Plasmodium vivax Tryptophan-Rich Antigen PvTRAg33.5 Contains Alpha Helical Structure and Multidomain Architecture

Tryptophan-rich proteins from several malarial parasites have been identified where they play an important role in host-parasite interaction. Structural characterization of these proteins is needed to develop them as therapeutic targets. Here, we describe a novel Plasmodium vivax tryptophan-rich protein named PvTRAg33.5. It is expressed by blood stage(s) of the parasite and its gene contains two exons. The exon 1 encodes for a 23 amino acids long putative signal peptide which is likely to be cleaved off whereas the exon 2 encodes for the mature protein of 252 amino acids. The mature protein contains B-cell epitopes which were recognized by the human immune system during P.vivax infection. The PvTRAg33.5 contains 24 (9.5%) tryptophan residues and six motifs whose patterns were similar among tryptophan-rich proteins. The modeled structure of the PvTRAg33.5 consists of a multidomain architecture which is stabilized by the presence of large number of tryptophan residues. The recombinant PvTRAg33.5 showed predominantly α helical structure and alpha helix to beta sheet transition at pH below 4.5. Protein acquires an irreversible non-native state at temperature more than 50°C at neutral pH. Its secondary and tertiary structures remain stable in the presence of 35% alcohol but these structures are destabilized at higher alcohol concentrations due to the disturbance of hydrophobic interactions between tryptophanyl residues. These structural changes in the protein might occur during its translocation to interact with other proteins at its final destination for biological function such as erythrocyte invasion

Identification of Enriched Driver Gene Alterations in Subgroups of Non-Small Cell Lung Cancer Patients Based on Histology and Smoking Status

BACKGROUND: Appropriate patient selection is needed for targeted therapies that are efficacious only in patients with specific genetic alterations. We aimed to define subgroups of patients with candidate driver genes in patients with non-small cell lung cancer. METHODS: Patients with primary lung cancer who underwent clinical genetic tests at Guangdong General Hospital were enrolled. Driver genes were detected by sequencing, high-resolution melt analysis, qPCR, or multiple PCR and RACE methods. RESULTS: 524 patients were enrolled in this study, and the differences in driver gene alterations among subgroups were analyzed based on histology and smoking status. In a subgroup of non-smokers with adenocarcinoma, EGFR was the most frequently altered gene, with a mutation rate of 49.8%, followed by EML4-ALK (9.3%), PTEN (9.1%), PIK3CA (5.2%), c-Met (4.8%), KRAS (4.5%), STK11 (2.7%), and BRAF (1.9%). The three most frequently altered genes in a subgroup of smokers with adenocarcinoma were EGFR (22.0%), STK11 (19.0%), and KRAS (12.0%). We only found EGFR (8.0%), c-Met (2.8%), and PIK3CA (2.6%) alterations in the non-smoker with squamous cell carcinoma (SCC) subgroup. PTEN (16.1%), STK11 (8.3%), and PIK3CA (7.2%) were the three most frequently enriched genes in smokers with SCC. DDR2 and FGFR2 only presented in smokers with SCC (4.4% and 2.2%, respectively). Among these four subgroups, the differences in EGFR, KRAS, and PTEN mutations were statistically significant. CONCLUSION: The distinct features of driver gene alterations in different subgroups based on histology and smoking status were helpful in defining patients for future clinical trials that target these genes. This study also suggests that we may consider patients with infrequent alterations of driver genes as having rare or orphan diseases that should be managed with special molecularly targeted therapies

Genome-Wide Screening for Genetic Alterations in Esophageal Cancer by aCGH Identifies 11q13 Amplification Oncogenes Associated with Nodal Metastasis

Esophageal squamous cell carcinoma (ESCC) is highly prevalent in China and other Asian countries, as a major cause of cancer-related mortality. ESCC displays complex chromosomal abnormalities, including multiple structural and numerical aberrations. Chromosomal abnormalities, such as recurrent amplifications and homozygous deletions, directly contribute to tumorigenesis through altering the expression of key oncogenes and tumor suppressor genes.To understand the role of genetic alterations in ESCC pathogenesis and identify critical amplification/deletion targets, we performed genome-wide 1-Mb array comparative genomic hybridization (aCGH) analysis for 10 commonly used ESCC cell lines. Recurrent chromosomal gains were frequently detected on 3q26-27, 5p15-14, 8p12, 8p22-24, 11q13, 13q21-31, 18p11 and 20q11-13, with frequent losses also found on 8p23-22, 11q22, 14q32 and 18q11-23. Gain of 11q13.3-13.4 was the most frequent alteration in ESCC. Within this region, CCND1 oncogene was identified with high level of amplification and overexpression in ESCC, while FGF19 and SHANK2 was also remarkably over-expressed. Moreover, a high concordance (91.5%) of gene amplification and protein overexpression of CCND1 was observed in primary ESCC tumors. CCND1 amplification/overexpression was also significantly correlated with the lymph node metastasis of ESCC.These findings suggest that genomic gain of 11q13 is the major mechanism contributing to the amplification. Novel oncogenes identified within the 11q13 amplicon including FGF19 and SHANK2 may play important roles in ESCC tumorigenesis

Mechanisms Underlying Hypoxia Tolerance in Drosophila melanogaster: hairy as a Metabolic Switch

Hypoxia-induced cell injury has been related to multiple pathological conditions. In order to render hypoxia-sensitive cells and tissues resistant to low O2 environment, in this current study, we used Drosophila melanogaster as a model to dissect the mechanisms underlying hypoxia-tolerance. A D. melanogaster strain that lives perpetually in an extremely low-oxygen environment (4% O2, an oxygen level that is equivalent to that over about 4,000 m above Mt. Everest) was generated through laboratory selection pressure using a continuing reduction of O2 over many generations. This phenotype is genetically stable since selected flies, after several generations in room air, survive at this low O2 level. Gene expression profiling showed striking differences between tolerant and naïve flies, in larvae and adults, both quantitatively and qualitatively. Up-regulated genes in the tolerant flies included signal transduction pathways (e.g., Notch and Toll/Imd pathways), but metabolic genes were remarkably down-regulated in the larvae. Furthermore, a different allelic frequency and enzymatic activity of the triose phosphate isomerase (TPI) was present in the tolerant versus naïve flies. The transcriptional suppressor, hairy, was up-regulated in the microarrays and its binding elements were present in the regulatory region of the specifically down-regulated metabolic genes but not others, and mutations in hairy significantly reduced hypoxia tolerance. We conclude that, the hypoxia-selected flies: (a) altered their gene expression and genetic code, and (b) coordinated their metabolic suppression, especially during development, with hairy acting as a metabolic switch, thus playing a crucial role in hypoxia-tolerance

Helicobacter pylori and oesophageal and gastric cancers in a prospective study in China

In a cohort of 29 584 residents of Linxian, China, followed from 1985 to 2001, we conducted a case–cohort study of the magnitude of the association of Helicobacter pylori seropositivity with cancer risk in a random sample of 300 oesophageal squamous cell carcinomas, 600 gastric cardia adenocarcinomas, all 363 diagnosed gastric non-cardia adenocarcinomas, and a random sample of the entire cohort (N=1050). Baseline serum was evaluated for IgG antibodies to whole-cell and CagA H. pylori antigens by enzyme-linked immunosorbent assay. Risks of both gastric cardia and non-cardia cancers were increased in individuals exposed to H. pylori (Hazard ratios (HRs) and 95% confidence intervals=1.64; 1.26–2.14, and 1.60; 1.15–2.21, respectively), whereas risk of oesophageal squamous cell cancer was not affected (1.17; 0.88–1.57). For both cardia and non-cardia cancers, HRs were higher in younger individuals. With longer time between serum collection to cancer diagnosis, associations became stronger for cardia cancers but weaker for non-cardia cancers. CagA positivity did not modify these associations. The associations between H. pylori exposure and gastric cardia and non-cardia adenocarcinoma development were equally strong, in contrast to Western countries, perhaps due to the absence of Barrett's oesophagus and oesophageal adenocarcinomas in Linxian, making all cardia tumours of gastric origin, rather than a mixture of gastric and oesophageal malignancies

Transitional Probability-Based Model for HPV Clearance in HIV-1-Positive Adolescent Females

BACKGROUND: HIV-1-positive patients clear the human papillomavirus (HPV) infection less frequently than HIV-1-negative. Datasets for estimating HPV clearance probability often have irregular measurements of HPV status and risk factors. A new transitional probability-based model for estimation of probability of HPV clearance was developed to fully incorporate information on HIV-1-related clinical data, such as CD4 counts, HIV-1 viral load (VL), highly active antiretroviral therapy (HAART), and risk factors (measured quarterly), and HPV infection status (measured at 6-month intervals). METHODOLOGY AND FINDINGS: Data from 266 HIV-1-positive and 134 at-risk HIV-1-negative adolescent females from the Reaching for Excellence in Adolescent Care and Health (REACH) cohort were used in this study. First, the associations were evaluated using the Cox proportional hazard model, and the variables that demonstrated significant effects on HPV clearance were included in transitional probability models. The new model established the efficacy of CD4 cell counts as a main clearance predictor for all type-specific HPV phylogenetic groups. The 3-month probability of HPV clearance in HIV-1-infected patients significantly increased with increasing CD4 counts for HPV16/16-like (p<0.001), HPV18/18-like (p<0.001), HPV56/56-like (p = 0.05), and low-risk HPV (p<0.001) phylogenetic groups, with the lowest probability found for HPV16/16-like infections (21.60±1.81% at CD4 level 200 cells/mm(3), p<0.05; and 28.03±1.47% at CD4 level 500 cells/mm(3)). HIV-1 VL was a significant predictor for clearance of low-risk HPV infections (p<0.05). HAART (with protease inhibitor) was significant predictor of probability of HPV16 clearance (p<0.05). HPV16/16-like and HPV18/18-like groups showed heterogeneity (p<0.05) in terms of how CD4 counts, HIV VL, and HAART affected probability of clearance of each HPV infection. CONCLUSIONS: This new model predicts the 3-month probability of HPV infection clearance based on CD4 cell counts and other HIV-1-related clinical measurements

Stomatin Inhibits Pannexin-1-Mediated Whole-Cell Currents by Interacting with Its Carboxyl Terminal

The pannexin-1 (Panx1) channel (often referred to as the Panx1 hemichannel) is a large-conductance channel in the plasma membrane of many mammalian cells. While opening of the channel is potentially detrimental to the cell, little is known about how it is regulated under physiological conditions. Here we show that stomatin inhibited Panx1 channel activity. In transfected HEK-293 cells, stomatin reduced Panx1-mediated whole-cell currents without altering either the total or membrane surface Panx1 protein expression. Stomatin coimmunoprecipitated with full-length Panx1 as well as a Panx1 fragment containing the fourth membrane-spanning domain and the cytosolic carboxyl terminal. The inhibitory effect of stomatin on Panx1-mediated whole-cell currents was abolished by truncating Panx1 at a site in the cytosolic carboxyl terminal. In primary culture of mouse astrocytes, inhibition of endogenous stomatin expression by small interfering RNA enhanced Panx1-mediated outward whole-cell currents. These observations suggest that stomatin may play important roles in astrocytes and other cells by interacting with Panx1 carboxyl terminal to limit channel opening

Hypermethylation of the DLC1 CpG island does not alter gene expression in canine lymphoma

<p>Abstract</p> <p>Background</p> <p>This study is a comparative epigenetic evaluation of the methylation status of the <it>DLC1 </it>tumor suppressor gene in naturally-occurring canine lymphoma. Canine non-Hodgkin's lymphoma (NHL) has been proposed to be a relevant preclinical model that occurs spontaneously and may share causative factors with human NHL due to a shared home environment. The canine <it>DLC1 </it>mRNA sequence was derived from normal tissue. Using lymphoid samples from 21 dogs with NHL and 7 normal dogs, the methylation status of the promoter CpG island of the gene was defined for each sample using combined bisulfite restriction analysis (COBRA), methylation-specific PCR (MSP), and bisulfite sequencing methods. Relative gene expression was determined using real-time PCR.</p> <p>Results</p> <p>The mRNA sequence of canine <it>DLC1 </it>is highly similar to the human orthologue and contains all protein functional groups, with 97% or greater similarity in functional regions. Hypermethylation of the 5' and 3' flanking regions of the promoter was statistically significantly associated with the NHL phenotype, but was not associated with silencing of expression or differences in survival.</p> <p>Conclusion</p> <p>The canine <it>DLC1 </it>is constructed highly similarly to the human gene, which has been shown to be an important tumor suppressor in many forms of cancer. As in human NHL, the promoter CpG island of <it>DLC1 </it>in canine NHL samples is abnormally hypermethylated, relative to normal lymphoid tissue. This study confirms that hypermethylation occurs in canine cancers, further supporting the use of companion dogs as comparative models of disease for evaluation of carcinogenesis, biomarker diagnosis, and therapy.</p