Estimation of parent specific DNA copy number in tumors using high-density genotyping arrays

Chromosomal gains and losses comprise an important type of genetic change in tumors, and can now be assayed using microarray hybridization-based experiments. Most current statistical models for DNA copy number estimate total copy number, which do not distinguish between the underlying quantities of the two inherited chromosomes. This latter information, sometimes called parent specific copy number, is important for identifying allele-specific amplifications and deletions, for quantifying normal cell contamination, and for giving a more complete molecular portrait of the tumor. We propose a stochastic segmentation model for parent-specific DNA copy number in tumor samples, and give an estimation procedure that is computationally efficient and can be applied to data from the current high density genotyping platforms. The proposed method does not require matched normal samples, and can estimate the unknown genotypes simultaneously with the parent specific copy number. The new method is used to analyze 223 glioblastoma samples from the Cancer Genome Atlas (TCGA) project, giving a more comprehensive summary of the copy number events in these samples. Detailed case studies on these samples reveal the additional insights that can be gained from an allele-specific copy number analysis, such as the quantification of fractional gains and losses, the identification of copy neutral loss of heterozygosity, and the characterization of regions of simultaneous changes of both inherited chromosomes.     

Terpenoid and phenolic metabolites from the fungus xylaria sp. associated with termite nests

Three new sesquiterpene acids, xylaric acids A-C (1-3, resp.), and a new tetralone (=3,4-dihydronaphthalen-1(2H)-one) derivative, 4, along with nine known compounds, xylaric acid D (5), hydroheptelidic acid (6), gliocladic acid (7), chlorine heptelidic acid (8), trichoderonic acid A (9), 16-(α-D-mannopyranosyloxy)isopimar-7-en-19-oic acid (10), 16-(α-D-glucopyranosyloxy)isopimar-7-en-19-oic acid (11), 5-carboxymellein (12), and naphthalen-1,8-diol 1-O-α-D-glucopyranoside (13) have been isolated from the solid culture of the ascomycete fungus Xylaria sp. associated with termite nest. The structures of these compounds were elucidated primarily by NMR experiments. The absolute configurations of compounds 1-3 and 5-9 were determined by combination of X-ray data and CD spectral analysis. The absolute configuration of 4 was assigned by Snatzke's method. Compounds 8 and 11 showed slight cytotoxicities against two cell lines A549 and SGC7901.     

Enhanced contact allergen- and UVB-induced keratinocyte apoptosis in the absence of CD95/Fas/Apo-1

FAS/CD95/Apo-1 is a ubiquitously expressed cell-surface receptor involved in the initiation of programmed cell death. Its function in epidermal keratinocytes has been incompletely defined. Available evidence from in vitro studies points to important roles of Fas in the pathogenesis of contact dermatitis and in keratinocyte apoptosis induced by ultraviolet light. To define functions of Fas in the epidermis in vivo, we have generated mice with epidermis-specific deletion of the fas gene and tested its requirement for 2,4-dinitrofluorobenzene-induced contact dermatitis and for ultraviolet light B (UVB)-induced keratinocyte apoptosis. We report here our unexpected finding that keratinocyte apoptosis induced by both a contact allergen and UVB irradiation was significantly enhanced in Fas-negative epidermis. Expression of Fas by epidermal keratinocytes was neither necessary for the normal development of contact hypersensitivity of the skin, nor required for keratinocyte apoptosis following UVB irradiation. Our study results thus show that in the epidermis in vivo Fas exerts antiapoptotic effects that outweigh its proapoptotic role in contact hypersensitivity responses of the skin and in the tissue response of the epidermis to UVB irradiation.     

Molecular cloning, characterization and expression of a C-type lectin cDNA in Chinese mitten crab, Eriocheir sinensis

C-type lectins are pattern-recognition proteins which are functionally important for pathogen recognition and immune regulation in vertebrates and invertebrates. In this study, a lectin cDNA named as Es-Lectin was cloned and characterized from the Chinese mitten crab, Eriocheir sinensis. The full-length sequence of this Es-Lectin cDNA was 651 bp, including an open reading frame of 483 bp encoding 160 amino acids. The predicted molecular weight of the Es-Lectin was 11.8 kDa. A typical signal peptide of 21 amino acids was deduced at the N-terminus of the predicted protein. This Es-Lectin belongs to a C-type lectin and contains six cysteines, a conserved EPN motif (Glu-Pro-Asn) and an imperfect WND (Trp-Asn-Asp) motif (FND, Phe-Asn-Asp). This Es-Lectin had 55% and 32% identity with other two C-type lectins in E. sinensis, and 29-36% homology with decapods. Although the Es-Lectin was also expressed in gill, hepatopancreas, intestine, muscle and stomach, its expression in haemocytes was the greatest. The expression of Es-Lectins in haemocytes increased at 1.5 h after the Aeromonas hydrophila challenge. After a slight decrease, the Es-Lectin expression in haemocytes significantly increased at 48 h post-challenge. The diverse distribution of Es-Lectin and its enhancement by bacterial challenge indicate that C-type lectins are important in the innate immune response to bacterial infection, and can be activated for innate immune response in crab at the initial stage after pathogen infection.     

Microarray analysis of differentially expressed microRNAs in non-regressed and regressed bovine corpus luteum tissue; microRNA-378 may suppress luteal cell apoptosis by targeting the interferon gamma receptor 1 gene

MicroRNAs (miRNAs) are small non-coding endogenous RNA molecules that down-regulate the expression of target genes in a sequence-dependent manner. Recent studies indicated that miRNAs are mechanistically involved in the regulation of the mammalian corpus luteum (CL). However, few studies have profiled the different miRNA expression patterns in bovine non-regressed and regressed CL. In this study, miRNA microarray was employed to investigate the different miRNA expression patterns in bovine CL. Among the 13 differentially expressed miRNAs, seven were preferentially expressed in non-regressed CL, while six miRNAs were more highly expressed in regressed CL. Real-time RT-PCR was used to validate the microarray results. Mir-378 miRNA, known to be associated with apoptosis, was 8.54-fold (P < 0.01) up-regulated in non-regressed CL, and the interferon gamma receptor 1 (IFNGR1) gene, which potentially plays a role in apoptosis of the luteal cell, was predicted to be the target of mir-378. The results of real-time RT-PCR of mir-378 and western blot analysis of the IFNGR1 protein at different stages of CL development showed that mir-378 decreased the expression of IFNGR1 protein but not IFNGR1 mRNA. Taken together, our data support a direct role for miRNA in apoptosis of bovine CL.     

Knockdown of ribosomal protein S7 causes developmental abnormalities via p53 dependent and independent pathways in zebrafish

Ribosomal proteins (RPs), structural components of the ribosome involved in protein synthesis, are of significant importance in all organisms. Previous studies have suggested that some RPs may have other functions in addition to assembly of the ribosome. The small ribosomal subunits RPS7, has been reported to modulate the mdm2-p53 interaction. To further investigate the biological functions of RPS7, we used morpholino antisense oligonucleotides (MO) to specifically knockdown RPS7 in zebrafish. In RPS7-deficient embryos, p53 was activated, and its downstream target genes and biological events were induced, including apoptosis and cell cycle arrest. Hematopoiesis was also impaired seriously in RPS7-deficient embryos, which was confirmed by the hemoglobin O-dianisidine staining of blood cells, and the expression of scl, gata1 and α-E1 globin were abnormal. The matrix metalloproteinase (mmp) family genes were also activated in RPS7 morphants, indicating that improper cell migration might also cause development defects. Furthermore, simultaneously knockdown of the p53 protein by co-injecting a p53 MO could partially reverse the abnormal phenotype in the morphants. These results strengthen the hypothesis that specific ribosomal proteins regulate p53 and that their deficiency affects hematopoiesis. Moreover, our data implicate that RPS7 is a regulator of matrix metalloproteinase (mmp) family in zebrafish system. These specific functions of RPS7 may provide helpful clues to study the roles of RPs in human disease.     

Heterogeneity of primary glioblastoma cells in the expression of caspase-8 and the response to TRAIL-induced apoptosis

Recent studies suggest that cancer stem cells (CSCs) are responsible for cancer resistance to therapies. We therefore investigated how glioblastoma-derived CSCs respond to the treatment of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). Neurospheres were generated from glioblastomas, characterized for CSC properties including self-renewal, cell differentiation and xenograft formation capacity, and analyzed for TRAIL-induced apoptosis, CASP8 genomic status, and caspase-8 protein expression. The neurosphere NSC326 was sensitive to TRAIL-induced apoptosis as evidenced by cell death and caspase-8, -3, and -7 enzymatic activities. In contrast, however, the neurosphere NSC189 was TRAIL-resistant. G-banding analysis identified five chromosomally distinguishable cell populations in the neurospheres. Fluorescence in situ hybridization revealed the variation of chromosome 2 copy number in these populations and the loss of CASP8 locus in 2q33-34 region in a small set of cell populations in the neurosphere. Immunohistochemistry of NSC189 cell blocks revealed the lack of caspase-8 protein in a subset of neurosphere cells. Western blotting and immunohistochemistry of human glioblastoma tumors demonstrated the expression of caspase-8 protein in the vast majority of the tumors as compared to normal human brain tissues that lack the caspase-8 expression. This study shows heterogeneity of glioblastomas and derived CSCs in the genomic status of CASP8, expression of caspase-8, and thus responsiveness to TRAIL-induced apoptosis. Clinic trials may consider genomic analysis of the cancer tissue to identify the genomic loss of CASP8 and use it as a genomic marker to predict the resistance of glioblastomas to TRAIL apoptosis pathway-targeted therapies.     

Scale specific determinants of tree diversity in an old growth temperate forest in China

The major processes generating pattern in plant community composition depend upon the spatial scale and resolution of observation, therefore understanding the role played by spatial scale on species patterns is of major concern. In this study, we investigate how well environmental (topography and soil variables) and spatial variables explain variation in species composition in a 25-ha temperate forest in northeastern China. We used new variation partitioning approaches to discover the spatial scale (using multi-scale spatial PCNM variables) at which environmental heterogeneity and other spatially structured processes influence tree community composition. We also test the effect of changing grain of the study (i.e. quadrat size) on the variation partitioning results. Our results indicate that (1) species composition in the Changbai mixed broadleaf-conifer forest was controlled mainly by spatially structured soil variation at broad scales, while at finer scales most of the explained variation was of a spatial nature, pointing to the importance of biotic processes. (2) These results held at all sampling grains. However, reducing quadrat size progressively reduced both spatially and environmentally explained variance. This probably partly reflects increasing stochasticity in species abundances, and the smaller proportion of quadrats occupied by each species, when quadrat size is reduced. The results suggest that environmental heterogeneity (i.e. niche processes) and other biotic processes such as dispersal work together, but at different spatial scales, to build up diversity patterns.