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1.
Nat Commun ; 11(1): 198, 2020 01 10.
Article in English | MEDLINE | ID: mdl-31924792

ABSTRACT

The neural crest gives rise to numerous cell types, dysfunction of which contributes to many disorders. Here, we report that adenosine deaminase acting on RNA (ADAR1), responsible for adenosine-to-inosine editing of RNA, is required for regulating the development of two neural crest derivatives: melanocytes and Schwann cells. Neural crest specific conditional deletion of Adar1 in mice leads to global depigmentation and absence of myelin from peripheral nerves, resulting from alterations in melanocyte survival and differentiation of Schwann cells, respectively. Upregulation of interferon stimulated genes precedes these defects, which are associated with the triggering of a signature resembling response to injury in peripheral nerves. Simultaneous extinction of MDA5, a key sensor of unedited RNA, rescues both melanocytes and myelin defects in vitro, suggesting that ADAR1 safeguards neural crest derivatives from aberrant MDA5-mediated interferon production. We thus extend the landscape of ADAR1 function to the fields of neural crest development and disease.


Subject(s)
Adenosine Deaminase/metabolism , Melanocytes/metabolism , Neural Crest/metabolism , Schwann Cells/metabolism , Adenosine Deaminase/genetics , Animals , Cell Differentiation , Disease Models, Animal , Gene Deletion , Gene Expression Regulation, Developmental , Heart , Interferons/metabolism , Mice , Mice, Knockout , Neurogenesis , RNA Editing , Sciatic Nerve/cytology , Skin/pathology , Transcriptome , Up-Regulation
2.
Hum Mol Genet ; 24(17): 4933-47, 2015 Sep 01.
Article in English | MEDLINE | ID: mdl-26060192

ABSTRACT

SOX10 is a transcription factor with well-known functions in neural crest and oligodendrocyte development. Mutations in SOX10 were first associated with Waardenburg-Hirschsprung disease (WS4; deafness, pigmentation defects and intestinal aganglionosis). However, variable phenotypes that extend beyond the WS4 definition are now reported. The neurological phenotypes associated with some truncating mutations are suggested to be the result of escape from the nonsense-mediated mRNA decay pathway; but, to date, no mechanism has been suggested for missense mutations, of which approximately 20 have now been reported, with about half of the latter shown to be redistributed to nuclear bodies of undetermined nature and function in vitro. Here, we report that p54NRB, which plays a crucial role in the regulation of gene expression during many cellular processes including differentiation, interacts synergistically with SOX10 to regulate several target genes. Interestingly, this paraspeckle protein, as well as two other members of the Drosophila behavior human splicing (DBHS) protein family, co-localize with SOX10 mutants in nuclear bodies, suggesting the possible paraspeckle nature of these foci or re-localization of the DBHS members to other subnuclear compartments. Remarkably, the co-transfection of wild-type and mutant SOX10 constructs led to the sequestration of wild-type protein in mutant-induced foci. In contrast to mutants presenting with additional cytoplasmic re-localization, those exclusively found in the nucleus alter synergistic activity between SOX10 and p54NRB. We propose that such a dominant negative effect may contribute to or be at the origin of the unique progressive and severe neurological phenotype observed in affected patients.


Subject(s)
Genetic Association Studies , Mutation, Missense , Nuclear Matrix-Associated Proteins/metabolism , Octamer Transcription Factors/metabolism , Phenotype , RNA-Binding Proteins/metabolism , SOXE Transcription Factors/genetics , SOXE Transcription Factors/metabolism , Cell Line , Cell Nucleus/metabolism , DNA-Binding Proteins , Gene Expression , Humans , Melanoma/genetics , Melanoma/metabolism , Nuclear Matrix-Associated Proteins/genetics , Octamer Transcription Factors/genetics , Protein Binding , Protein Transport , RNA-Binding Proteins/genetics , Waardenburg Syndrome/genetics , Waardenburg Syndrome/metabolism
3.
Hum Mutat ; 35(3): 303-7, 2014 Mar.
Article in English | MEDLINE | ID: mdl-24357527

ABSTRACT

A deletion encompassing several SOX10 enhancers was recently identified in a patient presenting with Waardenburg syndrome type 4 (WS4), which is defined as a combination of Hirschsprung disease (HSCR, intestinal aganglionosis) and WS (deafness and pigmentation defects). The expression patterns of some of the known SOX10 enhancers in animal models led to the speculation that endophenotypes of WS4 may be linked to mutations within some of these sequences. The present study investigated deletions and point mutations within four SOX10 enhancers in 144 unexplained isolated HSCR cases. One deletion and two point mutations affecting binding sites for known neural crest transcription factors were identified. In vitro functional analysis revealed that the first point mutation disrupts autoregulation by SOX10, whereas the second affects AP2a and SOX10 synergistic activity. The present findings suggest that the mutations within SOX10 enhancers contribute to isolated HSCR.


Subject(s)
Regulatory Sequences, Nucleic Acid , SOXE Transcription Factors/genetics , Waardenburg Syndrome/genetics , Base Sequence , Female , Hirschsprung Disease , Humans , Infant , Male , Molecular Sequence Data , Point Mutation , Sequence Deletion , Transcription Factors/genetics
4.
J Nutr Biochem ; 24(3): 586-94, 2013 Mar.
Article in English | MEDLINE | ID: mdl-22818713

ABSTRACT

Cardiac hypertrophy is the main response of the heart to various extrinsic and intrinsic stimuli, and it is characterized by specific molecular and phenotypic changes. Recent in vitro and in vivo studies indicate the involvement of reactive oxygen species in the hypertrophic response. In this study, silibinin, a plant flavonolignan extracted from milk thistle with potent antioxidant activity, was evaluated for its effects in (a) preventing hydrogen peroxide (H2O2)-induced cellular damage and (b) blocking the phenylephrine-induced hypertrophic response. Using the in vitro model of embryonic rat heart-derived H9c2 cells, we showed that silibinin has a rather safe profile as concentrations up to 200µM did not affect cell viability. Pretreatment of H9c2 cells with silibinin resulted in better protection of H9c2 cells under conditions of H2O2-induced cellular stress compared to untreated cells as indicated by cell viability and DNA fragmentation assays. Furthermore, silibinin attenuated the phenylephrine-induced hypertrophic response as evidenced by the measurement of cell surface, up-regulation of atrial natriuretic peptide and increase of cellular protein levels. Moreover, silibinin repressed the phenylephrine-induced phosphorylation of ERK1/2 kinases, while it appeared to inhibit the weakly activated by phenylephrine phosphorylation of Akt. Based on our results, silibinin may attenuate the phenylephrine-induced hypertrophic response of H9c2 cells via antioxidant mechanisms involving mainly the inhibition of the intracellular signaling pathways mediated by ERK1/2 MAPKs and Akt.


Subject(s)
Cardiomegaly/drug therapy , Heart/drug effects , Oxidative Stress/drug effects , Phenylephrine/adverse effects , Plant Extracts/pharmacology , Silymarin/pharmacology , Animals , Antioxidants , Cardiomegaly/chemically induced , Cardiomegaly/pathology , Carrier Proteins/genetics , Carrier Proteins/metabolism , Cell Line , Cell Survival , DNA Fragmentation , Heart/physiology , Hydrogen Peroxide/adverse effects , Hydrogen Peroxide/metabolism , Silybum marianum/chemistry , Mitogen-Activated Protein Kinase 1/genetics , Mitogen-Activated Protein Kinase 1/metabolism , Mitogen-Activated Protein Kinase 3/genetics , Mitogen-Activated Protein Kinase 3/metabolism , Phosphorylation , Proto-Oncogene Proteins c-akt/genetics , Proto-Oncogene Proteins c-akt/metabolism , Rats , Reactive Oxygen Species , Reverse Transcriptase Polymerase Chain Reaction , Rhodamines/analysis , Rhodamines/metabolism , Signal Transduction , Silybin , Up-Regulation
5.
Atherosclerosis ; 220(1): 257-64, 2012 Jan.
Article in English | MEDLINE | ID: mdl-22056215

ABSTRACT

AIM: Recently, the concept that high density lipoprotein (HDL) quality is an important parameter for atheroprotection is gaining ground, though little data exists so far to support it. In an attempt to identify measurable qualitative parameters of HDL associated with increased risk for premature myocardial infarction (MI), we studied the structural characteristics of HDL from patients who survived an MI at a young age (≤35 years). METHODS AND RESULTS: We studied 20 MI patients and 20 healthy control subjects. HDL of patients had reduced apolipoprotein A-I (apoA-I), apolipoprotein M, and paraoxonase 1 levels and significantly elevated apolipoprotein C-III (apoCIII) levels (all p<0.05). Specifically, the HDL apoA-I/apoC-III ratio was 0.24±0.01 in patients versus 4.88±0.90 in controls (p<0.001). These structural alterations correlated with increased oxidation potential of HDL of the MI group compared to controls (2.5-fold, p=0.026). Electron microscopy showed no significant difference in average HDL particle diameter between the two groups though a significant difference existed in HDL diameter distribution, suggesting the presence of different HDL subpopulations in MI and control subjects. Indeed, non-denaturing two-dimensional electrophoresis revealed that MI patients had reduced pre-ß1(α), pre-ß1(b) and α(2), and elevated α(1), α(3), and pre-α(4) HDL. CONCLUSIONS: Reduction in the HDL apoA-I/apoC-III ratio, changes in the HDL subpopulation distribution and an increase in HDL oxidation potential correlated with the development of MI in young patients. The possibility that such changes may serve as markers for the early identification of young individuals at high risk for an acute coronary event should be further explored.


Subject(s)
Lipoproteins, HDL/blood , Myocardial Infarction/blood , Adult , Age of Onset , Analysis of Variance , Apolipoprotein A-I/blood , Apolipoprotein C-III/blood , Aryldialkylphosphatase/blood , Biomarkers/blood , Blotting, Western , Case-Control Studies , Chi-Square Distribution , Early Diagnosis , Electrophoresis, Gel, Two-Dimensional , Enzyme-Linked Immunosorbent Assay , Female , Greece/epidemiology , Humans , Lipoproteins, HDL/chemistry , Lipoproteins, HDL/classification , Lipoproteins, HDL/ultrastructure , Male , Microscopy, Electron , Myocardial Infarction/epidemiology , Oxidation-Reduction , Particle Size , Predictive Value of Tests , Risk Assessment , Risk Factors , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
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