Structure of Vibrio Phage XM1, a Simple Contractile DNA Injection Machine.

Antibiotic resistance poses a growing risk to public health, requiring new tools to combat pathogenic bacteria. Contractile injection systems, including bacteriophage tails, pyocins, and bacterial type VI secretion systems, can efficiently penetrate cell envelopes and become potential antibacterial agents. Bacteriophage XM1 is a dsDNA virus belonging to the Myoviridae family and infecting Vibrio bacteria. The XM1 virion, made of 18 different proteins, consists of an icosahedral head and a contractile tail, terminated with a baseplate. Here, we report cryo-EM reconstructions of all components of the XM1 virion and describe the atomic structures of 14 XM1 proteins. The XM1 baseplate is composed of a central hub surrounded by six wedge modules to which twelve spikes are attached. The XM1 tail contains a fewer number of smaller proteins compared to other reported phage baseplates, depicting the minimum requirements for building an effective cell-envelope-penetrating machine. We describe the tail sheath structure in the pre-infection and post-infection states and its conformational changes during infection. In addition, we report, for the first time, the in situ structure of the phage neck region to near-atomic resolution. Based on these structures, we propose mechanisms of virus assembly and infection.

Surveillance of the Antimicrobial Susceptibility and Molecular Characteristics of Neisseria gonorrhoeae Isolates Collected in Changsha, China from 2016 to 2021.

Antibiotic treatment is critical for individuals infected with gonorrhea and preventing disease transmission. This study aimed to analyze the antimicrobial susceptibility and molecular epidemiological characteristics of Neisseria gonorrhoeae isolates in Changsha, China. A total of 271 N.gonorrhoeae isolates collected from the clinical laboratories of two hospitals between 2016 and 2021 were analyzed for antimicrobial susceptibility using the agar dilution method. N. gonorrhoeae multi-antigen sequence typing (NG-MAST) was conducted for genotyping, and phylogenetic analysis was performed using the porB and tbpB sequences. The results showed that antimicrobial resistance against ciprofloxacin, tetracycline, and penicillin was high, and these drugs are no longer recommended for the treatment of gonorrhea. All isolates were susceptible to spectinomycin. However, in 2016-2021, a total of 15 (5.5%) ceftriaxone (CRO)-resistant strains and 31 (11.4%) isolates with decreased susceptibility to CRO were found, and the resistance rate to azithromycin had reached 7.1% in 2016-2017. Epidemiologically, the mosaic penA allele was identified in all CRO-resistant isolates. Based on NG-MAST, ST5061 was the most prevalent ST. Phylogenetic analysis suggested that the resistant isolates did not cluster independently. Despite focus on the local situation, this study raises the need for better gonorrhea medication and highlights that CRO may not be adequate as first-line treatment for gonorrhea in Changsha.

Breviscapine suppresses the growth and metastasis of prostate cancer through regulating PAQR4-mediated PI3K/Akt pathway.

OBJECTIVES: Prostate cancer, one of the most frequently diagnosed tumors of men, leads to poor quality of life. Previous studies have shown that breviscapine (BRE) exerts therapeutic activity in malignant tumors. However, the role and mechanism of BRE exhibit an anti-tumor effect on prostate cancer are largely unknown. METHODS: The mRNA and protein levels in prostate cancer tissues and cell lines were measured using RT-qPCR, western blot, and immunohistochemical staining, respectively. Cell proliferation, invasion, and migration in both PC3 and DU145 cells were evaluated using CCK-8 and Transwell assay. The effect of BRE on cell proliferation and metastasis by regulating the PAQR4-mediated PI3K/Akt pathway in vitro and in vivo was determined. RESULTS: PAQR4 was significantly overexpressed in prostate cancer tissues and cell lines, which was positively correlated with poor prognosis. Knockdown of PAQR4 inhibited the proliferation, invasion, migration, and epithelial-mesenchymal transition (EMT) of both PC3 and DU145 cells. Mechanistically, BRE treatment significantly suppressed the malignant biological behavior of both prostate cancer cells by downregulating PAQR4 and blocking the PI3K/Akt pathway. In vivo experiments, BRE administration remarkably inhibited tumor growth and metastasis in a xenograft model of prostate cancer. CONCLUSION: Our findings revealed that BRE exerts anti-tumor and anti-metastasis roles in prostate cancer by inhibiting PAQR4-mediated PI3K/Akt pathway, which provides a new therapeutic agent for prostate cancer clinical treatment.

The novel TP53 3'-end methylation pattern associated with its expression would be a potential biomarker for breast cancer detection.

BACKGROUND: Deficiency or silence of TP53 is an early event in breast tumorigenesis. Aberrant methylation and mutation in regulatory regions were considered as crucial regulators of gene expression. METHODS: Using multiplex-PCR and next-generation sequencing technology, we analyzed TP53 mutation spectrum in its promoter region. Using PCR target sequence enrichment and next-generation bisulfite sequencing technology, we analyzed the methylation profile of the promoter and 3'-end regions of TP53 gene in paired breast tumor and normal tissues from 120 breast cancer patients. The expression of TP53 and the flanking gene ATP1B2 was explored with qPCR method in the same cohort. RESULTS: No promoter mutation of TP53 gene was found in the cohort of the 120 breast cancer patients. The 3'-end of TP53 gene was hyper-methylated (average 78.71%) compared with the promoter region (average less than 1%) in breast tumor tissues. TP53 was significantly lower expressed (P = 1.68E-15) and hyper-methylated in 3'-end (P = 1.82E-18) in tumor. Negative cis correlation was found between the TP53 expression and its 3'-end methylation (P = 9.02E-8, R = 0.337). TP53 expression was significantly associated with PR status (P = 0.0128), Ki67 level (P = 0.0091), and breast cancer subtypes (P = 0.0109). TP53 3'-end methylation and expression showed a good performance in discriminating breast cancer and normal tissues with an AUC of 0.930. CONCLUSIONS: The 3'-end methylation of TP53 might be a crucial regulator for its expression in breast cancer, suggesting that TP53 3'-end hyper-methylation associated with its lower expression could be a potential biomarker for breast cancer diagnosis.

Homeobox B8 Targets Sterile Alpha Motif Domain-Containing Protein 9 and Drives Glioma Progression.

Gliomas are the most commonly occurring tumors of the central nervous system. Glioblastoma multiforme (GBM) is the most malignant and aggressive brain cancer in adults. Further understanding of the mechanisms underlying the aggressive nature of GBM is urgently needed. Here we identified homeobox B8 (HOXB8), a member of the homeobox family, as a crucial contributor to the aggressiveness of GBM. Data mining of publicly accessible RNA sequence datasets and our patient cohorts confirmed a higher expression of HOXB8 in the tumor tissue of GBM patients, and a strong positive correlation between the expression level and pathological grading of tumors and a negative correlation between the expression level and the overall survival rate. We next showed that HOXB8 promotes the proliferation and migration of glioblastoma cells and is crucial for the activation of the PI3K/AKT pathway and expression of epithelial-mesenchymal transition-related genes, possibly through direct binding to the promoter of SAMD9 (Sterile Alpha Motif Domain-Containing Protein 9) and activating its transcription. Collectively, we identified HOXB8 as a critical contributor to the aggressiveness of GBM, which provides insights into a potential therapeutic target for GBM and opens new avenues for improving its treatment outcome.

C1Q/TNF-related protein 4 expression correlates with herpes simplex encephalitis progression.

BACKGROUND: Herpes simplex encephalitis (HSE), an acute inflammatory disease of the central nervous system is caused by the herpes simplex virus infection. HSE occurs at any age, and it is often accompanied by high mortality and neurological dysfunction. The C1Q/TNF-related protein (CTRP) family, usually contains a homotrimeric structure, which comprises the N-terminal signal peptide and the C-terminal C1q globular domain. It has been demonstrated that CTRPs play pivotal roles in the inflammation process. CTRP4 is a member of the CTRP family and contains two C1q globular domains. Moreover, evidence shows that the recombinant human CTRP4 (rhCTRP4) protein exerts satisfactory anti-inflammatory effects in experimental colitis models via the NF-κB pathway. However, its role in inflammation-related neurological diseases remains unknown. METHODS: The purpose of this study is to evaluate the expression of CTRP4 and its correlation with HSE progression. We determined the serum CTRP4 levels in a normal brain, tuberculous meningitis (TBM), bacterial meningitis (BM) and HSE. RESULTS: We found that compared to a normal brain, TBM and BM, CTRP4 was significantly increased in HSE. Moreover, in the course of HSE, serum interleukin (IL-6) and necrosis factor-α (TNF-α) were also increased and were closely associated with CTRP4 expression. CTRP4 expression was examined by immunohistochemistry (IHC) in the normal control brain tissues, HSE, TBM and BM brain tissues. High positively expression of CTRP4 was found in HSE. In the normal brain tissue, TBM, and BM brain tissues, CTRP4 showed a weak expression. In the clinical evaluation, CTRP4 expression correlated closely with an ascending stage of the disease [mini-mental state examination (MMSE) evaluation, MRI imaging). CONCLUSIONS: Our findings suggest that CTRP4 is highly expressed in HSE and is closely related to the progression of HSE. Thus, CTRP4 may serve as a potential severity index for HSE and targeting CTRP4 might be a promising therapeutic strategy against HSE.

MARVELD1 attenuates arsenic trioxide-induced apoptosis in liver cancer cells by inhibiting reactive oxygen species production.

BACKGROUND: Arsenic trioxide (As2O3) is widely used for the treatment of acute promyelocytic leukemia (APL), and more recently, has also been applied to solid tumors. However, there are a fraction of patients with solid tumors, such as liver cancer, who respond to As2O3 treatment poorly. The underlying mechanisms for this remain unclear. METHODS: We determined the suitable concentration of drugs by IC50. Cell Counting Kit-8 (CCK-8) and flow cytometry were used to analyze the apoptosis. Morphological changes of the cells were observed by laser scanning confocal microscopy. Furthermore, reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) were detected by flow cytometry. Quantitative polymerase chain reaction (qPCR) and Western blot tests were conducted to detect the mRNA and protein levels in different groups. Finally, a xenograft tumor assay and histopathological analysis were performed to evaluate the MARVELD1 function in cell proliferation and apoptosis. RESULTS: Here, we show that MARVELD1 enhances the therapeutic effects of epirubicin, while inducing the strong resistance of liver cancer cells to As2O3 treatment. We further demonstrate that the As2O3-induced apoptosis was inhibited by MARVELD1 overexpression (24 h Vector vs. MARVELD1 =30.58% vs. 17.41%, P<0.01; 48 h Vector vs. MARVELD1 =46.50% vs. 21.02%, P<0.01), possibly through inhibiting ROS production by enhancing TRXR1 expression. In vivo, we found a significantly increased size (Vector vs. MARVELD1 =203.90±21.92 vs. 675.70±37.84 mm3, P<0.001) and weight (Vector vs. MARVELD1 =0.19±0.02 vs. 0.58±0.05 g, P<0.001) of tumors with high expression of MARVELD1 after As2O3 treatment. Consistently, a higher expression of MARVELD1 predicted a poor prognosis for liver cancer patients. CONCLUSIONS: Our data identified a unique role of MARVELD1 in As2O3-induced apoptosis and As2O3 cancer therapy resistance.

Diversity of the microbial community and cultivable protease-producing bacteria in the sediments of the Bohai Sea, Yellow Sea and South China Sea.

The nitrogen (N) cycle is closely related to the stability of marine ecosystems. Microbial communities have been directly linked to marine N-cycling processes. However, systematic research on the bacterial community composition and diversity involved in N cycles in different seas is lacking. In this study, microbial diversity in the Bohai Sea (BHS), Yellow Sea (YS) and South China Sea (SCS) was surveyed by targeting the hypervariable V4 regions of the 16S rRNA gene using next-generation sequencing (NGS) technology. A total of 2,505,721 clean reads and 15,307 operational taxonomic units (OTUs) were obtained from 86 sediment samples from the three studied China seas. LEfSe analysis demonstrated that the SCS had more abundant microbial taxa than the BHS and YS. Diversity indices demonstrated that Proteobacteria and Planctomycetes were the dominant phyla in all three China seas. Canonical correspondence analysis (CCA) indicated that pH (P = 0.034) was the principal determining factors, while the organic matter content, depth and temperature had a minor correlated with the variations in sedimentary microbial community distribution. Cluster and functional analyses of microbial communities showed that chemoheterotrophic and aerobic chemoheterotrophic microorganisms widely exist in these three seas. Further research found that the cultivable protease-producing bacteria were mainly affiliated with the phyla Proteobacteria, Firmicutes and Bacteroidetes. It was very clear that Pseudoalteromonadaceae possessed the highest relative abundance in the three sea areas. The predominant protease-producing genera were Pseudoalteromonas and Bacillus. These results shed light on the differences in bacterial community composition, especially protease-producing bacteria, in these three China seas.

Comprehensive analysis of serum tumor markers and BRCA1/2 germline mutations in Chinese ovarian cancer patients.

BACKGROUND: The serum tumor markers has been widely used in ovarian cancer diagnosis. BRCA1/2 germline mutations are the most common predisposing factors for ovarian cancer development. This study aimed to comprehensively investigate serum tumor markers and BRCA1/2 germline mutations and analyze their associations with ovarian cancer. METHODS: Levels of 11 serum tumor markers were examined in ovarian cancer patients and controls with benign gynecologic diseases. By integrating multiplex PCR and next-generation sequencing technologies, BRCA1/2 germline mutations were analyzed and confirmed by Sanger sequencing. The discriminative models with serum tumor markers and BRCA1/2 mutation status were constructed for ovarian cancer detection and patient stratification. RESULTS: Among 11 markers, six of them were significantly elevated and only beta-human chorionic gonadotropin (ß-HCG) was significantly reduced in ovarian cancer patients. A total of 54 (23.3%) ovarian cancer patients were found to harbor BRCA1/2 deleterious mutations, and BRCA1/2 mutations were significantly associated with Hereditary Breast and Ovarian Cancer-related tumors and family history of cancer. Carbohydrate antigen 125 showed a good performance in ovarian cancer detection as a single marker (AUC = 0.799), while a panel of eight markers showed a good performance in BRCA1 mutation detection with an AUC value of 0.974. In addition, a panel of five serum tumor markers combined with BRCA1/2 mutation status showed a good performance in lymph node metastasis prediction (AUC = 0.843). CONCLUSIONS: We found the association between BRCA1/2 germline mutation status and serum tumor marker levels, and identified discriminative models that combined serum tumor markers with BRCA1/2 mutation status for ovarian cancer detection and patient stratification.

Prevalence and spectrum of AKT1, PIK3CA, PTEN and TP53 somatic mutations in Chinese breast cancer patients.

Breast cancer, one of the most frequently occurring cancers worldwide, is the leading cause of cancer-related death among women. AKT1, PIK3CA, PTEN and TP53 mutations were common observed in breast cancer representing potential clinical biomarkers for cancer classification and treatment. A comprehensive knowledge of AKT1, PIK3CA, PTEN and TP53 mutations in breast cancer was still insufficient in Chinese population. In this study, the complete coding regions and exon-intron boundaries of AKT1, PIK3CA, PTEN and TP53 genes were sequenced in paired breast tumor and normal tissues from 313 Chinese breast cancer patients using microfluidic PCR-based target enrichment and next-generation sequencing technology. Total 120 somatic mutations were identified in 190 of the 313 patients (60.7%), with the mutation frequency of AKT1 as 3.2%, PIK3CA as 36.4%, PTEN as 4.8%, and TP53 as 33.9%. Among these mutations, 1 in PIK3CA (p.I69N), 3 in PTEN (p.K62X, c.635-12_636delTTAACCATGCAGAT and p.N340IfsTer4) and 5 in TP53 (p.Q136AfsTer5, p.K139_P142del, p.Y234dup, p.V274LfsTer31 and p.N310TfsTer35) were novel. Notably, PIK3CA somatic mutations were significantly associated with ER-positive or PR-positive tumors. TP53 somatic mutations were significantly associated with ER-negative, PR-negative, HER2-positive, BRCA1 mutation, Ki67 high expression and basal-like tumors. Our findings provided a comprehensive mutation profiling of AKT1, PIK3CA, PTEN and TP53 genes in Chinese breast cancer patients, which have potential implications in clinical management.

Analysis of BRCA1/2 mutation spectrum and prevalence in unselected Chinese breast cancer patients by next-generation sequencing.

PURPOSE: BRCA1 and BRCA2 (BRCA1/2) are two major high-penetrance breast cancer predisposition genes, mutations in which can lead to high risks and early onset of breast cancer. This study was performed to comprehensively investigate the spectrum and prevalence of BRCA1/2 mutations in unselected Chinese breast cancer patients and evaluate the associations of BRCA1/2 mutations with related clinicopathological characteristics of the tumors. METHODS: By integrating microfluidic PCR-based target enrichment and next-generation sequencing, paired tumor and normal tissues from 313 unselected breast cancer patients were analyzed for both germline and somatic mutations of BRCA1/2 genes in Chinese Han population. RESULTS: Total 5 BRCA1 and 8 BRCA2 deleterious germline mutations were detected in 5 (1.60%) and 12 (3.83%) of the 313 patients, respectively. The entire frequency of deleterious germline mutations of BRCA1/2 was 5.43%. Among them, c.1069A > T and c.3418_3419insTGACTACT in BRCA1, c.8474_8487delCATACCCTATACAG and c.6547delG in BRCA2 were novel. In addition, 32 germline variants of unknown significance in 31 (9.90%) of the 313 patients were identified. We also detected 13 somatic mutations in ten patients (3.19%), including 4 (1.28%) deleterious mutations (c.1575delT, c.2677C > T, c.7024C > T, and c.7672G > T in BRCA2) and 5 novel mutations (c.4728A > G and c.4820T > C in BRCA1; c.2527G > A, c.4069C > G and c.7672G > T in BRCA2). Notably, BRCA1 mutation carriers were significantly younger, and more likely to be ER negative and basal-like breast cancers. CONCLUSIONS: Our study provided a reliable and effective platform for BRCA1/2 genetic testing, and suggested that there was a relatively high prevalence and special spectrum of BRCA1/2 mutations in unselected Chinese breast cancer patients.

Integrated analysis of promoter methylation and expression of telomere related genes in breast cancer.

Telomeres at the ends of eukaryotic chromosomes play a critical role in tumorgenesis. Using microfluidic PCR and next-generation bisulfite sequencing technology, we investigated the promoter methylation of 29 telomere related genes in paired tumor and normal tissues from 184 breast cancer patients. The expression of significantly differentially methylated genes was quantified using qPCR method.We observed that the average methylation level of the 29 telomere related genes was significant higher in tumor than that in normal tissues (P = 4.30E-21). A total of 4 genes (RAD50, RTEL, TERC and TRF1) showed significant hyper-methylation in breast tumor tissues. RAD51D showed significant methylation difference among the four breast cancer subtypes. The methylation of TERC showed significant association with ER status of breast cancer. The expression profiles of the 4 hyper-methylated genes showed significantly reduced expression in tumor tissues. The integration analysis of methylation and expression of these 4 genes showed a good performance in breast cancer prediction (AUC = 0.947).Our results revealed the methylation pattern of telomere related genes in breast cancer and suggested a novel 4-gene panel might be a valuable biomarker for breast cancer diagnosis.

Integrated analysis of promoter mutation, methylation and expression of AKT1 gene in Chinese breast cancer patients.

BACKGROUND: As downstream mediators of PI3K /PTEN /AKT /mTORC1 pathway, the AKT isoforms play critical roles in tumorgenesis. Although the pleiotropic effects of AKT1 in breast cancer have been reported, the genetic and epigenetic characteristics of AKT1 promoter region in breast cancer remains to be identified. In this study we aimed to investigate the promoter mutation spectrum, methylation and gene expression pattern of AKT1 and their relationship with breast cancer. METHODS: By using PCR target sequence enrichment and next-generation sequencing technology, we sequenced AKT1 promoter region in pairs of breast tumor and normal tissues from 95 unselected Chinese breast cancer patients. The methylation of the promoter region and the expression profile of AKT1 in the same cohort were detected with bisulfite next-generation sequencing and qPCR, respectively. RESULTS: We identified 28 somatic mutations in 23 of the 95 (24.2%) breast cancer samples. And 19 of the 28 mutations were located in transcription factor (TF) binding sites. In the 23 patients with somatic mutations, no significant change of methylation or expression was found comparing with other patients. AKT1 promoter region was significantly hypo-methylated in tumor compared with matched normal tissue (P = 0.0014) in the 95 patients. The expression of AKT1 was significantly suppressed in tumor tissue (P = 0.0375). In clinicopathological factor analysis, AKT1 showed significant hypo-methylation (P = 0.0249) and suppressed expression (P = 0.0375) in HER2 negative subtype. And a trend of decrease in expression level (P = 0.0624) of AKT1 in the ER negative subtype was observed, which is significantly decreased in basal-like breast tumor (P = 0.0328). CONCLUSIONS: Hypo-methylation and suppressed expression of AKT1 was observed to be associated with breast cancer in our cohort. The methylation and expression of AKT1 were both significantly associated with HER2 status. The promoter mutation of AKT1 did not show significant association with its methylation and expression status. These results suggested that the promoter mutation, methylation and gene expression of AKT1 may play distinct roles in tumorgenesis of breast cancer and the integrated analysis of methylation and expression of AKT1 might serve as potential biomarkers for diagnosis and classification of breast cancer.

Integrated analysis of gene expression and methylation profiles of 48 candidate genes in breast cancer patients.

PURPOSE: Gene-specific methylation and expression have shown biological and clinical importance for breast cancer diagnosis and prognosis. Integrated analysis of gene methylation and gene expression may identify genes associated with biology mechanism and clinical outcome of breast cancer and aid in clinical management. METHODS: Using high-throughput microfluidic quantitative PCR, we analyzed the expression profiles of 48 candidate genes in 96 Chinese breast cancer patients and investigated their correlation with gene methylation and associations with breast cancer clinical parameters. RESULTS: Breast cancer-specific gene expression alternation was found in 25 genes with significant expression difference between paired tumor and normal tissues. A total of 9 genes (CCND2, EGFR, GSTP1, PGR, PTGS2, RECK, SOX17, TNFRSF10D, and WIF1) showed significant negative correlation between methylation and gene expression, which were validated in the TCGA database. Total 23 genes (ACADL, APC, BRCA2, CADM1, CAV1, CCND2, CST6, EGFR, ESR2, GSTP1, ICAM5, NPY, PGR, PTGS2, RECK, RUNX3, SFRP1, SOX17, SYK, TGFBR2, TNFRSF10D, WIF1, and WRN) annotated with potential TFBSs in the promoter regions showed negative correlation between methylation and expression. In logistics regression analysis, 31 of the 48 genes showed improved performance in disease prediction with combination of methylation and expression coefficient. CONCLUSIONS: Our results demonstrated the complex correlation and the possible regulatory mechanisms between DNA methylation and gene expression. Integration analysis of methylation and expression of candidate genes could improve performance in breast cancer prediction. These findings would contribute to molecular characterization and identification of biomarkers for potential clinical applications.

Methylation analysis of plasma cell-free DNA for breast cancer early detection using bisulfite next-generation sequencing.

Circulating cell-free DNA (cfDNA) has been considered as a potential biomarker for non-invasive cancer detection. To evaluate the methylation levels of six candidate genes (EGFR, GREM1, PDGFRB, PPM1E, SOX17, and WRN) in plasma cfDNA as biomarkers for breast cancer early detection, quantitative analysis of the promoter methylation of these genes from 86 breast cancer patients and 67 healthy controls was performed by using microfluidic-PCR-based target enrichment and next-generation bisulfite sequencing technology. The predictive performance of different logistic models based on methylation status of candidate genes was investigated by means of the area under the ROC curve (AUC) and odds ratio (OR) analysis. Results revealed that EGFR, PPM1E, and 8 gene-specific CpG sites showed significantly hypermethylation in cancer patients' plasma and significantly associated with breast cancer (OR ranging from 2.51 to 9.88). The AUC values for these biomarkers were ranging from 0.66 to 0.75. Combinations of multiple hypermethylated genes or CpG sites substantially improved the predictive performance for breast cancer detection. Our study demonstrated the feasibility of quantitative measurement of candidate gene methylation in cfDNA by using microfluidic-PCR-based target enrichment and bisulfite next-generation sequencing, which is worthy of further validation and potentially benefits a broad range of applications in clinical oncology practice. Quantitative analysis of methylation pattern of plasma cfDNA by next-generation sequencing might be a valuable non-invasive tool for early detection of breast cancer.

Methylation profiling of 48 candidate genes in tumor and matched normal tissues from breast cancer patients.

Gene-specific methylation alterations in breast cancer have been suggested to occur early in tumorigenesis and have the potential to be used for early detection and prevention. The continuous increase in worldwide breast cancer incidences emphasizes the urgent need for identification of methylation biomarkers for early cancer detection and patient stratification. Using microfluidic PCR-based target enrichment and next-generation bisulfite sequencing technology, we analyzed methylation status of 48 candidate genes in paired tumor and normal tissues from 180 Chinese breast cancer patients. Analysis of the sequencing results showed 37 genes differentially methylated between tumor and matched normal tissues. Breast cancer samples with different clinicopathologic characteristics demonstrated distinct profiles of gene methylation. The methylation levels were significantly different between breast cancer subtypes, with basal-like and luminal B tumors having the lowest and the highest methylation levels, respectively. Six genes (ACADL, ADAMTSL1, CAV1, NPY, PTGS2, and RUNX3) showed significant differential methylation among the 4 breast cancer subtypes and also between the ER +/ER- tumors. Using unsupervised hierarchical clustering analysis, we identified a panel of 13 hypermethylated genes as candidate biomarkers that performed a high level of efficiency for cancer prediction. These 13 genes included CST6, DBC1, EGFR, GREM1, GSTP1, IGFBP3, PDGFRB, PPM1E, SFRP1, SFRP2, SOX17, TNFRSF10D, and WRN. Our results provide evidence that well-defined DNA methylation profiles enable breast cancer prediction and patient stratification. The novel gene panel might be a valuable biomarker for early detection of breast cancer.

Interaction of TLR-IFN and HLA polymorphisms on susceptibility of chronic HBV infection in Southwest Han Chinese.

BACKGROUND & AIMS: The toll-like receptor-interferon (TLR-IFN) signalling pathway plays a crucial role in HBV infection. Human leucocyte antigen (HLA) polymorphisms are associated with chronic HBV infection by genome wide association study (GWAS). We aimed to explore interaction between TLR-IFN and HLA gene polymorphisms in susceptibility of chronic HBV infection. METHODS: In the Chinese Southwest Han population, 1191 chronic HBV infection patients and 273 HBV clearance were selected. A total of 39 single nucleotide polymorphism loci in 23 genes of the TLR-IFN pathway and four HLA polymorphism loci associated with chronic HBV infection identified by GWAS were selected for genotyping. SNPStats, QVALUE, and multifactor dimensionality reduction were used for statistical analysis. RESULTS: A significant association was seen in several of the TLR-IFN pathway genes, TLR9 rs352140 (OR = 0.70, P = 0.0088), IL1B rs16944 (OR = 0.67, P = 0.016), IL12B rs3212227 (OR = 1.38, P = 0.021), IFNGR1 rs3799488 (OR = 1.48, P = 0.0048), IFNGR2 rs1059293 (OR = 0.27, P = 0.011), MX1 rs467960 (OR = 0.68, P = 0.022), as well as four loci in HLA, rs3077 (OR = 0.55, P < 0.0001), rs2856718 (OR = 0.60, P = 4e-04), rs9277535 (OR = 0.54, P < 0.0001) and rs7453920 (OR = 0.43, P < 0.0001). A synergistic relationship was seen between rs9277535 and rs16944 (0.13%), rs1143623 and rs6613 (0.10%). The combination of rs9277535 in HLA and rs16944 in IL1B was the best model to predict chronic HBV infection (testing accuracy = 0.6040, P = 0.0010, cross-validation consistency = 10/10). CONCLUSIONS: TLR-IFN pathway gene polymorphisms are associated with chronic HBV infection. Interactions with polymorphisms in these genes may be one mechanism by which HLA polymorphisms influence susceptibility to chronic HBV infection, as specific single nucleotide polymorphism combinations are highly predictive of chronic HBV infection.

Local selection of human populations shapes complex evolution patterns of CXCL10 gene.

CXC motif chemokine 10 (CXCL10) is a small cytokine belonging to the CXC chemokine family, and it is secreted by several cell types in response to IFN-γ and regulates immune responses through the recruitment and activation of lymphocytes. As CXCL10 is very important in T-cell immunity and infectious diseases, we studied the effect of natural selection on the CXCL10 locus. By sequencing 74 individuals from three human populations, we found a complex pattern of natural selection acting on the CXCL10 locus. We discovered a signature of balancing selection in the European population with a significant positive Tajima's D value (2.57, P=0.005) and an excess of intermediate frequency alleles. However, we observed an excess of high frequency-derived alleles and a significant Fay and Wu's test statistics (P=0.015) in the Chinese population, which suggests that recent selective sweeps under positive selection had occurred. Also, there are a lot of alleles showing great frequency difference among populations. These results demonstrate that local selection has shaped CXCL10 evolution and indicates that there exist different actions of natural selection on the CXCL10 locus in different populations. This study provides insights into the likely relative roles of natural selection and population history in shaping today's genetic variation at the CXCL10 locus, indicates the relationship between adaptation to past infection and predisposition to autoimmunity in modern populations, improves our understanding of CXCL10 evolution, and motivates further investigations of the role of CXCL10 in infectious diseases and autoimmune diseases.

Exome sequencing identifies compound heterozygous mutations in CYP4V2 in a pedigree with retinitis pigmentosa.

Retinitis pigmentosa (RP) is a heterogeneous group of progressive retinal degenerations characterized by pigmentation and atrophy in the mid-periphery of the retina. Twenty two subjects from a four-generation Chinese family with RP and thin cornea, congenital cataract and high myopia is reported in this study. All family members underwent complete ophthalmologic examinations. Patients of the family presented with bone spicule-shaped pigment deposits in retina, retinal vascular attenuation, retinal and choroidal dystrophy, as well as punctate opacity of the lens, reduced cornea thickness and high myopia. Peripheral venous blood was obtained from all patients and their family members for genetic analysis. After mutation analysis in a few known RP candidate genes, exome sequencing was used to analyze the exomes of 3 patients III2, III4, III6 and the unaffected mother II2. A total of 34,693 variations shared by 3 patients were subjected to several filtering steps against existing variation databases. Identified variations were verified in the rest family members by PCR and Sanger sequencing. Compound heterozygous c.802-8_810del17insGC and c.1091-2A>G mutations of the CYP4V2 gene, known as genetic defects for Bietti crystalline corneoretinal dystrophy, were identified as causative mutations for RP of this family.

RNA-seq analysis of prostate cancer in the Chinese population identifies recurrent gene fusions, cancer-associated long noncoding RNAs and aberrant alternative splicings.

There are remarkable disparities among patients of different races with prostate cancer; however, the mechanism underlying this difference remains unclear. Here, we present a comprehensive landscape of the transcriptome profiles of 14 primary prostate cancers and their paired normal counterparts from the Chinese population using RNA-seq, revealing tremendous diversity across prostate cancer transcriptomes with respect to gene fusions, long noncoding RNAs (long ncRNA), alternative splicing and somatic mutations. Three of the 14 tumors (21.4%) harbored a TMPRSS2-ERG fusion, and the low prevalence of this fusion in Chinese patients was further confirmed in an additional tumor set (10/54=18.5%). Notably, two novel gene fusions, CTAGE5-KHDRBS3 (20/54=37%) and USP9Y-TTTY15 (19/54=35.2%), occurred frequently in our patient cohort. Further systematic transcriptional profiling identified numerous long ncRNAs that were differentially expressed in the tumors. An analysis of the correlation between expression of long ncRNA and genes suggested that long ncRNAs may have functions beyond transcriptional regulation. This study yielded new insights into the pathogenesis of prostate cancer in the Chinese population.