Nuclear S-nitrosylation impacts tissue regeneration in zebrafish.

Despite the importance of nitric oxide signaling in multiple biological processes, its role in tissue regeneration remains largely unexplored. Here, we provide evidence that inducible nitric oxide synthase (iNos) translocates to the nucleus during zebrafish tailfin regeneration and is associated with alterations in the nuclear S-nitrosylated proteome. iNos inhibitors or nitric oxide scavengers reduce protein S-nitrosylation and impair tailfin regeneration.

Nuclear -Nitrosylation Defines an Optimal Zone for Inducing Pluripotency.

Background: We found that cell-autonomous innate immune signaling causes global changes in the expression of epigenetic modifiers to facilitate nuclear reprogramming to pluripotency. A role of -nitrosylation by inducible nitric oxide (NO) synthase, an important effector of innate immunity, has been previously described in the transdifferentiation of fibroblasts to endothelial cells. Accordingly, we hypothesized that -nitrosylation might also have a role in nuclear reprogramming to pluripotency.

Increase in Local Protein Concentration by Field-Inversion Gel Electrophoresis.

Proteins that migrate through cross-linked polyacrylamide gels (PAGs) under the influence of a constant electric field experience negative factors, such as diffusion and nonspecific trapping in the gel matrix. These negative factors reduce protein concentrations within a defined gel volume with increasing migration distance and, therefore, decrease protein recovery efficiency. Here, we describe the enhancement of protein separation efficiency for up to twofold in conventional one-dimensional PAG electrophoresis (1D PAGE), two-dimensional (2D) PAGE, and native PAGE by implementing pulses of inverted electric field during gel electrophoresis.

CLN8 is an endoplasmic reticulum cargo receptor that regulates lysosome biogenesis.

Organelle biogenesis requires proper transport of proteins from their site of synthesis to their target subcellular compartment. Lysosomal enzymes are synthesized in the endoplasmic reticulum (ER) and traffic through the Golgi complex before being transferred to the endolysosomal system, but how they are transferred from the ER to the Golgi is unknown. Here, we show that ER-to-Golgi transfer of lysosomal enzymes requires CLN8, an ER-associated membrane protein whose loss of function leads to the lysosomal storage disorder, neuronal ceroid lipofuscinosis 8 (a type of Batten disease).

Development and rescue of human familial hypercholesterolaemia in a xenograft mouse model.

Diseases of lipid metabolism are a major cause of human morbidity, but no animal model entirely recapitulates human lipoprotein metabolism. Here we develop a xenograft mouse model using hepatocytes from a patient with familial hypercholesterolaemia caused by loss-of-function mutations in the low-density lipoprotein receptor (LDLR). Like familial hypercholesterolaemia patients, our familial hypercholesterolaemia liver chimeric mice develop hypercholesterolaemia and a 'humanized' serum profile, including expression of the emerging drug targets cholesteryl ester transfer protein and apolipoprotein (a), for which no genes exist in mice.

Silencing BMI1 eliminates tumor formation of pediatric glioma CD133+ cells not by affecting known targets but by down-regulating a novel set of core genes.

Clinical outcome of children with malignant glioma remains dismal. Here, we examined the role of over-expressed BMI1, a regulator of stem cell self-renewal, in sustaining tumor formation in pediatric glioma stem cells. Our investigation revealed BMI1 over-expression in 29 of 54 (53.

Genome-wide analysis of DNA copy number alterations and loss of heterozygosity in intracranial germ cell tumors.

Backgrounds: Intracranial germ cell tumors (GCTs) are rare and heterogeneous with very little is known about their pathogenesis and underlying genetic abnormalities.

Procedures: In order to identify candidate genes and pathways which are involved in the pathogenesis of these tumors, we have profiled 62 intracranial GCTs for DNA copy number alterations (CNAs) and loss of heterozygosity (LOH) by using single nucleotide polymorphism (SNP) array and quantitative real time PCR (qPCR).

Results: Initially 27 cases of tumor tissues with matched blood samples were fully analyzed by SNP microarray and qPCR.

Immunodepletion plasma proteomics by tripleTOF 5600 and Orbitrap elite/LTQ-Orbitrap Velos/Q exactive mass spectrometers.

Plasma proteomic experiments performed rapidly and economically using several of the latest high-resolution mass spectrometers were compared. Four quantitative hyperfractionated plasma proteomics experiments were analyzed in replicates by two AB SCIEX TripleTOF 5600 and three Thermo Scientific Orbitrap (Elite/LTQ-Orbitrap Velos/Q Exactive) instruments. Each experiment compared two iTRAQ isobaric-labeled immunodepleted plasma proteomes, provided as 30 labeled peptide fractions, and 480 LC-MS/MS runs delivered >250 GB of data in 2 months.

Histone induced platelet aggregation is inhibited by normal albumin.

Introduction: Histones are small, nuclear proteins that serve to package DNA. Recent reports suggest that extracellular histones, including histone H4, may contribute to the pathogenesis of sepsis; they promote platelet aggregation and thrombosis when released into the circulation during inflammation or cell death. The mechanisms by which the body minimizes the deleterious effects of circulating histones are unclear.

Curcumin modulates DNA methylation in colorectal cancer cells.

Aim: Recent evidence suggests that several dietary polyphenols may exert their chemopreventive effect through epigenetic modifications. Curcumin is one of the most widely studied dietary chemopreventive agents for colon cancer prevention, however, its effects on epigenetic alterations, particularly DNA methylation, remain unclear. Using systematic genome-wide approaches, we aimed to elucidate the effect of curcumin on DNA methylation alterations in colorectal cancer cells.

Global gene expression profiling confirms the molecular fidelity of primary tumor-based orthotopic xenograft mouse models of medulloblastoma.

We previously showed that primary tumor-based orthotopic xenograft mouse models of medulloblastoma replicated the histopathological phenotypes of patients' original tumors. Here, we performed global gene expression profiling of 11 patient-specific xenograft models to further determine whether the xenograft tumors were molecularly accurate during serial subtransplantations in mouse brains and whether they represented all the molecular subtypes of medulloblastoma that were recently described. Analysis of the transcriptomes of 9 pairs of matched passage I xenografts and patients' tumors revealed high correlation coefficients (r(2) > 0.

Boswellic acid induces epigenetic alterations by modulating DNA methylation in colorectal cancer cells.

Accumulating evidence suggests that chemopreventive effects of some dietary polyphenols may in part be mediated by their ability to influence epigenetic mechanisms in cancer cells. Boswellic acids, derived from the plant Boswellia serrata, have long been used for the treatment of various inflammatory diseases due to their potent anti-inflammatory activities. Recent preclinical studies have also suggested that this compound has anti-cancer potential against various malignancies.

De novo constitutional MLH1 epimutations confer early-onset colorectal cancer in two new sporadic Lynch syndrome cases, with derivation of the epimutation on the paternal allele in one.

Lynch syndrome is an autosomal dominant cancer predisposition syndrome classically caused by germline mutations of the mismatch repair genes, MLH1, MSH2, MSH6 and PMS2. Constitutional epimutations of the MLH1 gene, characterized by soma-wide methylation of a single allele of the promoter and allelic transcriptional silencing, have been identified in a subset of Lynch syndrome cases lacking a sequence mutation in MLH1. We report two individuals with no family history of colorectal cancer who developed that disease at age 18 and 20 years.

Solid-phase profiling of proteins.

Protein array technologies refer to any fabrication and use of any arrays containing multiple proteins captured on solid surfaces. This valuable tool is used for high-throughput protein-protein interaction studies, protein marker discovery and other applications. This unit provides basic protocols for biomarker discovery and protein-protein interactions.

Recursive SVM feature selection and sample classification for mass-spectrometry and microarray data.

Background: Like microarray-based investigations, high-throughput proteomics techniques require machine learning algorithms to identify biomarkers that are informative for biological classification problems. Feature selection and classification algorithms need to be robust to noise and outliers in the data.

Results: We developed a recursive support vector machine (R-SVM) algorithm to select important genes/biomarkers for the classification of noisy data.