that functional lncRNAs variants in GWAS-identified loci might alter expression level of genes associated with persistent HBV infection and hepatocellular carcinoma (HCC). Integrated bioinformatics approaches were used to prioritize potentially functional variants and a two-stage case–control study (2473 HBV positive HCC patients, 2248 persistent HBV carriers and 2294 spontaneously recovered subjects) was performed to assess the roles of these variants. The rs2844512 G > C variant in LINC01149 was identified to facilitate HBV spontaneous recovery (OR = 0.84, 95% CI = 0.77–0.92) but increase the risk of HCC (OR = 1.21, 95% CI = 1.11–1.32) in combined samples. Subsequent biological assays indicated this variant created a binding site for miR-128-3p and upregulated MICA expression by serving as a miRNA sponge, which might recruit NK-cells to lyse infected cells, but release highly soluble MICA by shedding to induce NK-cells exhaustion and tumor immune evasion. These findings highlight a regulatory circuit between LINC01149 and MICA, mediating by miR-128-3p, and the important role of upregulated MICA in conferring susceptibility to persistent HBV infection and HCC.

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.

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.

The SLC10A1 Ser267Phe (S267F) variant has been reported to severely inhibit hepatitis B virus (HBV) infection and taurocholate transport activity. This study aimed to clarify the effects of this variant on HBV infection and bile acid metabolism. SLC10A1 S267F was genotyped in 2907 HBV-exposed subjects (including HBV persistent carriers and spontaneously recovered subjects) and 1364 unexposed subjects (HBV marker-negative subjects), followed by replication I, comprising 914 exposed subjects and 1123 unexposed subjects, and replication II, comprising 355 children born to HBsAg-positive mothers (226 HBV-infected children and 129 controls). Intriguingly, SLC10A1 AA was observed only in the unexposed group, but not in the exposed group. The SLC10A1 A allele consistently decreased HBV infection risk compared with the G allele (OR = 0.76, 95% CI: 0.64-0.90 in combined samples). In addition, children with the SLC10A1 GA genotype had a reduced risk of perinatal transmission (OR = 0.31, 95% CI: 0.14-0.71). Moreover, unexposed subjects with the SLC10A1 AA genotype exhibited decreased serum total cholesterol and low-density lipoprotein cholesterol compared to those with the GG or GA genotypes (P = 2.975 × 10-4 and 0.004, respectively). The study highlighted the role of the SLC10A1 S267F variant in the loss of the ability to support HBV infection and taurocholate transport activity. Subjects with the AA genotype may escape from HBV infection and present decreased cholesterol levels as a consequence of impaired bile acid uptake.

Pancreatic cancer is one of the deadliest malignancies with few early detection tests or effective therapies. PI3K‐AKT signaling is recognized to modulate cancer progression. We previously identified that a genetic variant in PKN1 increased pancreatic cancer risk through the PKN1/FAK/PI3K/AKT pathway. In order to investigate the associations between genetic variations in that pathway and pancreatic cancer prognosis, we conducted a two‐stage survival analysis in a total of 547 Chinese pancreatic cancer patients. Consequently, a variant, rs13167294 A>C in PIK3R1, was significantly associated with poor survival in both stages and with hazard ratio being 1.32 (95% CI = 1.13‐1.56, P = 0.0007) in the combined analysis. Function annotation and prediction suggested that genetic variants in this locus might affect overall survival of pancreatic cancer patients by regulating PIK3R1 expression.

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); P(multiplicative-interaction) = 1.20 × 10(−22); P(additive-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-β, 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.

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.

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.

Background: Telomere dysfunction triggers cellular senescence and constitutes a driving force for cancer initiation. Genetic variants in genes involved in telomere maintenance may contribute to colorectal cancer susceptibility. Methods: In this study, we firstly captured germline mutations in 192 patients with colorectal cancer by sequencing the coding regions of 13 core components implicated in telomere biology. Five potential functional variants were then genotyped and assessed in a case–control set with 3,761 colorectal cancer cases and 3,839 healthy controls. The promising association was replicated in additional 6,765 cases and 6,906 controls. Functional experiments were used to further clarify the potential function of the significant variant and uncover the underlying mechanism in colorectal cancer development. Results: The two-stage association studies showed that a rare missense variant rs149418249 (c.C1520T and p.P507L) in the 11th exon of TPP1 (also known as ACD, gene ID 65057) was significantly associated with colorectal cancer risk with the ORs being 2.90 [95% confidence interval (CI), 1.04–8.07; P = 0.041], 2.50 (95% CI, 1.04–6.04; P = 0.042), and 2.66 (95% CI, 1.36–5.18; P = 0.004) in discovery, replication, and the combined samples, respectively. Further functional annotation indicated that the TPP1 P507L substitution interrupted TPP1–TIN2 interaction, impaired telomerase processivity, and shortened telomere length, which subsequently facilitated cell proliferation and promoted colorectal cancer development. Conclusions: A rare variant P507L in TPP1 confers increased risk of colorectal cancer through interrupting TPP1–TIN2 interaction, impairing telomerase processivity, and shrinking telomere length. Impact: These findings emphasize the important role of telomere dysfunction in colorectal cancer development, and provide new insights about the prevention of this type of cancer. Cancer Epidemiol Biomarkers Prev; 27(9); 1029–35. ©2018 AACR.

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.

Multiple single nucleotide polymorphisms (SNPs) have been found to be highly correlated with colorectal cancer (CRC) risk. However, the variants identified thus far only explain a small part of the cases, suggesting the existence of many uncharacterised genetic determinants. In this study, using the multilevel 'omics' data provided in The Cancer Genome Atlas, we systematically performed expression quantitative trait locus (eQTL) analysis for CRC and identified nine SNPs with significant effects on mRNA expression (correlation |r| > 0.3 and FDR

Background MYC is a well-established cancer driver gene regulating the expression of numerous genes, indicating that polymorphisms in MYC response elements could affect tumorigenesis through altering MYC regulation. We performed integrative multistage study to evaluate the effects of variants in MYC response elements and colorectal cancer (CRC) risk. Patients and methods We systematically integrated ChIP-Seq, DNase-Seq and transcription factor motif data to screen variants with potential ability to affect the MYC binding affinity. Then, we conducted a two-stage case–control study, totally consisting of 4830 CRC cases and 4759 controls in Chinese population to identify risk polymorphisms and interactions. The effects of risk variants were confirmed by functional assays in CRC LoVo, SW480 and HCT15 cells. Results We identified a novel polymorphism rs11777210 in KBTBD11 significantly associated with CRC susceptibility (P = 2.43 × 10−12). Notably, we observed a significant interaction between rs11777210 and MYC nearby rs6983267 (P-multi = 0.003, P-add = 0.005), subjects carrying rs6983267 GG and rs11777210 CC genotypes showing higher susceptibility to CRC (2.83-fold) than those carrying rs6983267 TT and rs11777210 TT genotypes. We further demonstrated that rs6983267 T > G increased MYC expression, and MYC bound to and negatively regulated KBTBD11 expression when the rs11777210 C risk allele was present. KBTBD11 was downregulated in tumor tissues, and KBTBD11 knockdown promoted cell proliferation and inhibited cell apoptosis. Conclusion The rs11777210 is a potential predictive biomarker of CRC susceptibility, and KBTBD11 functions as a putative tumor suppressor in tumorigenesis. Our study highlighted the high CRC risk of people carrying rs6983267 G and rs11777210 C alleles, and provided possible biological mechanism of the interaction.

Genome-wide association studies have identified common variants associated with risk of esophageal squamous cell carcinoma (ESCC). However, these common variants cannot explain all heritability of ESCC. Here we report an exome-wide interrogation of 3,714 individuals with ESCC and 3,880 controls for low-frequency susceptibility loci, with two independent replication samples comprising 7,002 cases and 8,757 controls. We found six new susceptibility loci in CCHCR1, TCN2, TNXB, LTA, CYP26B1 and FASN (P = 7.77 × 10−24 to P = 1.49 × 10−11), and three low-frequency variants had relatively high effect size (odds ratio > 1.5). Individuals with the rs138478634-GA genotype had significantly lower levels of serum all-trans retinoic acid, an anticancer nutrient, than those with the rs138478634-GG genotype (P = 0.0004), most likely due to an enhanced capacity of variant CYP26B1 to catabolize this agent. These findings emphasize the important role of rare coding variants in the development of ESCC. Exome-wide analyses identify low-frequency coding variants associated with esophageal squamous cell carcinoma. One of the risk variants, in CYP26B1, is associated with enhanced enzymatic activity and lower levels of all-trans retinoic acid in serum.

// Jiaoyuan Li 1 , Hui Liu 2 , Li Zou 3 , Juntao Ke 1 , Yi Zhang 1 , Ying Zhu 1 , Yang Yang 1 , Yajie Gong 1 , Jianbo Tian 1 , Danyi Zou 1 , Xiating Peng 1 , Jing Gong 1 , Rong Zhong 1 , Kun Huang 4 , Jiang Chang 5, * and Xiaoping Miao 1, * 1 Department of Epidemiology and Biostatistics, School of Public Health, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China 2 National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, Wuhan, China 3 Department of Health Care, Bao’an Maternal and Child Health Hospital, Shenzhen, China 4 Tongji School of Pharmacy, Huazhong University of Science and Technology, Wuhan, China 5 Key Laboratory of Environment and Health, Ministry of Education & Ministry of Environmental Protection, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China * These authors have contributed equally to this work Correspondence to: Xiaoping Miao, email: miaoxp@mail.hust.edu.cn Jiang Chang, email: changjiang815@hust.edu.cn Keywords: TGF-β, GREM1, untranslated region, microRNA, colorectal cancer Received: February 25, 2017 Accepted: April 26, 2017 Published: May 23, 2017 ABSTRACT The transforming growth factor beta (TGF-β) pathway has been implicated in carcinogenesis of intestinal canal. Except for common variants indentified by genome-wide association studies, variants with lower frequency can also explain a part of the disease heritability, especially those in gene regulatory regions. In this study, we searched for colorectal cancer (CRC) related functional low-frequency variants (minor allele frequency 1-5%) in untranslated regions (UTR) involved in the TGF-β signaling using a next-generation sequencing based approach. A case-control study including 1,841 CRC cases and 1,837 controls was performed to identify CRC associated variants and biological experiments were applied to further explore the potential functions of the significant variants. Three low-frequency UTR variants were selected as our candidates and subsequent association analyses showed that a low-frequency variant rs12915554 in the 3’ UTR of GREM1 was significantly associated with CRC risk (Additive model: OR=1.43, 95%CI: 1.04-1.95, P =0.026). Functional annotations suggested that rs12915554 variation increased the expression of GREM1 by perturbing a hsa-miR-185-3p binding site. Moreover, higher expression level of GREM1 was investigated in colon tumor tissues compared with adjacent normal tissues using TCGA data. In conclusion, low-frequency UTR variant rs12915554 in the gene GREM1 was in relation to CRC susceptibility in a Chinese population and this variation might promote CRC development through enhancing GREM1 expression in a miRNA-mediated posttranscriptional manner.

Genome-wide association studies (GWASs) have identified dozens of susceptibility loci for colorectal cancer (CRC). However, most of them lack functional genetic variants and clear biological mechanisms. MicroRNAs (miRNAs) are small noncoding RNAs involved in a variety of physiological and tumorigenic processes. Here we hypothesized that single nucleotide polymorphisms (SNPs) that affect miRNAs biogenesis and binding, could contribute to CRC risk in the Chinese population. To locate miRNA-related SNPs in established GWAS loci, we initially screened out five candidate SNPs using a systematic bioinformatics analysis. Then, we performed a two-stage case-control study consisting of 2347 cases and 3390 controls, and found a positive polymorphism rs1062044, which presented consistently significant associations with CRC in both stages, and with an odds ratio (OR) = 1.32 (95% confidence interval (95%CI) = 1.18-1.49, P = 3.43E-06) under the dominant model in the combined study. Further luciferase reporter gene assays indicated that the variant G allele obviously improved the specific binding between miR-423-5p and the gene LAMC1. These findings suggested that the functional SNP rs1062044 at 1q25.3 might be a genetic modifier for the occurrence and development of CRC.

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.

// Ying Zhu 1, * , Kan Zhai 2, * , Juntao Ke 1 , Jiaoyuan Li 1 , Yajie Gong 1 , Yang Yang 1 , Jianbo Tian 1 , Yi Zhang 1 , Danyi Zou 1 , Xiating Peng 1 , Jing Gong 1 , Rong Zhong 1 , Kun Huang 3 , Jiang Chang 1 , Xiaoping Miao 1 1 Key Laboratory of Environment and Health, Ministry of Education and Ministry of Environmental Protection, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China 2 Medical Research Center, Beijing Chao-Yang Hospital, Capital Medical University, Beijing, China 3 Tongji School of Pharmacy, Huazhong University of Science and Technology, Wuhan, China * These authors contribute equally to this work Correspondence to: Jiang Chang, email: changjiang815@hust.edu.cn Xiaoping Miao, email: Miaoxp@hust.edu.cn Keywords: prognosis, pancreatic cancer, BRCA, SNP Received: February 09, 2017 Accepted: March 15, 2017 Published: March 21, 2017 ABSTRACT Pancreatic cancer is a highly lethal disease with limited prognostic marker. BRAC1 and BRCA2 are two classic tumor suppressor genes which play an important role in DNA repair. Somatic mutations and germline genetic variants on BRCA1/2 have been found associated with the tumorigenesis of pancreatic cancer. However, the correlations between BRCA1/2 polymorphism and pancreatic cancer prognosis remained unknown. In this study, we genotyped three tag missense variants on BRCA1/2 in 603 sporadic pancreatic cancer patients in a Chinese population. We found rs1799966 on BRCA1 was associated with poor prognosis of pancreatic cancer patients with hazard ratio being 1.23 (95% CI: 1.09–1.40, P = 0.0010). Further stratification analyses showed that significant correlation was particularly in locally advanced stage patients with hazard ratio being 1.36 (95% CI: 1.13–1.64, P = 0.0014), but not in patients in local stage ( P = 0.1139) or metastatic stage ( P = 0.5185). Two missense variants (rs766173 and rs144848) on BRAC2 showed no significant correlation with pancreatic cancer patients’ overall survival. In conclusion, we identified a germline missense variant on BRAC1 significantly associated with poor prognosis of pancreatic cancer patients with locally advanced stage. These results may contribute to the precision medicine of this disease.

Genome-wide association studies (GWAS) have identified dozens of loci associated with colon and rectal adenocarcinoma risk. As tissue-specific super-enhancers (SE) play important roles in tumorigenesis, we systematically investigate SEs and inner variants in established GWAS loci to decipher the underlying biological mechanisms.Through a comprehensive bioinformatics analysis on multi-omics data, we screen potential single-nucleotide polymorphisms (SNP) in cancer-specific SEs, and then subject them to a two-stage case-control study containing 4,929 cases and 7,083 controls from the Chinese population. A series of functional assays, including reporter gene assays, electrophoretic mobility shift assays (EMSA), CRISPR-Cas9 genome editing, chromosome conformation capture (3C) assays, and cell proliferation experiments, are performed to characterize the variant's molecular consequence and target genes.The SNP rs11064124 in 12p13.31 is found significantly associated with the risk of colon and rectal adenocarcinoma with an odds ratio (OR) of 0.87 [95% confidence interval (CI), 0.82-0.92,Our integrative study highlights an SE polymorphism rs11064124 and two susceptibility genesThese findings suggest a novel insight for genetic pathogenesis of colon and rectal adenocarcinoma, involving transcriptional coactivation of diverse susceptibility genes via the SE element as a gene regulation hub.

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.

The N

// Yi Zhang 1, 2 , Yan Li 2 , Shaojie Jiang 3 , Wei Chen 1, 4 , Jiao Lou 1 , Juntao Ke 1 , Jiaoyuan Li 1 , Ying Zhu 1 , Yajie Gong 1 , Yang Yang 1 , Jianbo Tian 1 , Xiating Peng 1 , Danyi Zou 1 , Jing Gong 1 , Jiang Chang 1 , Xiaoping Miao 1 , Rong Zhong 1 1 Department of Epidemiology and Biostatistics and State Key Laboratory of Environment Health (Incubation), Ministry of Education Key Laboratory of Environment & Health, Ministry of Environmental Protection Key Laboratory of Environment and Health (Wuhan), School of Public Health, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, PR China 2 Department of Preventive Medicine, School of Public Health, Zunyi Medical University, Zunyi, Guizhou, China 3 Department of Gastroenterology, Jing Zhou Central Hospital, Jingzhou, China 4 Zhuhai Center for Chronic Disease Control, Zhuhai, China Correspondence to: Rong Zhong, email: rongzhong91@gmail.com Xiaoping Miao, email: miaoxp@mail.hust.edu.cn Keywords: breast cancer, FOXA1, genetic variants, susceptibility, case-control study Received: August 20, 2016 Accepted: October 19, 2016 Published: November 07, 2016 ABSTRACT The DNA-binding protein FOXA1 has been shown to regulate nearly all estrogen receptor-chromatin interactions, thereby influencing target gene expression levels in breast cancer (BC) cells. Recently, the rs4442975 T-allele, which disrupts the recruitment of FOXA1 and interacts with the IGFBP5 promoter, was associated to BC susceptibility in a European population. We conducted a hospital-based case-control study that included 1227 cases and 1285 controls to explore the potential association between rs4442975 and BC risk in Chinese Han population, and the effect of this SNP on BC progression was also observed in cases. No significant associations between rs4442975 and BC risk were observed under any genetic models, with an odds ratio of 0.96 (95% confidence interval = 0.81-1.15) under the additive model. When stratified based on estrogen or progesterone receptor expression, smoking or drinking habits, or menopausal status, similar negative associations were observed for all subgroups. No significant associations were observed between rs4442975 and traditional progression factors such as tumor size, nodal status, distant metastasis, or TNM staging. These results reveal that rs4442975 may not confer a risk of BC occurrence or progression in the Chinese Han population, which indicates a distinct association related to genetic heterogeneity across ethnic populations.

Biliary atresia (BA) is a major neonatal cholestatic disease and main indication for pediatric liver transplantation in the world. Recently, GPC1 has been implicated as a risk gene for BA by genetic studies and follow-up functional experiments on zebrafish.Two common genetic variants of GPC1, rs2292832 and rs3828336, were selected systematically through 'SNPinfo', and were examined using TaqMan Genotyping Assays for association studies in a Chinese population containing 134 cases and 618 controls.Of the two single nucleotide polymorphisms (SNPs), we found a significantly decreased BA risk associated with rs2292832 (additive model: OR=0.638, 95% CI: 0.467-0.873, P=0.005), and a marginal effect for rs3828336 (heterozygous model: OR=0.564, 95% CI: 0.312-1.020, P=0.058). The haplotype analysis indicated that either Crs2292832-Crs3828336Trs3828336 or Trs2292832-Trs3828336 conferred a protective effect from BA (OR=0.569, 95% CI=0.414-0.783, P0.001; OR=0.528, 95% CI: 0.301-0.926, P=0.026). Moreover, bioinformatics analysis suggested that rs2292832 altered GPC1 expression via effect on transcription-factor-binding sites (TFBS) of upstream binding transcription factor (UBTF), as a regulatory DNA variation in Deoxyribonuclease I (DNase I) hypersensitive sites (DHSs).Common variants of GPC1 gene were genetically involved in BA risk.