Correction: Brain region-specific alterations of RNA editing in PDE8A mRNA in suicide decedents.

Author forgot to attach a supplementary doc file which includes the supplementary methods and supplementary figure legends.

Brain region-specific alterations of RNA editing in PDE8A mRNA in suicide decedents.

Phosphodiesterases (PDE) are key modulators of signal transduction and are involved in inflammatory cell activation, memory and cognition. There is a two-fold decrease in the expression of phosphodiesterase 8A (PDE8A) in the temporal cortex of major depressive disorder (MDD) patients. Here, we studied PDE8A mRNA-editing profile in two architectonically distinct neocortical regions in a clinically well-characterized cohort of age- and sex-matched non-psychiatric drug-free controls and depressed suicide decedents. By using capillary electrophoresis single-stranded conformational polymorphism (CE-SSCP), a previously validated technique to identify A-to-I RNA modifications, we report the full editing profile of PDE8A in the brain, including identification of two novel editing sites. Editing of PDE8A mRNA displayed clear regional difference when comparing dorsolateral prefrontal cortex (BA9) and anterior cingulate cortex (BA24). Furthermore, we report significant intra-regional differences between non-psychiatric control individuals and depressed suicide decedents, which could discriminate the two populations. Taken together, our results (i) highlight the importance of immune/inflammatory markers in major depressive disorder and suicide and (ii) establish a direct relationship between A-to-I RNA modifications of peripheral markers and A-to-I RNA editing-related modifications in brain. This work provides the first immune response-related brain marker for suicide and could pave the way for the identification of a blood-based biomarker that predicts suicidal behavior.

Genome engineering empowers the diatom Phaeodactylum tricornutum for biotechnology.

Diatoms, a major group of photosynthetic microalgae, have a high biotechnological potential that has not been fully exploited because of the paucity of available genetic tools. Here we demonstrate targeted and stable modifications of the genome of the marine diatom Phaeodactylum tricornutum, using both meganucleases and TALE nucleases. When nuclease-encoding constructs are co-transformed with a selectable marker, high frequencies of genome modifications are readily attained with 56 and 27% of the colonies exhibiting targeted mutagenesis or targeted gene insertion, respectively. The generation of an enhanced lipid-producing strain (45-fold increase in triacylglycerol accumulation) through the disruption of the UDP-glucose pyrophosphorylase gene exemplifies the power of genome engineering to harness diatoms for biofuel production.

In vitro screening for drug-induced depression and/or suicidal adverse effects: a new toxicogenomic assay based on CE-SSCP analysis of HTR2C mRNA editing in SH-SY5Y cells.

Many drugs in clinical trials, or already on the market, can have psychiatric side effects, independently of their therapeutic indication (e.g., Acomplia, Taranabant, and Roaccutane). There is currently no in vitro or in vivo approved test for the detection/prediction of such adverse effects, and the Food and Drugs Administration (FDA) can only issue general alerts on specific therapeutic classes. The development of a screening assay is therefore of real interest. The anti-viral and anti-tumor action of human interferon-alpha (hIFNα) is associated with a variety of neuropsychiatric side effects, including major depression, suicidal ideation and suicide. RNA editing of the serotonin 2C receptor (HTR2C) by adenosine deaminases acting on RNA (ADARs) is a post-transcriptional modification, the regulation of which is altered in depressed suicide victims. In this study, we show that in the SH-SY5Y neuroblastoma cell line, hIFNα specifically activates the ADAR1a isoform and thereby modifies the HTR2C mRNA editing profile. As this hIFNα-induced altered profile partly overlaps with that observed in the brain of depressed suicide victims, we investigated whether it could be used as a signature to identify drugs with depression and/or suicidal side effects. By means of the Biocortech proprietary screening assay, which allows the relative quantification of all the edited HTR2C isoforms in a sample, we blind-tested the effect of 50 marketed molecules on HTR2C mRNA editing in SH-SY5Y cells and identified 17 compounds with an IFN-like editing profile. This new toxicogenomic assay can identify compounds with potential psychiatric adverse events with a positive predictive value of 90 %.

In vitro and in vivo pharmacological profile of AS057278, a selective d-amino acid oxidase inhibitor with potential anti-psychotic properties.

Non-competitive N-methyl-d-aspartate (NMDA) blockers induce schizophrenic-like behavior in healthy volunteers and exacerbate symptomatology in schizophrenic patients. Hence, a compound able to enhance NMDA neurotransmission by increasing levels of d-serine, an endogenous full agonist at the glycine site of the NMDA receptors, could have anti-psychotic activity. One way to increase d-serine levels is the inhibition of d-amino acid oxidase (DAAO), the enzyme responsible for d-serine oxidation. Indeed AS057278, a potent in vitro (IC(50)=0.91 microM) and ex vivo (ED(50)=2.2-3.95 microM) DAAO inhibitor, was able to increase d-serine fraction in rat cortex and midbrain (10 mg/kg i.v.). AS057278 was able to normalize phencyclidine (PCP)-induced prepulse inhibition after acute (80 mg/kg) and chronic (20 mg/kg b.i.d.) oral administration in mice. Finally, AS057278 after oral chronic treatment (10 mg/kg b.i.d.) was able to normalize PCP-induced hyperlocomotion. These results suggest that AS057278 has the potential to anti-psychotic action toward both cognitive and positive symptoms of schizophrenia.

Identification of new drug sensitivity genes using genetic suppressor elements: protein arginine N-methyltransferase mediates cell sensitivity to DNA-damaging agents.

Genetic suppressor elements (GSEs) are cDNA fragments encoding either truncated proteins, acting as dominant-negative mutants, or inhibitory antisense RNA segments counteracting with the gene from which they are derived. To identify genes controlling the cell response to cytotoxic agents, a normalized retroviral library of randomly fragmented cDNAs from Chinese hamster cell line DC-3F was screened for GSEs conferring resistance to the topoisomerase II inhibitor 9-OH-ellipticine. From 218 cDNA fragments isolated, 11 functional GSEs, corresponding to at least 8 independent genes, were selected. The gene corresponding to the most abundant GSE encodes two proteins, p77 and p82, highly homologous to proteins detected in various species and carrying the sequence motifs characteristic of the protein arginine N-methyltransferase family. Furthermore, a methylase activity was observed on myelin basic protein in immunoprecipitates of hemagglutinin-tagged p77 and p82. Therefore, p77 and p82 are the first identified members of a new protein arginine N-methyltransferase family. A decreased expression of these enzymes is associated with either resistance or hypersensitivity to a broad range of DNA-damaging agents. Our data indicate that down-regulation of these enzymes in the GSE-expressing cells would alter one or several steps downstream of the drug-target interaction in the drug-response pathway.

Genetic and physiological data implicating the new human gene G72 and the gene for D-amino acid oxidase in schizophrenia.

A map of 191 single-nucleotide polymorphism (SNPs) was built across a 5-Mb segment from chromosome 13q34 that has been genetically linked to schizophrenia. DNA from 213 schizophrenic patients and 241 normal individuals from Canada were genotyped with this marker set. Two 1,400- and 65-kb regions contained markers associated with the disease. Two markers from the 65-kb region were also found to be associated to schizophrenia in a Russian sample. Two overlapping genes G72 and G30 transcribed in brain were experimentally annotated in this 65-kb region. Transfection experiments point to the existence of a 153-aa protein coded by the G72 gene. This protein is rapidly evolving in primates, is localized to endoplasmic reticulum/Golgi in transfected cells, is able to form multimers and specifically binds to carbohydrates. Yeast two-hybrid experiments with the G72 protein identified the enzyme d-amino acid oxidase (DAAO) as an interacting partner. DAAO is expressed in human brain where it oxidizes d-serine, a potent activator of N-methyl-D-aspartate type glutamate receptor. The interaction between G72 and DAAO was confirmed in vitro and resulted in activation of DAAO. Four SNP markers from DAAO were found to be associated with schizophrenia in the Canadian samples. Logistic regression revealed genetic interaction between associated SNPs in vicinity of two genes. The association of both DAAO and a new gene G72 from 13q34 with schizophrenia together with activation of DAAO activity by a G72 protein product points to the involvement of this N-methyl-d-aspartate receptor regulation pathway in schizophrenia.

Identification and characterization of Moca-cyp. A Drosophila melanogaster nuclear cyclophilin.

Cyclophilins are enzymes catalyzing the cis-trans isomerization of peptidyl-prolyl bonds and belong to the enzyme class of peptidyl-prolyl cis-trans isomerases (PPIases), which includes two more families (FK506 binding proteins and parvulins). We report the characterization of a novel cyclophilin (Moca-cyp) isolated from Drosophila melanogaster. The single-copy Moca-cyp gene, which is localized on chromosome 3R, was cloned and sequenced. The sequence alignment of the gene against Moca-cyp cDNA allowed us to define its intron/exon structure and to identify a variant cDNA corresponding to an alternatively spliced mRNA. By embryo in situ RNA hybridization and immunostaining, we show that the expression of Moca-cyp is regulated during embryogenesis of Drosophila. The 120-kDa nuclear Moca-cyp protein belongs to a subfamily of large cyclophilins sharing structural and enzymatic features: their highly conserved N-terminal PPIase domain is extended by a positively charged and divergent C-terminal tail. Compared with cyclophilin 18, the enzymatic activity carried by the PPIase domain of Moca-cyp is low, exhibits characteristic substrate specificity, and shows a reduced sensitivity to the drug cyclosporin A (CsA). The reduced affinity for CsA is one of the typical features linking members of this subfamily and is probably the consequence of two amino acid substitutions within their active site. Another structural feature shared by members of this subfamily is a conserved polypeptidic segment ("moca" domain) that we report for the first time. The moca domain is located within the C-terminal tail and is the exclusive hallmark of a group of large cyclophilins found in multicellular organisms of the animal kingdom.