RESUMEN
Unrepaired DNA damage during embryonic development can be potentially inherited by a large population of cells. However, the quality control mechanisms that minimize the contribution of damaged cells to developing embryos remain poorly understood. Here, we uncovered an ATR- and CHK1-mediated transcriptional response to replication stress (RS) in mouse embryonic stem cells (ESCs) that induces genes expressed in totipotent two-cell (2C) stage embryos and 2C-like cells. This response is mediated by Dux, a multicopy retrogene defining the cleavage-specific transcriptional program in placental mammals. In response to RS, DUX triggers the transcription of 2C-like markers such as murine endogenous retrovirus-like elements (MERVL) and Zscan4. This response can also be elicited by ETAA1-mediated ATR activation in the absence of RS. ATR-mediated activation of DUX requires GRSF1-dependent post-transcriptional regulation of Dux mRNA. Strikingly, activation of ATR expands ESCs fate potential by extending their contribution to both embryonic and extra-embryonic tissues. These findings define a novel ATR dependent pathway involved in maintaining genome stability in developing embryos by controlling ESCs fate in response to RS.
Asunto(s)
Quinasa 1 Reguladora del Ciclo Celular (Checkpoint 1)/metabolismo , Animales , Proteínas de la Ataxia Telangiectasia Mutada/genética , Proteínas de la Ataxia Telangiectasia Mutada/metabolismo , Diferenciación Celular , Proliferación Celular/fisiología , Células Cultivadas , Quinasa 1 Reguladora del Ciclo Celular (Checkpoint 1)/genética , Quimera , Cromatografía Liquida , Clonación Molecular , Daño del ADN , Células Madre Embrionarias , Regulación de la Expresión Génica , Ratones , ARN Mensajero/genética , ARN Mensajero/metabolismo , Análisis de Secuencia de ARN , Espectrometría de Masas en TándemRESUMEN
INTRODUCTION: Recent report on existence of a stem region of hemagglutinin has arisen new hopes for vaccination of influenza A as it consist of a conserve fusion peptide shared across several influenza subtypes and can be targeted by human immune system. METHODS: Given that traditional vaccination based on live attenuated viruses often fails to surpass such viral infection, a great deal of attention has been devoted to develop a safe yet efficient system for vaccination influenza A. We believe that a natural bioshuttle can be recruited for spontaneous mass vaccination. RESULTS: Thus, here, we hypothesize that a bioengineered transgenic Hirudo medicinalis can be considered as an alive bioshuttle for in-situ vaccination against influenza A virus. By introducing the designated gene(s) encoding the target fragment (i.e., stem region of hemagglutinin), this microsurgeon can act as a rapid microproducer of viral proteins for in-house mass vaccination through imparting the necessary proteins such as those, naturally presented in leech's saliva. CONCLUSION: This peculiar bioshuttle can be easily exploited as a medical modality choice at home resulting in greater patient compliance.
RESUMEN
INTRODUCTION: Today, with the tremendous potential of genomics and other recent advances in science, the role of science to improve reliable DNA extraction methods is more relevant than ever before. The ideal process for genomic DNA extraction demands high quantities of pure, integral and intact genomic DNA (gDNA) from the sample with minimal co-extraction of inhibitors of downstream processes. Here, we report the development of a very rapid, less-hazardous, and high throughput protocol for extracting of high quality DNA from blood samples. METHODS: Dried, clotted and ethylene diamine tetra-acetic acid (EDTA) treated fresh and frozen blood samples were extracted using this method in which the quality and integrity of the extracted DNA were corroborated by agarose gel electrophoresis, PCR reaction and DNA digestion using restricted enzyme. The UV spectrophotometric and gel electrophoresis analysis resulted in high A260/A280 ratio (>1.8) with high intactness of DNA. RESULTS: PCR and DNA digestion experiments indicated that the final solutions of extracted DNA contained no inhibitory substances, which confirms that the isolated DNA is of good quality. CONCLUSION: The high quality and quantity of current method, no enzymatic processing and accordingly its low cost, make it appropriate for DNA extraction not only from human but also from animal blood samples in any molecular biology labs.