Repertoire of microRNAs in Epithelial Ovarian Cancer as Determined by Next Generation Sequencing of Small RNA cDNA Libraries

MicroRNAs (miRNAs) are small regulatory RNAs that are implicated in cancer pathogenesis and have recently shown promise as blood-based biomarkers for cancer detection. Epithelial ovarian cancer is a deadly disease for which improved outcomes could be achieved by successful early detection and enhanced understanding of molecular pathogenesis that leads to improved therapies. A critical step toward these goals is to establish a comprehensive view of miRNAs expressed in epithelial ovarian cancer tissues as well as in normal ovarian surface epithelial cells.We used massively parallel pyrosequencing (i.e., "454 sequencing") to discover and characterize novel and known miRNAs expressed in primary cultures of normal human ovarian surface epithelium (HOSE) and in tissue from three of the most common histotypes of ovarian cancer. Deep sequencing of small RNA cDNA libraries derived from normal HOSE and ovarian cancer samples yielded a total of 738,710 high-quality sequence reads, generating comprehensive digital profiles of miRNA expression. Expression profiles for 498 previously annotated miRNAs were delineated and we discovered six novel miRNAs and 39 candidate miRNAs. A set of 124 miRNAs was differentially expressed in normal versus cancer samples and 38 miRNAs were differentially expressed across histologic subtypes of ovarian cancer. Taqman qRT-PCR performed on a subset of miRNAs confirmed results of the sequencing-based study.This report expands the body of miRNAs known to be expressed in epithelial ovarian cancer and provides a useful resource for future studies of the role of miRNAs in the pathogenesis and early detection of ovarian cancer

Balanced translocation linked to psychiatric disorder, glutamate and cortical structure/function

Rare genetic variants of large effect can help elucidate the pathophysiology of brain disorders. Here we expand the clinical and genetic analyses of a family with a (1;11)(q42;q14.3) translocation multiply affected by major psychiatric illness and test the effect of the translocation on the structure and function of prefrontal, and temporal brain regions. The translocation showed significant linkage (LOD score 6.1) with a clinical phenotype that included schizophrenia, schizoaffective disorder, bipolar disorder, and recurrent major depressive disorder. Translocation carriers showed reduced cortical thickness in the left temporal lobe, which correlated with general psychopathology and positive psychotic symptom severity. They showed reduced gyrification in prefrontal cortex, which correlated with general psychopathology severity. Translocation carriers also showed significantly increased activation in the caudate nucleus on increasing verbal working memory load, as well as statistically significant reductions in the right dorsolateral prefrontal cortex glutamate concentrations. These findings confirm that the t(1;11) translocation is associated with a significantly increased risk of major psychiatric disorder and suggest a general vulnerability to psychopathology through altered cortical structure and function, and decreased glutamate levels

Joining S100 proteins and migration:for better or for worse, in sickness and in health

The vast diversity of S100 proteins has demonstrated a multitude of biological correlations with cell growth, cell differentiation and cell survival in numerous physiological and pathological conditions in all cells of the body. This review summarises some of the reported regulatory functions of S100 proteins (namely S100A1, S100A2, S100A4, S100A6, S100A7, S100A8/S100A9, S100A10, S100A11, S100A12, S100B and S100P) on cellular migration and invasion, established in both culture and animal model systems and the possible mechanisms that have been proposed to be responsible. These mechanisms involve intracellular events and components of the cytoskeletal organisation (actin/myosin filaments, intermediate filaments and microtubules) as well as extracellular signalling at different cell surface receptors (RAGE and integrins). Finally, we shall attempt to demonstrate how aberrant expression of the S100 proteins may lead to pathological events and human disorders and furthermore provide a rationale to possibly explain why the expression of some of the S100 proteins (mainly S100A4 and S100P) has led to conflicting results on motility, depending on the cells used. © 2013 Springer Basel

Biological function in the twilight zone of sequence conservation

Abstract Strong DNA conservation among divergent species is an indicator of enduring functionality. With weaker sequence conservation we enter a vast ‘twilight zone’ in which sequence subject to transient or lower constraint cannot be distinguished easily from neutrally evolving, non-functional sequence. Twilight zone functional sequence is illuminated instead by principles of selective constraint and positive selection using genomic data acquired from within a species’ population. Application of these principles reveals that despite being biochemically active, most twilight zone sequence is not functional

Advancing schizophrenia drug discovery : optimizing rodent models to bridge the translational gap

Although our knowledge of the pathophysiology of schizophrenia has increased, treatments for this devastating illness remain inadequate. Here, we critically assess rodent models and behavioural end points used in schizophrenia drug discovery and discuss why these have not led to improved treatments. We provide a perspective on how new models, based on recent advances in the understanding of the genetics and neural circuitry underlying schizophrenia, can bridge the translational gap and lead to the development of more effective drugs. We conclude that previous serendipitous approaches should be replaced with rational strategies for drug discovery in integrated preclinical and clinical programmes. Validation of drug targets in disease-based models that are integrated with translationally relevant end point assessments will reduce the current attrition rate in schizophrenia drug discovery and ultimately lead to therapies that tackle the disease process

Xenopus phospho-CDK7/cyclin H expressed in baculoviral-infected insect cells.

The cyclin-dependent kinase-activating kinase (CAK) catalyzes the phosphorylation of the cyclin-dependent protein kinases (CDKs) on a threonine residue (Thr160 in human CDK2). The reaction is an obligatory step in the activation of the CDKs. In higher eukaryotes, the CAK complex has been characterized in two forms. The first consists of three subunits, namely CDK7, cyclin H, and an assembly factor called MAT1, while the second consists of phospho-CDK7 and cyclin H. Phosphorylation of CDK7 is essential for cyclin association and kinase activity in the absence of the assembly factor MAT1. The Xenopus laevis CDK7 phosphorylation sites are located on the activation segment of the kinase at residues Ser170 and at Thr176 (the latter residue corresponding to Thr160 in human CDK2). We report the expression and purification of X. laevis CDK7/cyclin H binary complex in insect cells through coinfection with the recombinant viruses, AcCDK7 and Accyclin H. Quantities suitable for crystallization trials have been obtained. The purified CDK7/cyclin H binary complex phosphorylated CDK2 and CDK2/cyclin A but did not phosphorylate histone H1 or peptide substrates based on the activation segments of CDK7 and CDK2. Analysis by mass spectrometry showed that coexpression of CDK7 with cyclin H in baculoviral-infected insect cells results in phosphorylation of residues Ser170 and Thr176 in CDK7. It is assumed that phosphorylation is promoted by kinase(s) in the insect cells that results in the correct, physiologically significant posttranslational modification. We discuss the occurrence of in vivo phosphorylation of proteins expressed in baculoviral-infected insect cells

Population Genomics on the Fly: Recent Advances in Drosophila

Drosophila melanogaster, a small dipteran of African origin, represents one of the best-studied model organisms. Early work in this system has uniquely shed light on the basic principles of genetics and resulted in a versatile collection of genetic tools that allow to uncover mechanistic links between genotype and phenotype. Moreover, given its worldwide distribution in diverse habitats and its moderate genome-size, Drosophila has proven very powerful for population genetics inference and was one of the first eukaryotes whose genome was fully sequenced. In this book chapter, we provide a brief historical overview of research in Drosophila and then focus on recent advances during the genomic era. After describing different types and sources of genomic data, we discuss mechanisms of neutral evolution including the demographic history of Drosophila and the effects of recombination and biased gene conversion. Then, we review recent advances in detecting genome-wide signals of selection, such as soft and hard selective sweeps. We further provide a brief introduction to background selection, selection of noncoding DNA and codon usage and focus on the role of structural variants, such as transposable elements and chromosomal inversions, during the adaptive process. Finally, we discuss how genomic data helps to dissect neutral and adaptive evolutionary mechanisms that shape genetic and phenotypic variation in natural populations along environmental gradients. In summary, this book chapter serves as a starting point to Drosophila population genomics and provides an introduction to the system and an overview to data sources, important population genetic concepts and recent advances in the field