Angelman Syndrome Protein UBE3A Interacts with Primary Microcephaly Protein ASPM, Localizes to Centrosomes and Regulates Chromosome Segregation

Many proteins associated with the phenotype microcephaly have been localized to the centrosome or linked to it functionally. All the seven autosomal recessive primary microcephaly (MCPH) proteins localize at the centrosome. Microcephalic osteodysplastic primordial dwarfism type II protein PCNT and Seckel syndrome (also characterized by severe microcephaly) protein ATR are also centrosomal proteins. All of the above findings show the importance of centrosomal proteins as the key players in neurogenesis and brain development. However, the exact mechanism as to how the loss-of-function of these proteins leads to microcephaly remains to be elucidated. To gain insight into the function of the most commonly mutated MCPH gene ASPM, we used the yeast two-hybrid technique to screen a human fetal brain cDNA library with an ASPM bait. The analysis identified Angelman syndrome gene product UBE3A as an ASPM interactor. Like ASPM, UBE3A also localizes to the centrosome. The identification of UBE3A as an ASPM interactor is not surprising as more than 80% of Angelman syndrome patients have microcephaly. However, unlike in MCPH, microcephaly is postnatal in Angelman syndrome patients. Our results show that UBE3A is a cell cycle regulated protein and its level peaks in mitosis. The shRNA knockdown of UBE3A in HEK293 cells led to many mitotic abnormalities including chromosome missegregation, abnormal cytokinesis and apoptosis. Thus our study links Angelman syndrome protein UBE3A to ASPM, centrosome and mitosis for the first time. We suggest that a defective chromosome segregation mechanism is responsible for the development of microcephaly in Angelman syndrome.

Annotated Chromosome Maps for Renal Disease

A combination of linkage analyses and association studies are currently employed to promote the identification of genetic factors contributing to inherited renal disease. We have standardized and merged complex genetic data from disparate sources, creating unique chromosomal maps to enhance genetic epidemiological investigations. This database and novel renal maps effectively summarize genomic regions of suggested linkage, association, or chromosomal abnormalities implicated in renal disease. Chromosomal regions associated with potential intermediate clinical phenotypes have been integrated, adding support for particular genomic intervals. More than 500 reports from medical databases, published scientific literature, and the World Wide Web were interrogated for relevant renal-related information. Chromosomal regions highlighted for prioritized investigation of renal complications include 3q13-26, 6q22-27, 10p11-15, 16p11-13, and 18q22. Combined genetic and physical maps are effective tools to organize genetic data for complex diseases. These renal chromosome maps provide insights into renal phenotype-genotype relationships and act as a template for future genetic investigations into complex renal diseases. New data from individual researchers and/or future publications can be readily incorporated to this resource via a user-friendly web-form accessed from the website: www.qub.ac.uk/neph-res/CORGI/index.php.

Suggestive association with ocular phoria at chromosome 6p22

PURPOSE. We conducted a genome-wide association study to identify genetic factors that contribute to the etiology of heterophoria.

METHODS. We measured near and far vertical and horizontal phorias in 988 healthy adults aged 16 to 40 using the Keystone telebinocular with plates 5218 and 5219. We regressed degree of phoria against genotype at 642758 genetic loci. To control for false positives, we applied the conservative genome-wide permutation test to our data.

RESULTS. A locus at 6p22.2 was found to be associated with the degree of near horizontal phoria (P = 2.3 × 10 ). The P value resulting from a genome-wide permutation test was 0.014.

CONCLUSIONS. The strongest association signal arose from an intronic region of the gene ALDH5A1, which encodes the mitochondrial enzyme succinic semialdehyde dehydrogenase (SSADH), an enzyme involved in γ-aminobutyric acid metabolism. Succinic semialdehyde dehydrogenase deficiency, resulting from mutations of ALDH5A1, causes a variety of neural and behavioral abnormalities, including strabismus. Variation in ALDH5A1 is likely to contribute to degree of horizontal phoria.

Prematuration of bovine oocytes with butyrolactone I reversibly arrests meiosis without increasing meiotic abnormalities after in vitro maturation

Objectives: Asynchrony between nuclear and cytoplasmic maturation, and possibly damage to the oocyte meiotic spindle, limits the application of in vitro maturation (IVM) in assisted reproduction. Several studies have suggested that Prematuration with meiosis blockers may improve oocyte quality after IVM, favoring early embryogenesis. Thus, we investigated the effect of Prematuration with the nuclear maturation inhibitor butyrolactone I (BLI) on the meiotic spindle and chromosomal configuration of bovine oocytes. Study design: Immature oocytes obtained from cows slaughtered in a slaughterhouse (n = 840) were divided into the following groups: (1) control (n = 325), submitted only to IVM in TCM199 for 24 h; (2) BLI 18 h (n = 208) submitted to meiotic blockage with 100 mu M BLI for 24 h (Prematuration) and then induction of IVM in TCM199 for 18 h; and (3) BLI 24 h (n = 307), pre-matured with 100 mu m BLI for 24 h followed by 24 h of IVM in TCM199. The oocytes were then fixed, stained by immunofluorescence for morphological visualization of both microtubules and chromatin, and evaluated. Results: Meiotic arrest occurred in 90.2% of the oocytes cultured with BLI. Maturation rates were similar for all groups (80.3%, 73.6% and 82.7% for the control, BLI 18 h and BLI 24 h groups, respectively). We observed 81.3% normal oocytes in metaphase II in the control group, and 80.0% and 81.2% in the BLI 18 h and BLI 24 h groups, respectively. The incidence of meiotic anomalies did not differ between groups (18.7%, 20.0% and 18.8% for the control, BLI 18 h and BLI 24 h, respectively). Conclusion: Prematuration with butyrolactone I reversibly arrests meiosis without damaging the meiotic spindle or the chromosome distribution of bovine oocytes after in vitro maturation. (c) 2009 Elsevier Ireland Ltd. All rights reserved.

A Rare Case of Trisomy 15pter-q21.2 Due to a De Novo Marker Chromosome

Supernumerary marker chromosomes (sSMC) may or may not be associated with an abnormal phenotype, depending on the presence of euchromatin, on their chromosomal origin and whether they are inherited. Over 80% of sSMCs are derived from acrocentric chromosomes and half of them include the short arm of chromosome 15. Generally, they appear as bisatellited isodicentric marker chromosomes, most of them are symmetric. These chromosomes are normally originated de novo and are associated with mild to severe intellectual disability but not with physical abnormalities. We report on a patient with an SMC studied using classical and molecular cytogenetic procedures (G and C banding, NOR staining, painting and centromeric fluorescent in situ hybridization (FISH), BAC-FISH, and SKY). The MLPA technique and DNA polymorphic markers were used in order to identify its parental origin. The marker chromosome, monosatellited and monocentric, was found to be derived from a maternal chromosome 15 and was defined as 15pter-q21.2. This is the report of the largest de novo monosatellited 15q marker chromosome ever published presenting detailed cytogenetic and clinical data. It was associated with a phenotype including cardiac defect, absence of septum pellucidum, and dysplasia of the corpus callosum. (C) 2010 Wiley-Liss, Inc.

Loss of Y-chromosome does not correlate with age at onset of head and neck carcinoma: a case-control study

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Genotoxicity and mutagenicity of water contaminated with tannery effluents, as evaluated by the micronucleus test and comet assay using the fish Oreochromis niloticus and chromosome aberrations in onion root-tips

Cytotoxicity of metals is important because some metals are potential mutagens able to induce tumors in humans and experimental animals. Chromium can damage DNA in several ways, including DNA double strand breaks (DSBs) which generate chromosomal aberrations, micronucleus formation, sister chromatid exchange, formation of DNA adducts and alterations in DNA replication and transcription. In our study, water samples from three sites in the Córrego dos Bagres stream in the Franca municipality of the Brazilian state of São Paulo were subjected to the comet assay and micronucleus test using erythrocytes from the fish Oreochromis niloticus. Nuclear abnormalities of the erythrocytes included blebbed, notched and lobed nuclei, probably due to genotoxic chromium compounds. The greatest comet assay damage occurred with water from a chromium-containing tannery effluent discharge site, supporting the hypothesis that chromium residues can be genotoxic. The mutagenicity of the water samples was assessed using the onion root-tip cell assay, the most frequent chromosomal abnormalities observed being: c-metaphases, stick chromosome, chromosome breaks and losses, bridged anaphases, multipolar anaphases, and micronucleated and binucleated cells. Onion root-tip cell mutagenicity was highest for water samples containing the highest levels of chromium.

Allelic imbalance studies of chromosome 9 suggest major differences in chromosomal instability among nonmelanoma skin carcinomas

CONTEXTO: Vários estudos de perda de heterozigozidade na região 9p21-p22, que abriga os genes supressores tumorais CDKN2a/p16INK4a, p19ARF e p15INK4b, têm sido realizados em uma ampla série de tumores humanos, incluindo os melanomas familiares. Perdas e ganhos em outras regiões do cromossomo 9 também têm sido observados com freqüência e podem indicar mecanismos adicionais no processo de tumorigênese dos carcinomas basocelulares da pele. OBJETIVO: Investigar o equilíbrio alélico existente na região 9p21-p22 em carcinomas basocelulares. TIPO DE ESTUDO: Análise molecular de marcadores de microssatélites em tumores e controles. LOCAL: Dois serviços de dermatologia de atendimento terciário em universidades públicas de São Paulo e o Laboratório de Genética Molecular do Câncer da Universidade Estadual de Campinas (UNICAMP), Brasil. PARTICIPANTES: Examinamos 13 casos benignos, incluindo 4 queratoses solares, 3 queratoacantomas, 3 nevos melanocíticos, 2 doenças de Bowen e 1 neurofibroma cutâneo, além de 58 tumores malignos da pele: 14 de células escamosas, 40 carcinomas basocelulares e 4 melanomas; em pacientes consecutivamente encaminhados à clínica de Dermatologia da Unicamp e que concordaram em participar do estudo. VARIÁVEIS ESTUDADAS: O tumor principal e uma porção normal de pele não-adjacente foram removidos cirurgicamente de pacientes que consecutivamente procuraram os ambulatórios de dermatologia da Universidade Estadual de Campinas (UNICAMP) e da Universidade Estadual de São Paulo (Unesp), São Paulo, por causa de lesões cutâneas. Extraímos DNA tanto de tecido tumoral como do correspondente tecido normal de cada paciente. Para amplificar regiões de repetição polimórfica de microssatélites do cromossomo 9, foram utilizados quatro pares de primers, sendo dois deles destinados à região 9p21-p22. RESULTADOS: Identificamos oito casos (20%) de desequilíbrio alélico entre os carcinomas basocelulares, sendo dois casos de perda de heterozigozidade e seis casos de instabilidade de microssatélite na região 9p21-p22. Outros marcadores também mostravam anormalidades em três destes tumores, enquanto nenhuma alteração foi detectada entre os casos benignos e nos outros tumores malignos. CONCLUSÃO: Esta dependência fenotípica sugere que existem diferenças importantes no comportamento das formas mais comuns de tumores cutâneos não-melanocíticos em relação à sua tendência para instabilidade de microssatélite no cromossomo 9. Considerando-se que os genes CDKN2a/p16INK4a, p19ARF e p15INK4b não parecem responsáveis pelas anormalidades observadas, outros genes em 9p21-p22 podem estar envolvidos na etiopatogenia e na progressão dos carcinomas basocelulares.

Cytogenetics of three breeds of river buffalo (Bubalus bubalis L.), with evidence of a fragile site on the X chromosome

The cytogenetic study of 182 river buffalo (Bubalus bubalis L., 2n=50) of Murrah, Mediterranean and Jaffarabadi breeds, from the State of São Paulo, was carried out to characterize their chromosomes and to detect possible chromosomal abnormalities. The karyotypes were indistinguishable with conventional staining as well as with C and replication R banding techniques. In about 44% of the sample (8 males and 72 females), an X marker chromosome due to a fragile site was shown. The frequency of metaphases expressing the fragility site on the X was highly variable, from 2.86 to 41.03%. In females, the fragile site, rarely appeared on both X chromosomes. Most of the metaphases showed only 1 marker chromosome. In R-banded metaphases using 5-bromodeoxyuridine (BrdU) treatment, it corresponded in general to the late replicating X chromosome. No correlation between the X fragile site and altered phenotype was found. Structural and numerical chromosome rearrangements were ruled out in the present sample of buffalo. (C) 1998 by Elsevier B.V.

Interstitial long-arm deletion of chromosome 7 and ectrodactyly

An interstitial deletion of 7q21 was found in a boy with mental retardation, microcephaly, convergent strabismus, micrognathia, genital anomalies, and other findings, including ectrodactyly.

Clonal monosomy of chromosome 21 in a case of myelodysplastic syndrome

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Induction of mitotic and chromosomal abnormalities on Allium cepa cells by pesticides imidacloprid and sulfentrazone and the mixture of them

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Loss of Y-chromosome does not correlate with age at onset of head and neck carcinoma: a case-control study

Loss of Y-chromosome has been correlated with older age in males. Furthermore, current evidence indicates that Y-chromosome loss also occurs in several human tumors, including head and neck carcinomas. However, the association between Y nullisomy and the occurrence of neoplasias in elderly men has not been well established. In the present study, the association between Y-chromosome loss and head and neck carcinomas was evaluated by comparison to cells from peripheral blood lymphocytes and normal mucosa of cancer-free individuals matched for age using dual-color fluorescence in situ hybridization. Twenty-one patients ranging in age from 28 to 68 years were divided into five-year groups for comparison with 16 cancer-free individuals matched for age. The medical records of all patients were examined to obtain clinical and histopathological data. None of the patients had undergone radiotherapy or chemotherapy before surgery. In all groups, the frequency of Y-chromosome loss was higher among patients than among normal reference subjects (P < 0.0001) and was not age-dependent. These data suggest that Y-chromosome loss is a tumor-specific alteration not associated with advanced age in head and neck carcinomas.

Interphase Chromosome Flow-FISH

A 2-day method using flow cytometry and FISH for interphase cells was developed to detect monosomy 7 cells in myelodysplastic syndrome patients. The method, Interphase Chromosome Flow-FISH (IC Flow-FISH), involves fixation of leukocytes from blood, membrane permeabilization, hybridization of cellular DNA with peptide nucleic acid probes with cells intact, and analysis by flow cytometry. Hundreds to thousands of monosomy 7 cells were consistently detected from 10-20 mL of blood in patients with monosomy 7. Proportions of monosomy 7 cells detected in IC Flow-FISH were compared with results from conventional cytogenetics; identification of monosomy 7 populations was verified with FACS; and patient and donor cells were mixed to test for sensitivity. IC Flow-FISH allows for detecting monosomy 7 without requiring bone marrow procurement or the necessity of metaphase spreads, and wider applications to other chromosomal abnormalities are in development. (Blood. 2012; 120(15): e54-e59)