A further case of a Prader-Willi syndrome phenotype in a patient with Angelman syndrome molecular defect

Angelman syndrome (AS) and Prader-Willi syndrome (PWS) are distinct human neurogenetic disorders; however, a clinical overlap between AS and PWS has been identified. We report on a further case of a patient showing the PWS phenotype with the AS molecular defect. Despite the PWS phenotype, the DNA methylation analysis of SNRPN revealed an AS pattern. Cytogenetic and FISH analysis showed normal chromosomes 15 and microsatellite analysis showed heterozygous loci inside and outside the 15q11-13 region. The presence of these atypical cases could be more frequent than previously expected and we reinforce that the DNA methylation analysis is important for the correct diagnosis of severe mental deficiency, congenital hypotonia and obesity

Role of NFKB2 on the early myeloid differentiation of CD34+ hematopoietic stem/progenitor cells

To better understand the early events regulating lineage-specific hematopoietic differentiation, we analyzed the transcriptional profiles of CD34+ human hematopoietic stem and progenitor cells (HSPCs) subjected to differentiation stimulus. CD34+ cells were cultured for 12 and 40 h in liquid cultures with supplemented media favoring myeloid or erythroid commitment. Serial analysis of gene expression (SAGE) was employed to generate four independent libraries. By analyzing the differentially expressed regulated transcripts between the un-stimulated and the stimulated CD34+ cells, we observed a set of genes that was initially up-regulated at 12 h but were then down-regulated at 40 h, exclusively after myeloid stimulus. Among those we found transcripts for NFKB2, RELB, IL1B, LTB, LTBR, TNFRSF4, TGFB1, and IKBKA. Also, the inhibitor NFKBIA (IKBA) was more expressed at 12 h. All those transcripts code for signaling proteins of the nuclear factor kappaB pathway. NFKB2 is a subunit of the NF-kappa B transcription factor that with RELB mediates the non-canonical NF-kappa B pathway. Interference RNA (RNAi) against NFKB1, NFKB2 and control RNAi were transfected into bone marrow CD34+HSPC. The percentage and the size of the myeloid colonies derived from the CD34+ cells decreased after inhibition of NFKB2. Altogether, our results indicate that NFKB2 gene has a role in the early commitment of CD34+HSPC towards the myeloid lineage. (C) 2010 International Society of Differentiation. Published by Elsevier Ltd. All rights reserved.Fundacao de Amparo a Pesquisa do Estadode Sao Paulo (FAPESP)[2002/08273-1]Fundacao de Amparo a Pesquisa do Estadode Sao Paulo (FAPESP)[1998/14247-6]CNPq, Brazi

Clinical and molecular characterization of Brazilian families with von Hippel-Lindau disease: a need for delineating genotype-phenotype correlation

von Hippel-Lindau (VHL) disease is an autosomal dominant hereditary cancer syndrome that predisposes to the development of a variety of benign and malignant tumours, especially cerebellar haemangioblastomas, retinal angiomas and clear-cell renal cell carcinomas (RCC). The etiology and manifestations are due to germline and somatic mutations in the VHL tumour suppressor gene. VHL disease is classified into type 1 and type 2, showing a clear genotype-phenotype correlation, as type 2 is associated with phaeochromocytoma and essentially caused by missense mutations. The aim of this study is to characterize the phenotype and genotype of families with VHL disease. Eighteen of twenty patients from ten unrelated families underwent genetic testing, nine of them fulfilled VHL disease criteria and one had an apparently sporadic cerebellar haemangioblastoma. Four different germline mutations in the VHL gene were identified: c.226_228delTTC (p.Phe76del); c.217C > T (p.Gln73X); IVS1-1 G > A and IVS2-1 G > C. The first three mutations were associated with type 1 disease and the last one with type 2B, which had never been identified in the germline. The transcriptional processing of a novel splice-site mutation was characterised. Three type 1 VHL families showed large deletions of the VHL gene, two of them encompassed the FANCD2/C3orf10 genes and were not associated with renal lesions. We also suggest that such families should be subclassified according to the risk of RCC and the extent of the VHL gene deletions. This study highlights the need for a through clinical and molecular characterisation of families with VHL disease to better delineate its genotype-phenotype correlation.Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP)Conselho Nacional de Desenvolvimento Cientifico e Tecnologico (CNPq)Fundacao Hemocentro de Ribeirao Preto (FUNDHERP

Contribution of SLC26A4 to the molecular diagnosis of nonsyndromic prelingual sensorineural hearing loss in a Brazilian cohort

Abstract Objective Hereditary hearing loss (HL) is the most common sensorineural disorder in humans. Besides mutations in GJB2 and GJB6 genes, pathogenic variants in the SLC26A4 gene have been reported as a cause of hereditary HL due to its role in the physiology of the inner ear. In this research we wanted to investigate the prevalence of mutations in SLC26A4 in Brazilian patients with nonsyndromic prelingual sensorineural HL. We applied the high-resolution melting technique to screen 88 DNA samples from unrelated deaf individuals that were previously screened for GJB2, GJB6 and MT-RNR1 mutations. Results The frequency of mutations in the SLC26A4 gene was 28.4%. Two novel mutations were found: p.Ile254Val and p.Asn382Lys. The mutation c.-66C>G (rs17154282) in the promoter region of SLC26A4, was the most frequent mutation found and was significantly associated with nonsyndromic prelingual sensorineural HL. After mutations in the GJB2, GJB6 and mitochondrial genes, SLC26A4 mutations are considered the next most common cause of hereditary HL in Brazilian as well as in other populations, which corroborates with our data. Furthermore, we suggest the inclusion of the SCL26A4 gene in the investigation of hereditary HL since there was an increase in the frequency of the mutations found, up to 22.7%

Mitochondrial genome instability in colorectal adenoma and adenocarcinoma

Zanette, Dalila Lucíola “Documento produzido em parceria ou por autor vinculado à Fiocruz, mas não consta à informação no documento”.Submitted by Ana Maria Fiscina Sampaio ([email protected]) on 2017-09-11T19:26:35Z No. of bitstreams: 1 Araujo LF Mitochondrial genome....pdf: 1180653 bytes, checksum: 46ae00e6b7e596048911d407b473eb6a (MD5)Approved for entry into archive by Ana Maria Fiscina Sampaio ([email protected]) on 2017-09-11T19:41:06Z (GMT) No. of bitstreams: 1 Araujo LF Mitochondrial genome....pdf: 1180653 bytes, checksum: 46ae00e6b7e596048911d407b473eb6a (MD5)Made available in DSpace on 2017-09-11T19:41:06Z (GMT). No. of bitstreams: 1 Araujo LF Mitochondrial genome....pdf: 1180653 bytes, checksum: 46ae00e6b7e596048911d407b473eb6a (MD5) Previous issue date: 2015The National Council for Scientific and Technological Development (CNPq), grant #573754/2008-0; by grants #2008/57877-3 and #2013/08135-2, São Paulo Research Foundation (FAPESP); and by Research Support of the University Sao Paulo, CISBi-NAP/USP #12.1.25441.01.2.University of São Paulo. Ribeirão Preto Medical School. Department of Genetics. Ribeirão Preto, SP, Brazil / Center for Cell-Based. Therapy Regional Blood Center of Ribeirão Preto. Ribeirão Preto, SP, Brazil / National Institute of Science and Technology in Stem cell and Cell Therapy. Ribeirão Preto, SP, BrazilUniversity of São Paulo. Ribeirão Preto Medical School. Department of Genetics. Ribeirão Preto, SP, Brazil / Center for Cell-Based. Therapy Regional Blood Center of Ribeirão Preto. Ribeirão Preto, SP, Brazil / National Institute of Science and Technology in Stem cell and Cell Therapy. Ribeirão Preto, SP, BrazilUniversity of São Paulo. Ribeirão Preto Medical School. Department of Genetics. Ribeirão Preto, SP, Brazil / Center for Cell-Based. Therapy Regional Blood Center of Ribeirão Preto. Ribeirão Preto, SP, Brazil / National Institute of Science and Technology in Stem cell and Cell Therapy. Ribeirão Preto, SP, BrazilCenter for Cell-Based. Therapy Regional Blood Center of Ribeirão Preto. Ribeirão Preto, SP, Brazil / National Institute of Science and Technology in Stem cell and Cell Therapy. Ribeirão Preto, SP, BrazilUniversity of São Paulo. Ribeirão Preto Medical School. Department of Genetics. Ribeirão Preto, SP, Brazil / Center for Cell-Based. Therapy Regional Blood Center of Ribeirão Preto. Ribeirão Preto, SP, Brazil / National Institute of Science and Technology in Stem cell and Cell Therapy. Ribeirão Preto, SP, BrazilUniversity of São Paulo. Ribeirão Preto Medical School. Department of Genetics. Ribeirão Preto, SP, Brazil / Center for Cell-Based. Therapy Regional Blood Center of Ribeirão Preto. Ribeirão Preto, SP, Brazil / National Institute of Science and Technology in Stem cell and Cell Therapy. Ribeirão Preto, SP, BrazilCenter for Cell-Based. Therapy Regional Blood Center of Ribeirão Preto. Ribeirão Preto, SP, Brazil / National Institute of Science and Technology in Stem cell and Cell Therapy. Ribeirão Preto, SP, BrazilUniversity of São Paulo. Ribeirão Preto Medical School. Department of Genetics. Ribeirão Preto, SP, Brazil / Center for Cell-Based. Therapy Regional Blood Center of Ribeirão Preto. Ribeirão Preto, SP, Brazil / National Institute of Science and Technology in Stem cell and Cell Therapy. Ribeirão Preto, SP, Brazil / Center for Medical Genomics. Center for Integrative Systems Biology. Ribeirão Preto, SP, BrazilUniversity of São Paulo. Ribeirão Preto Medical School. Department of Genetics. Ribeirão Preto, SP, Brazil / Center for Cell-Based. Therapy Regional Blood Center of Ribeirão Preto. Ribeirão Preto, SP, Brazil / National Institute of Science and Technology in Stem cell and Cell Therapy. Ribeirão Preto, SP, Brazil / Center for Medical Genomics. Center for Integrative Systems Biology. Ribeirão Preto, SP, BrazilCenter for Cell-Based. Therapy Regional Blood Center of Ribeirão Preto. Ribeirão Preto, SP, Brazil / National Institute of Science and Technology in Stem cell and Cell Therapy. Ribeirão Preto, SP, Brazil / Center for Medical Genomics. Center for Integrative Systems Biology. Ribeirão Preto, SP, BrazilCenter for Cell-Based. Therapy Regional Blood Center of Ribeirão Preto. Ribeirão Preto, SP, Brazil / National Institute of Science and Technology in Stem cell and Cell Therapy. Ribeirão Preto, SP, Brazil / Center for Medical Genomics. Center for Integrative Systems Biology. Ribeirão Preto, SP, Brazil / University of São Paulo State. Faculty of Pharmaceutical Science of Araraquara. Department of Clinical Analysis. Araraquara, SP, BrazilUniversity of São Paulo. Ribeirão Preto Medical School. Department of Genetics. Ribeirão Preto, SP, Brazil / Center for Cell-Based. Therapy Regional Blood Center of Ribeirão Preto. Ribeirão Preto, SP, Brazil / National Institute of Science and Technology in Stem cell and Cell Therapy. Ribeirão Preto, SP, Brazil / Center for Medical Genomics. Center for Integrative Systems Biology. Ribeirão Preto, SP, BrazilMitochondrial dysfunction is regarded as a hallmark of cancer progression. In the current study, we evaluated mitochondrial genome instability and copy number in colorectal cancer using Next Generation Sequencing approach and qPCR, respectively. The results revealed higher levels of heteroplasmy and depletion of the relative mtDNA copy number in colorectal adenocarcinoma. Adenocarcinoma samples also presented an increased number of mutations in nuclear genes encoding proteins which functions are related with mitochondria fusion, fission and localization. Moreover, we found a set of mitochondrial and nuclear genes, which cooperate in the same mitochondrial function simultaneously mutated in adenocarcinoma. In summary, these results support an important role for mitochondrial function and genomic instability in colorectal tumorigenesis

Placenta-Enriched LincRNAs <i>MIR503HG</i> and <i>LINC00629</i> Decrease Migration and Invasion Potential of JEG-3 Cell Line

<div><p><i>LINC00629</i> and <i>MIR503HG</i> are long intergenic non-coding RNAs (lincRNAs) mapped on chromosome X (Xq26), a region enriched for genes associated with human reproduction. Genes highly expressed in normal reproductive tissues and cancers (CT genes) are well known as potential tumor biomarkers. This study aimed to characterize the structure, expression, function and regulation mechanism of <i>MIR503HG</i> and <i>LINC00629</i> lincRNAs. According to our data, <i>MIR503HG</i> expression was almost exclusive to placenta and <i>LINC00629</i> was highly expressed in placenta and other reproductive tissues. Further analysis, using a cancer cell lines panel, showed that <i>MIR503HG and LINC00629</i> were expressed in 50% and 100% of the cancer cell lines, respectively. <i>MIR503HG</i> was expressed predominantly in the nucleus of JEG-3 choriocarcinoma cells. We observed a positively correlated expression between <i>MIR503HG</i> and <i>LINC00629</i>, and between the lincRNAs and neighboring miRNAs. Also, both <i>LINC00629</i> and <i>MIR503GH</i> could be negatively regulated by DNA methylation in an indirect way. Additionally, we identified new transcripts for <i>MIR503HG</i> and <i>LINC00629</i> that are relatively conserved when compared to other primates. Furthermore, we found that overexpression of <i>MIR503HG2</i> and the three-exon <i>LINC00629</i> new isoforms decreased invasion and migration potential of JEG-3 tumor cell line. In conclusion, our results suggest that lincRNAs <i>MIR503HG</i> and <i>LINC00629</i> impaired migration and invasion capacities in a choriocarcinoma <i>in vitro</i> model, indicating a potential role in human reproduction and tumorigenesis. Moreover, the <i>MIR503HG</i> expression pattern found here could indicate a putative new tumor biomarker.</p></div

1031-1034delTAAC (Leu125Stop): a novel familial <it>UBE3A</it> mutation causing Angelman syndrome in two siblings showing distinct phenotypes

<p>Abstract</p> <p>Background</p> <p>More than 50 mutations in the <it>UBE3A</it> gene (E6-AP ubiquitin protein ligase gene) have been found in Angelman syndrome patients with no deletion, no uniparental disomy, and no imprinting defect.</p> <p>Case Presentation</p> <p>We here describe a novel <it>UBE3A</it> frameshift mutation in two siblings who have inherited it from their asymptomatic mother. Despite carrying the same <it>UBE3A</it> mutation, the proband shows a more severe phenotype whereas his sister shows a milder phenotype presenting the typical AS features.</p> <p>Conclusions</p> <p>We hypothesized that the mutation Leu125Stop causes both severe and milder phenotypes. Potential mechanisms include: i) maybe the proband has an additional problem (genetic or environmental) besides the <it>UBE3A</it> mutation; ii) since the two siblings have different fathers, the <it>UBE3A</it> mutation is interacting with a different genetic variant in the proband that, by itself, does not cause problems but in combination with the <it>UBE3A</it> mutation causes the severe phenotype; iii) this <it>UBE3A</it> mutation alone can cause either typical AS or the severe clinical picture seen in the proband.</p

Evolutionary conservation of the new isoforms.

<p>A. Sequence obtained from <i>MIR503HG2</i> isoform (<i>H</i>. <i>sapiens</i>) aligned to human genome using BLAT tool (<a href="http://genome.ucsc.edu/" target="_blank">http://genome.ucsc.edu</a>. <i>version</i>: <i>Feb</i>. <i>2009</i>, <i>CRCh37/hg19)</i> with similar sequences in <i>Saimiri boliviensis</i> (S. <i>boliviensis</i>), <i>Callithrix jacchus</i> (<i>C</i>. <i>jacchus</i>), <i>Nomascus leucogenys</i> (<i>N</i>. <i>leucogenys</i>) and <i>Papio anubis</i> (<i>P</i>. <i>anubis</i>) found in NCBI-BLAST. B. Sequences obtained from <i>LINC0026</i> isoforms comprising two exons (<i>H</i>. <i>sapiens</i>. 2) and three exons (<i>H</i>. <i>sapiens</i>. 3) aligned to human genome using BLAT tool (<a href="http://genome.ucsc.edu/" target="_blank">http://genome.ucsc.edu</a>. <i>version</i>: <i>Feb</i>. <i>2009</i>, <i>CRCh37/hg19)</i> with similar sequences in <i>Gorilla gorilla</i> (<i>G</i>. <i>gorilla</i>), <i>Pan troglodytes</i> (<i>P</i>. <i>troglodytes</i>) and <i>Pan paniscus</i> (<i>P</i>. <i>paniscus</i>) found in NCBI-BLAST.</p

<i>MIR503HG2</i> or <i>LINC00269</i> were overexpressed in JEG-3 cell line.

<p>Plasmidial expression vectors containing <i>MIR503HG2</i> or <i>LINC00629</i> full-length sequences were transfected into JEG-3 cell line. Relative expression was analyzed by RT-qPCR normalized by the geometric mean from <i>GAPDH</i> and <i>HPRT</i> endogenous genes. The empty plasmid was used as a control. *p <0.05 and **p <0.01 (<i>t</i>-test).</p

5-Aza-2-deoxycytidine (5-Aza-dC) treatment does not affect methylation status of the CpG islands near to putative promoter region of both genes.

<p>Representative normalized melt curves from samples treated with demethylating agent 5-Aza-dC. A-B. DNA analyzed from the CpG island at the promoter region of <i>MIR503HG</i> gene in DLD1 (a) and HCC1954 (b) cell lines. C. DNA analyzed from the CpG island at the promoter region of <i>LINC00629</i> gene in DLD1 cell line. Arrows indicate curves that correspond the percentage of methylation from reference, treatment, and control DNA samples. Images were obtained from High-ReSolution Melt Software v2.</p