H4K20me1 and H3K27me3 are concurrently loaded onto the inactive X chromosome but dispensable for inducing gene silencing.

During X chromosome inactivation (XCI), in female placental mammals, gene silencing is initiated by the Xist long non-coding RNA. Xist accumulation at the X leads to enrichment of specific chromatin marks, including PRC2-dependent H3K27me3 and SETD8-dependent H4K20me1. However, the dynamics of this process in relation to Xist RNA accumulation remains unknown as is the involvement of H4K20me1 in initiating gene silencing.

The role of Xist-mediated Polycomb recruitment in the initiation of X-chromosome inactivation.

Xist RNA has been established as the master regulator of X-chromosome inactivation (XCI) in female eutherian mammals, but its mechanism of action remains unclear. By creating novel Xist-inducible mutants at the endogenous locus in male mouse embryonic stem (ES) cells, we dissect the role of the conserved A-B-C-F repeats in the initiation of XCI. We find that transcriptional silencing can be largely uncoupled from Polycomb repressive complex 1 and complex 2 (PRC1/2) recruitment, which requires B and C repeats.

The Implication of Early Chromatin Changes in X Chromosome Inactivation.

During development, the precise relationships between transcription and chromatin modifications often remain unclear. We use the X chromosome inactivation (XCI) paradigm to explore the implication of chromatin changes in gene silencing. Using female mouse embryonic stem cells, we initiate XCI by inducing Xist and then monitor the temporal changes in transcription and chromatin by allele-specific profiling.

Consequences of VHL Loss on Global DNA Methylome.

In clear-cell renal cell carcinoma (ccRCC), loss of von Hippel-Lindau (VHL) tumour suppressor gene and reduced oxygen tension promote stabilisation of hypoxia-inducible factor (HIF) family of transcription factors, which promote changes in the expression of genes that contribute to oncogenesis. Multiple studies have demonstrated significant perturbations in DNA methylation in ccRCC via largely unclear mechanisms that modify the transcriptional output of tumour cells. Here, we show that the methylation status of the CpG dinucleotide within the consensus hypoxia-responsive element (HRE) markedly influences the binding of HIF and that the loss of VHL results in significant alterations in the DNA methylome.

Ultrasensitive detection and identification of BRAF V600 mutations in fresh frozen, FFPE, and plasma samples of melanoma patients by E-ice-COLD-PCR.

A number of molecular diagnostic methods have been developed for the detection and identification of mutations in tumor samples, which are important for the choice of treatment in the context of personalized medicine. For the treatment of metastatic melanoma, Vemurafenib is recommended for patients with BRAF V600 activating mutations. However, the different assays developed to date for the detection of these mutations lack sensitivity or specificity or do not allow a sequencing-based identification or validation of the mutation.