Loss of the Chr16p11.2 ASD candidate gene QPRT leads to aberrant neuronal differentiation in the SH-SY5Y neuronal cell model / Denise HASLINGER in Molecular Autism, 9 (2018)  

Public ISBD [article]  inMolecular Autism > 9 (2018) . - 56p. Titre : Loss of the Chr16p11.2 ASD candidate gene QPRT leads to aberrant neuronal differentiation in the SH-SY5Y neuronal cell model Type de document : texte imprimé Auteurs : Denise HASLINGER, Auteur ; Regina WALTES, Auteur ; Afsheen YOUSAF, Auteur ; Silvia LINDLAR, Auteur ; Isabel SCHNEIDER, Auteur ; Chai K. LIM, Auteur ; Meng-Miao TSAI, Auteur ; Boyan K. GARVALOV, Auteur ; Amparo ACKER-PALMER, Auteur ; Nicolas KREZDORN, Auteur ; Björn ROTTER, Auteur ; Till ACKER, Auteur ; Gilles J. GUILLEMIN, Auteur ; Simone FULDA, Auteur ; Christine M. FREITAG, Auteur ; Andreas G. CHIOCCHETTI, Auteur Article en page(s) : 56p. Langues : Anglais (eng) Mots-clés : 16p11.2  Autism  CRISPR/Cas9  Kynurenine  Quinolinate phosphoribosyltransferase  Quinolinic acid  Sholl analysis  has been positively reviewed by the ethic's committee Frankfurt (No 267/09).All  authors agree to publish the presented work.All authors declare that they have no  competing interests.Springer Nature remains neutral with regard to jurisdictional  claims in published maps and institutional affiliations. Index. décimale : PER Périodiques Résumé : Background: Altered neuronal development is discussed as the underlying pathogenic mechanism of autism spectrum disorders (ASD). Copy number variations of 16p11.2 have recurrently been identified in individuals with ASD. Of the 29 genes within this region, quinolinate phosphoribosyltransferase (QPRT) showed the strongest regulation during neuronal differentiation of SH-SY5Y neuroblastoma cells. We hypothesized a causal relation between this tryptophan metabolism-related enzyme and neuronal differentiation. We thus analyzed the effect of QPRT on the differentiation of SH-SY5Y and specifically focused on neuronal morphology, metabolites of the tryptophan pathway, and the neurodevelopmental transcriptome. Methods: The gene dosage-dependent change of QPRT expression following Chr16p11.2 deletion was investigated in a lymphoblastoid cell line (LCL) of a deletion carrier and compared to his non-carrier parents. Expression of QPRT was tested for correlation with neuromorphology in SH-SY5Y cells. QPRT function was inhibited in SH-SY5Y neuroblastoma cells using (i) siRNA knockdown (KD), (ii) chemical mimicking of loss of QPRT, and (iii) complete CRISPR/Cas9-mediated knock out (KO). QPRT-KD cells underwent morphological analysis. Chemically inhibited and QPRT-KO cells were characterized using viability assays. Additionally, QPRT-KO cells underwent metabolite and whole transcriptome analyses. Genes differentially expressed upon KO of QPRT were tested for enrichment in biological processes and co-regulated gene-networks of the human brain. Results: QPRT expression was reduced in the LCL of the deletion carrier and significantly correlated with the neuritic complexity of SH-SY5Y. The reduction of QPRT altered neuronal morphology of differentiated SH-SY5Y cells. Chemical inhibition as well as complete KO of the gene were lethal upon induction of neuronal differentiation, but not proliferation. The QPRT-associated tryptophan pathway was not affected by KO. At the transcriptome level, genes linked to neurodevelopmental processes and synaptic structures were affected. Differentially regulated genes were enriched for ASD candidates, and co-regulated gene networks were implicated in the development of the dorsolateral prefrontal cortex, the hippocampus, and the amygdala. Conclusions: In this study, QPRT was causally related to in vitro neuronal differentiation of SH-SY5Y cells and affected the regulation of genes and gene networks previously implicated in ASD. Thus, our data suggest that QPRT may play an important role in the pathogenesis of ASD in Chr16p11.2 deletion carriers. En ligne : https://dx.doi.org/10.1186/s13229-018-0239-z Permalink : https://www.cra-rhone-alpes.org/cid/opac_css/index.php?lvl=notice_display&id=371 [article] Loss of the Chr16p11.2 ASD candidate gene QPRT leads to aberrant neuronal differentiation in the SH-SY5Y neuronal cell model [texte imprimé] / Denise HASLINGER, Auteur ; Regina WALTES, Auteur ; Afsheen YOUSAF, Auteur ; Silvia LINDLAR, Auteur ; Isabel SCHNEIDER, Auteur ; Chai K. LIM, Auteur ; Meng-Miao TSAI, Auteur ; Boyan K. GARVALOV, Auteur ; Amparo ACKER-PALMER, Auteur ; Nicolas KREZDORN, Auteur ; Björn ROTTER, Auteur ; Till ACKER, Auteur ; Gilles J. GUILLEMIN, Auteur ; Simone FULDA, Auteur ; Christine M. FREITAG, Auteur ; Andreas G. CHIOCCHETTI, Auteur . - 56p. Langues : Anglais (eng) in Molecular Autism > 9 (2018) . - 56p. Mots-clés : 16p11.2  Autism  CRISPR/Cas9  Kynurenine  Quinolinate phosphoribosyltransferase  Quinolinic acid  Sholl analysis  has been positively reviewed by the ethic's committee Frankfurt (No 267/09).All  authors agree to publish the presented work.All authors declare that they have no  competing interests.Springer Nature remains neutral with regard to jurisdictional  claims in published maps and institutional affiliations. Index. décimale : PER Périodiques Résumé : Background: Altered neuronal development is discussed as the underlying pathogenic mechanism of autism spectrum disorders (ASD). Copy number variations of 16p11.2 have recurrently been identified in individuals with ASD. Of the 29 genes within this region, quinolinate phosphoribosyltransferase (QPRT) showed the strongest regulation during neuronal differentiation of SH-SY5Y neuroblastoma cells. We hypothesized a causal relation between this tryptophan metabolism-related enzyme and neuronal differentiation. We thus analyzed the effect of QPRT on the differentiation of SH-SY5Y and specifically focused on neuronal morphology, metabolites of the tryptophan pathway, and the neurodevelopmental transcriptome. Methods: The gene dosage-dependent change of QPRT expression following Chr16p11.2 deletion was investigated in a lymphoblastoid cell line (LCL) of a deletion carrier and compared to his non-carrier parents. Expression of QPRT was tested for correlation with neuromorphology in SH-SY5Y cells. QPRT function was inhibited in SH-SY5Y neuroblastoma cells using (i) siRNA knockdown (KD), (ii) chemical mimicking of loss of QPRT, and (iii) complete CRISPR/Cas9-mediated knock out (KO). QPRT-KD cells underwent morphological analysis. Chemically inhibited and QPRT-KO cells were characterized using viability assays. Additionally, QPRT-KO cells underwent metabolite and whole transcriptome analyses. Genes differentially expressed upon KO of QPRT were tested for enrichment in biological processes and co-regulated gene-networks of the human brain. Results: QPRT expression was reduced in the LCL of the deletion carrier and significantly correlated with the neuritic complexity of SH-SY5Y. The reduction of QPRT altered neuronal morphology of differentiated SH-SY5Y cells. Chemical inhibition as well as complete KO of the gene were lethal upon induction of neuronal differentiation, but not proliferation. The QPRT-associated tryptophan pathway was not affected by KO. At the transcriptome level, genes linked to neurodevelopmental processes and synaptic structures were affected. Differentially regulated genes were enriched for ASD candidates, and co-regulated gene networks were implicated in the development of the dorsolateral prefrontal cortex, the hippocampus, and the amygdala. Conclusions: In this study, QPRT was causally related to in vitro neuronal differentiation of SH-SY5Y cells and affected the regulation of genes and gene networks previously implicated in ASD. Thus, our data suggest that QPRT may play an important role in the pathogenesis of ASD in Chr16p11.2 deletion carriers. En ligne : https://dx.doi.org/10.1186/s13229-018-0239-z Permalink : https://www.cra-rhone-alpes.org/cid/opac_css/index.php?lvl=notice_display&id=371

Protein signatures of oxidative stress response in a patient specific cell line model for autism / Andreas G. CHIOCCHETTI in Molecular Autism, (February 2014)  

Public ISBD [article]  inMolecular Autism > (February 2014) Titre : Protein signatures of oxidative stress response in a patient specific cell line model for autism Type de document : texte imprimé Auteurs : Andreas G. CHIOCCHETTI, Auteur ; Denise HASLINGER, Auteur ; Maximilian BOESCH, Auteur ; Thomas KARL, Auteur ; Stefan WIEMANN, Auteur ; Christine M. FREITAG, Auteur ; Fritz POUSTKA, Auteur ; Burghardt SCHEIBE, Auteur ; Johann BAUER, Auteur ; Helmut HINTNER, Auteur ; Michael BREITENBACH, Auteur ; Josef KELLERMANN, Auteur ; Friedrich LOTTSPEICH, Auteur ; Sabine M. KLAUCK, Auteur ; Lore BREITENBACH-KOLLER, Auteur Langues : Anglais (eng) Index. décimale : PER Périodiques Résumé : Known genetic variants can account for 10% to 20% of all cases with autism spectrum disorders (ASD). Overlapping cellular pathomechanisms common to neurons of the central nervous system (CNS) and in tissues of peripheral organs, such as immune dysregulation, oxidative stress and dysfunctions in mitochondrial and protein synthesis metabolism, were suggested to support the wide spectrum of ASD on unifying disease phenotype. Here, we studied in patient-derived lymphoblastoid cell lines (LCLs) how an ASD-specific mutation in ribosomal protein RPL10 (RPL10[H213Q]) generates a distinct protein signature. We compared the RPL10[H213Q] expression pattern to expression patterns derived from unrelated ASD patients without RPL10[H213Q] mutation. In addition, a yeast rpl10 deficiency model served in a proof-of-principle study to test for alterations in protein patterns in response to oxidative stress. Protein extracts of LCLs from patients, relatives and controls, as well as diploid yeast cells hemizygous for rpl10, were subjected to two-dimensional gel electrophoresis and differentially regulated spots were identified by mass spectrometry. Subsequently, Gene Ontology database (GO)-term enrichment and network analysis was performed to map the identified proteins into cellular pathways. The protein signature generated by RPL10[H213Q] is a functionally related subset of the ASD-specific protein signature, sharing redox-sensitive elements in energy-, protein- and redox-metabolism. In yeast, rpl10 deficiency generates a specific protein signature, harboring components of pathways identified in both the RPL10[H213Q] subjects' and the ASD patients' set. Importantly, the rpl10 deficiency signature is a subset of the signature resulting from response of wild-type yeast to oxidative stress. Redox-sensitive protein signatures mapping into cellular pathways with pathophysiology in ASD have been identified in both LCLs carrying the ASD-specific mutation RPL10[H213Q] and LCLs from ASD patients without this mutation. At pathway levels, this redox-sensitive protein signature has also been identified in a yeast rpl10 deficiency and an oxidative stress model. These observations point to a common molecular pathomechanism in ASD, characterized in our study by dysregulation of redox balance. Importantly, this can be triggered by the known ASD-RPL10[H213Q] mutation or by yet unknown mutations of the ASD cohort that act upstream of RPL10 in differential expression of redox-sensitive proteins. En ligne : http://dx.doi.org/10.1186/2040-2392-5-10 Permalink : https://www.cra-rhone-alpes.org/cid/opac_css/index.php?lvl=notice_display&id=227 [article] Protein signatures of oxidative stress response in a patient specific cell line model for autism [texte imprimé] / Andreas G. CHIOCCHETTI, Auteur ; Denise HASLINGER, Auteur ; Maximilian BOESCH, Auteur ; Thomas KARL, Auteur ; Stefan WIEMANN, Auteur ; Christine M. FREITAG, Auteur ; Fritz POUSTKA, Auteur ; Burghardt SCHEIBE, Auteur ; Johann BAUER, Auteur ; Helmut HINTNER, Auteur ; Michael BREITENBACH, Auteur ; Josef KELLERMANN, Auteur ; Friedrich LOTTSPEICH, Auteur ; Sabine M. KLAUCK, Auteur ; Lore BREITENBACH-KOLLER, Auteur. Langues : Anglais (eng) in Molecular Autism > (February 2014) Index. décimale : PER Périodiques Résumé : Known genetic variants can account for 10% to 20% of all cases with autism spectrum disorders (ASD). Overlapping cellular pathomechanisms common to neurons of the central nervous system (CNS) and in tissues of peripheral organs, such as immune dysregulation, oxidative stress and dysfunctions in mitochondrial and protein synthesis metabolism, were suggested to support the wide spectrum of ASD on unifying disease phenotype. Here, we studied in patient-derived lymphoblastoid cell lines (LCLs) how an ASD-specific mutation in ribosomal protein RPL10 (RPL10[H213Q]) generates a distinct protein signature. We compared the RPL10[H213Q] expression pattern to expression patterns derived from unrelated ASD patients without RPL10[H213Q] mutation. In addition, a yeast rpl10 deficiency model served in a proof-of-principle study to test for alterations in protein patterns in response to oxidative stress. Protein extracts of LCLs from patients, relatives and controls, as well as diploid yeast cells hemizygous for rpl10, were subjected to two-dimensional gel electrophoresis and differentially regulated spots were identified by mass spectrometry. Subsequently, Gene Ontology database (GO)-term enrichment and network analysis was performed to map the identified proteins into cellular pathways. The protein signature generated by RPL10[H213Q] is a functionally related subset of the ASD-specific protein signature, sharing redox-sensitive elements in energy-, protein- and redox-metabolism. In yeast, rpl10 deficiency generates a specific protein signature, harboring components of pathways identified in both the RPL10[H213Q] subjects' and the ASD patients' set. Importantly, the rpl10 deficiency signature is a subset of the signature resulting from response of wild-type yeast to oxidative stress. Redox-sensitive protein signatures mapping into cellular pathways with pathophysiology in ASD have been identified in both LCLs carrying the ASD-specific mutation RPL10[H213Q] and LCLs from ASD patients without this mutation. At pathway levels, this redox-sensitive protein signature has also been identified in a yeast rpl10 deficiency and an oxidative stress model. These observations point to a common molecular pathomechanism in ASD, characterized in our study by dysregulation of redox balance. Importantly, this can be triggered by the known ASD-RPL10[H213Q] mutation or by yet unknown mutations of the ASD cohort that act upstream of RPL10 in differential expression of redox-sensitive proteins. En ligne : http://dx.doi.org/10.1186/2040-2392-5-10 Permalink : https://www.cra-rhone-alpes.org/cid/opac_css/index.php?lvl=notice_display&id=227

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