RESUMEN
Genetic effects from radiation have been observed in a number of species to date. However, observations in humans are nearly nonexistent. In this review, possible reasons for the paucity of positive observations in humans are discussed. Briefly, it appears likely that radiation sensitivity for the induction of mutations varies among different genes, and that the specific genes that were used in the past with the specific locus test utilizing millions of mice may have simply been very responsive to radiation. In support of this notion, recent studies targeting the whole genome to detect copy number variations (deletions and duplications) in offspring derived from irradiated spermatogonia indicated that the mutation induction rate per genome is surprisingly lower than what would have been expected from previous results with specific locus tests, even in the mouse. This finding leads us to speculate that the lack of evidence for the induction of germline mutations in humans is not due to any kind of species differences between humans and mice, but rather to the lack of highly responsive genes in humans, which could be used for effective mutation screening purposes. Examples of such responsive genes are the mouse coat color genes, but in human studies many more genes with higher response rates are required because the number of offspring examined and the radiation doses received are smaller than in mouse studies. Unfortunately, such genes have not yet been found in humans. These results suggest that radiation probably induces germline mutations in humans but that the mutation induction rate is likely to be much lower than has been estimated from past specific locus studies in mice. Whole genome sequencing studies will likely shed light on this point in the near future.
Asunto(s)
Tolerancia a Radiación/genética , Animales , Cromosomas Humanos X/genética , Cromosomas Humanos X/efectos de la radiación , Sitios Genéticos/genética , Sitios Genéticos/efectos de la radiación , Genoma Humano/genética , Haplotipos/genética , Haplotipos/efectos de la radiación , Humanos , Ratones , Mutagénesis/efectos de la radiación , Tasa de Mutación , Especificidad de la EspecieRESUMEN
Chromatin modifications are long known as an essential part of the orchestrated response resulting in the repair of radiation-induced DNA double-strand breaks (DSBs). Only recently, however, the influence of the chromatin architecture itself on the DNA damage response has been recognised. Thus for heterochromatic DSBs the sensing and early recruitment of repair factors to the lesion occurs within the heterochromatic compartments, but the damage sites are subsequently relocated from the inside to the outside of the heterochromatin. While previous studies were accomplished at the constitutive heterochromatin of centromeric regions in mouse and flies, here we examine the DSB repair at the facultative heterochromatin of the inactive X chromosome (Xi) in humans. Using heavy ion irradiation we show that at later times after irradiation the DSB damage streaks bend around the Xi verifying that the relocation process is conserved between species and not specialised to repetitive sequences only. In addition, to measure chromatin relaxation at rare positions within the genome, we established live cell microscopy at the GSI microbeam thus allowing the aimed irradiation of small nuclear structures like the Xi. Chromatin decondensation at DSBs within the Xi is clearly visible within minutes as a continuous decrease of the DNA staining over time, comparable to the DNA relaxation revealed at DSBs in mouse chromocenters. Furthermore, despite being conserved between species, slight differences in the underlying regulation of these processes in heterochromatic DSBs are apparent.
Asunto(s)
Cromatina/genética , Cromosomas Humanos X/genética , Daño del ADN/genética , Reparación del ADN/genética , Fibroblastos/patología , Heterocromatina/genética , Animales , Cromosomas Humanos X/efectos de la radiación , Daño del ADN/efectos de la radiación , Femenino , Fibroblastos/metabolismo , Técnica del Anticuerpo Fluorescente , Histonas/genética , Histonas/metabolismo , Humanos , Ratones , Células 3T3 NIHRESUMEN
The ultraviolet (UV) component of sunlight is the main cause of skin cancer. More than 50% of all non-melanoma skin cancers and >90% of squamous cell carcinomas in the US carry a sunlight-induced mutation in the p53 tumor suppressor gene. These mutations have a strong tendency to occur at methylated cytosines. Ligation-mediated PCR (LMPCR) was used to compare at nucleotide resolution DNA photoproduct formation at dipyrimidine sites either containing or lacking a methylated cytosine. For this purpose, we exploited the fact that the X chromosome is methylated in females only on the inactive X chromosome, and that the FMR1 (fragile-X mental retardation 1) gene is methylated only in fragile-X syndrome male patients. Purified genomic DNA was irradiated with UVC (254nm), UVB (290-320nm) or monochromatic UVB (302 and 313nm) to determine the effect of different wavelengths on cyclobutane pyrimidine dimer (CPD) formation along the X-linked PGK1 (phosphoglycerate kinase 1) and FMR1 genes. We show that constitutive methylation of cytosine increases the frequency of UVB-induced CPD formation by 1.7-fold, confirming that methylation per se is influencing the probability of damage formation. This was true for both UVB sources used, either broadband or monochromatic, but not for UVC. Our data prove unequivocally that following UVB exposure methylated cytosines are significantly more susceptible to CPD formation compared with unmethylated cytosines.
Asunto(s)
Citosina/metabolismo , Metilación de ADN/efectos de la radiación , Dímeros de Pirimidina/metabolismo , Dímeros de Pirimidina/efectos de la radiación , Rayos Ultravioleta/efectos adversos , Secuencia de Bases , Células Cultivadas , Cromosomas Humanos X/metabolismo , Cromosomas Humanos X/efectos de la radiación , Daño del ADN , Cartilla de ADN/genética , Femenino , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/genética , Síndrome del Cromosoma X Frágil/genética , Síndrome del Cromosoma X Frágil/metabolismo , Humanos , Masculino , Fosfoglicerato Quinasa/genética , Inactivación del Cromosoma X/genéticaRESUMEN
Densely ionizing radiations interact with DNA to cause heavily clustered sites of damage that are difficult to repair correctly. We have been able to determine for the first time the breakpoints of several very large deletions induced by densely ionizing radiation in diploid human cells and show that damage clustering is reflected in the complexity of mutations. Intra- and interchromosomal insertions and inversions occur at the sites of some large deletions. Short sequence repeats are commonly found at the breakpoints, showing that microhomologies help patch damage sites. We suggest that novel fragments found in complex rearrangements derive from other sites of radiation damage in the same cell. These transmissible molecular changes are echoed by visible chromosome rearrangements many days after irradiation and are likely to contribute significantly to the carcinogenic properties of densely ionizing radiations.