Browsing by Subject "Mitosis"
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Item Open Access Early postzygotic mutations contribute to de novo variation in a healthy monozygotic twin pair(B M J Group, 2014) Dal, G. M.; Ergüner, B.; Saǧıroǧlu, M. S.; Yüksel, B.; Onat, O. E.; Alkan C.; Özçelik, T.Background: Human de novo single-nucleotide variation (SNV) rate is estimated to range between 0.82-1.70×10-8 mutations per base per generation. However, contribution of early postzygotic mutations to the overall human de novo SNV rate is unknown. Methods: We performed deep whole-genome sequencing (more than 30-fold coverage per individual) of the whole-blood-derived DNA samples of a healthy monozygotic twin pair and their parents. We examined the genotypes of each individual simultaneously for each of the SNVs and discovered de novo SNVs regarding the timing of mutagenesis. Putative de novo SNVs were validated using Sanger-based capillary sequencing. Results: We conservatively characterised 23 de novo SNVs shared by the twin pair, 8 de novo SNVs specific to twin I and 1 de novo SNV specific to twin II. Based on the number of de novo SNVs validated by Sanger sequencing and the number of callable bases of each twin, we calculated the overall de novo SNV rate of 1.31×10-8 and 1.01×10-8 for twin I and twin II, respectively. Of these, rates of the early postzygotic de novo SNVs were estimated to be 0.34×10-8 for twin I and 0.04×10-8 for twin II. Conclusions: Early postzygotic mutations constitute a substantial proportion of de novo mutations in humans. Therefore, genome mosaicism resulting from early mitotic events during embryogenesis is common and could substantially contribute to the development of diseases.Item Open Access Histone H3.3 regulates mitotic progression in mouse embryonic fibroblasts(Canadian Science Publishing, 2017) Ors, A.; Papin, C.; Favier, B.; Roulland, Y.; Dalkara, D.; Ozturk, M.; Hamiche, A.; Dimitrov, S.; Padmanabhan, K.H3.3 is a histone variant that marks transcription start sites as well as telomeres and heterochromatic sites on the genome. The presence of H3.3 is thought to positively correlate with the transcriptional status of its target genes. Using a conditional genetic strategy against H3.3B, combined with short hairpin RNAs against H3.3A, we essentially depleted all H3.3 gene expression in mouse embryonic fibroblasts. Following nearly complete loss of H3.3 in the cells, our transcriptomic analyses show very little impact on global gene expression or on the localization of histone variant H2A.Z. Instead, fibroblasts displayed slower cell growth and an increase in cell death, coincident with large-scale chromosome misalignment in mitosis and large polylobed or micronuclei in interphase cells. Thus, we conclude that H3.3 may have an important under-explored additional role in chromosome segregation, nuclear structure, and the maintenance of genome integrity. © 2017 Published by NRC Research Press.Item Open Access p53 mutation with frequent novel codons but not a mutator phenotype in BRCA1-and BRCA2-associated breast tumours(Nature Publishing Group, 1998) Crook, T.; Brooks, L. A.; Crossland, S.; Osin, P.; Barker, K. T.; Waller, J.; Philp, E.; Smith, P. D.; Yulug, I.; Peto, J.; Parker, G.; Allday, M. J.; Crompton, M. R.; Gusterson, B. A.The status of p53 was investigated in breast tumours arising in germ-line carriers of mutant alleles of BRCA1 and BRCA2 and in a control series of sporadic breast tumours. p53 expression was detected in 20/26 (77%) BRCA1-, 10/22 (45%) BRCA2-associated and 25/72 (35%) grade-matched sporadic tumours. Analysis of p53 sequence revealed that the gene was mutant in 33/50 (66%) BRCA-associated tumours, whereas 7/20 (35%) sporadic grade-matched tumours contained p53 mutation (P < 0.05). A number of the mutations detected in the BRCA-associated tumours have not been previously described in human cancer databases, whilst others occur extremely rarely. Analysis of additional genes, p16(INK4), Ki-ras and β-globin revealed absence or very low incidence of mutations, suggesting that the higher frequency of p53 mutation in the BRCA-associated tumours does not reflect a generalized increase in susceptibility to the acquisition of somatic mutation. Furthermore, absence of frameshift mutations in the polypurine tracts present in the coding sequence of the TGF β type II receptor (TGF β IIR) and Bax implies that loss of function of BRCA1 or BRCA2 does not confer a mutator phenotype such as that found in tumours with microsatellite instability (MSI). p21(Waf1) was expressed in BRCA-associated tumours regardless of p53 status and, furthermore, some tumours expressing wild-type p53 did not express detectable p21(Waf1). These data do not support, therefore, the simple model based on studies of BRCA-/- embryos, in which mutation of p53 in BRCA-associated tumours results in loss of p21(Waf1) expression and deregulated proliferation. Rather, they imply that proliferation of such tumours will be subject to multiple mechanisms of growth regulation.