Repeat instability at human minisatellites arising from meiotic recombination

Jeffreys, Alec J.; Neil, David L.; Neumann, Rita

doi:10.1093/emboj/17.14.4147

Cited by 108 publications

(93 citation statements)

References 35 publications

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“…Meiotic intra-allelic rearrangements have been observed at unstable minisatellites (Buard & Vergnaud 1994, Jeffreys et al 1994. The localization of these tandem-repeat DNA sequences in CO hotspots strongly suggests that these rearrangements are a product of the repair of meiotic DSB (Jeffreys et al 1998, Buard et al 2000. Though rearranged products resulting from intra-or interchromatids cannot be distinguished from each other, some of them might result from intra-chromatid events.…”

Section: Unexplored Dsb Repair Eventsmentioning

confidence: 99%

Regulating double-stranded DNA break repair towards crossover or non-crossover during mammalian meiosis

2007

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During meiosis the programmed induction of DNA double-stranded breaks (DSB) leads to crossover (CO) and non-crossover products (NCO). One key role of CO is to connect homologs before metaphase I and thus to ensure the proper reductional segregation. This role implies an accurate regulation of CO frequency with the establishment of at least one CO per chromosome arm. Current major challenges are to understand how CO and NCO formation are regulated and what is the role of NCO. We present here the current knowledge about CO and NCO and their regulation in mammals. CO density varies widely along chromosomes and their distribution is not random as they are subject to positive interference. As documented in the mouse and human, a significant excess of DSB are generated relative to the number of CO. In fact, evidence has been obtained for the formation of NCO products, for regulation of the choice of DSB repair towards CO or NCO and for a CO specific pathway. We discuss the roles of Msh4, Msh5 and Sycp1 which affect DSB repair and probably not only the CO pathway. We suggest that, in mammals, the regulation of NCO differs from that described in Saccharomyces cerevisiae.

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Section: Unexplored Dsb Repair Eventsmentioning

confidence: 99%

Regulating double-stranded DNA break repair towards crossover or non-crossover during mammalian meiosis

2007

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“…MS32 and C1NH outside and nested PCR reactions employed a long PCR protocol. 7 Locus-specific probes (c. 300 bp long) were synthesised at MS32 by amplification of 1 ng MS32 cosmid DNA 8 using primer pairs 32-5NF and 32-5R (probe 1), and 32-1.6F and 32-1.3NR (probe 2) for 20 cycles, denaturing at 948C for 15 s, annealing at 658C for 30 s and 1 min extension at 708C; and at C1NH by amplification of 200 ng genomic DNA using primers C1+2.4NF and C1+2.8R (probe 1), and C1+6.5F and C1+6.8NR (probe 2) for 30 cycles, denaturing at 968C for 1 min, annealing at 628C for 30 s and 1 min extension at 708C. At MS32 semi-nested PCR was carried out using primers 32-5F and 32-1.3NR for 26 cycles, denaturing at 968C for 15 s, annealing at 688C for 30 s and 6 min extension at 708C.…”

Section: Pcr Amplificationmentioning

confidence: 99%

“…Following amplification, 500 ng of pAT153-DdeI/BamHI marker DNA was added to each sample as an internal control and separated by agarose gel electrophoresis. DNA between 1.4 and 9.5 kb was sizefractionated 7 to enrich for alleles containing insertions of between 200 and 7000 bp (consensus Alu and L1 elements have lengths of c. 300 and 6000 bp respectively). Fifty per cent of the purified DNA was re-amplified for between 12 ± 16 cycles such that the residual progenitor allele was amplified to levels no greater than 1 ng, and again size fractionated.…”

Section: Pcr Amplificationmentioning

confidence: 99%

“…4 To date, such insertions and re-arrangements have only been characterised in patients, resulting in considerable uncertainties about the nature of these processes. We have therefore developed highly sensitive methods for mutation detection, analogous to methods devised for studying de novo mutation at highly unstable minisatellites, 7 and have used these in an attempt to detect dispersed repeat-mediated instability in somatic and germline DNA at three different loci.…”

Section: Introductionmentioning

confidence: 99%

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Attempts to detect retrotransposition and de novo deletion of Alus and other dispersed repeats at specific loci in the human genome

2001

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Dispersed repeat elements contribute to genome instability by de novo insertion and unequal recombination between repeats. To study the dynamics of these processes, we have developed single DNA molecule approaches to detect de novo insertions at a single locus and Alu-mediated deletions at two different loci in human genomic DNA. Validation experiments showed these approaches could detect insertions and deletions at frequencies below 10 76 per cell. However, bulk analysis of germline (sperm) and somatic DNA showed no evidence for genuine mutant molecules, placing an upper limit of insertion and deletion rates of 2610 77 and 3610 77 , respectively, in the individuals tested. Such re-arrangements at these loci therefore occur at a rate lower than that detectable by the most sensitive methods currently available.

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“…Indeed if, similar to the human minisatellites, 3 mutation at these loci were mainly attributed to meiotic gene-conversion like events [17], then the frequency of ESTR mutation in meiotic spermatocytes and post-meiotic spermatids should substantially exceed that in pre-meiotic spermatogonia. The data showing the lack of measurable effects of meiotic events on the frequency of ESTR mutation in the male germline is consistent with the results of our previous study which failed to detect any correlation between meiotic crossing over and ESTR mutation induction [18].…”

Section: Resultsmentioning

confidence: 99%

Stage-specificity of spontaneous mutation at a tandem repeat DNA locus in the mouse germline

Shanks¹,

Riou²,

Fouchet³

et al. 2008

Mutation Research/Fundamental and Molecular Mechanisms of Mutag

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Mouse expanded simple tandem repeat (ESTR) loci are the most unstable loci in the mouse genome. Despite the fact that over the last decade these loci have been extensively used for studying germline mutation induction in mice, to date little is known about the mechanisms underlying spontaneous and induced ESTR mutation. Here we used flow cytometry and single-molecule PCR to compare the frequency of ESTR mutation in four flow-sorted fractions of the mouse male germ cells -spermatogonia, spermatocytes I, round and elongated spermatids. The frequency and the spectrum of ESTR mutation did not significantly differ between different stages of mouse spermatogenesis. Considering these data and the results of other publications, we propose that spontaneous ESTR mutation is mostly attributed to replication slippage in spermatogonia and these loci may be regarded as a class of expanded microsatellites. IntroductionThe results of some recent publication show that mouse ESTR loci provide a sensitive technique for studying germline mutation induction following exposure to ionising radiation and chemical mutagens [1,2]. ESTR loci consist of long homogenous arrays of relatively short repeats (4-9 bp) and show a very high spontaneous mutation rate of length changes both in germline and somatic cells [3][4][5]. In this respect they clearly differ from another group of highly unstable tandem repeat DNA loci -the GC-rich human minisatellites, where the bulk of spontaneous mutations occur in the germline with very rare events taking place in the somatic cells [6]. Given this, ESTRs more closely resemble some microsatellite loci showing equally high instability in the germline and somatic tissues. In many publications it has been suggested, though not experimentally proven, that microsatellite mutation is most probably attributed to replication slippage [7]. However, numerous studies have also identified a unique class of the expanded disease-causing human microsatellites i.e., trinucleotide instability, which is clearly attributed to non-replication mechanisms [8]. It therefore remains unclear whether the very high spontaneous instability at ESTR loci is replication-driven or may be attributed to some other mechanisms. Here, to elucidate the still unknown mechanisms of ESTR spontaneous mutation, we have compared ESTR mutation frequencies in flow-sorted fractions of the male germ cells. Materials and methods Flow cytometryCBA/J male mice (Iffa Credo, Charles River, France) were used in this study. Cells were isolated from 3 month-old male mice using a two-step enzymatic digestion as previously described [9]. Magnetic-activated cell sorting (MACS) of α 6 -integrin positive cells was performed as we previously described [9] using rat anti-α 6 -integrin antibodies (GoH3, BD Biosciences) and mouse anti-rat Kappa antibody coupled to magnetic microbeads (Miltenyi Biotec, Paris, France). Testicular cell suspension, MACS α 6 -integrin-positive and negative cellular fractions were collected and suspended at 10 6 cells/ml for DNA stain...

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Repeat instability at human minisatellites arising from meiotic recombination

Cited by 108 publications

References 35 publications

Regulating double-stranded DNA break repair towards crossover or non-crossover during mammalian meiosis

Regulating double-stranded DNA break repair towards crossover or non-crossover during mammalian meiosis

Attempts to detect retrotransposition and de novo deletion of Alus and other dispersed repeats at specific loci in the human genome

Stage-specificity of spontaneous mutation at a tandem repeat DNA locus in the mouse germline

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