Project description:Chromosomes and plasmids are two forms of genetic carriers. Exogenous yeast artificial chromosomes are also considered as yeast centromeric plasmids in many cases. Here, we used state-of-the-art sequencing technologies to comprehensively profile the genetic, epigenetic, transcriptional and proteomic characteristics of an exogenous data-carrying chromosome. We found that the data-carrying DNA formed active chromatin with high chromatin accessibility and H3K4 tri-methylation levels. We also confirmed that the data-carrying chromosome had a circular shape in the nucleus and was arranged in the Rabl configuration, which may contribute to the self-replication and haploidy of the chromosome in vivo. The data-carrying chromosome displayed highly pervasive transcriptional ability and transcribed hundreds of non-coding RNAs. In summary, this work explores the chromatin epigenetic state, chromatin structure and transcriptional landscape of an exogenous artificial chromosome. The results demonstrated that the exogenous artificial chromosome did form a chromatin structure and was not a naked and incompact plasmid, which strengthen our understanding of artificial chromosomes.
Project description:While histone H3 lysine 27 trimethylation (H3K27Me3) is associated with gene silencing, whether H3K27Me3 demethylation affects transcription and cell differentiation in vivo has remained elusive. To investigate this, we conditionally inactivated the two H3K27Me3 demethylases, Jmjd3 and Utx, in non-dividing intrathymic CD4+ T cell precursors. We show that both enzymes redundantly promote H3K27Me3 removal at, and expression of, a specific subset of genes involved in terminal thymocyte differentiation, especially S1pr1, encoding a sphingosine-phosphate receptor required for thymocyte egress. Floxed alleles of the genes encoding Utx and Jmjd3 (Kdm6a and Kdm6b, respectively) were deleted in double positive (DP) thymocytes carrying a CD4 Cre transgene. Genome-wide H3K27Me3 ChipSeq was performed on (i) pre-selection (CD69lo) DP thymocytes from wild-type mice carrying an endogenous polyclonal TCR repertoire, (ii) mature (TCRhi CD24lo) CD4 SP thymocytes from wild type (Wt), Jmjd3KO, UtxKO and dKO mice carrying an endogenous polyclonal TCR repertoire and (iii) mature (Va2hi CD24lo) CD4 SP thymocytes from wild type and dKO mice carrying the OTII TCR transgene.
Project description:Chromosomes and plasmids are two forms of genetic carriers. Exogenous yeast artificial chromosomes are also considered as yeast centromeric plasmids in many cases. Here, we used state-of-the-art sequencing technologies to comprehensively profile the genetic, epigenetic, transcriptional and proteomic characteristics of an exogenous data-carrying chromosome. We found that the data-carrying DNA formed active chromatin with high chromatin accessibility and H3K4 tri-methylation levels. We also confirmed that the data-carrying chromosome had a circular shape in the nucleus and was arranged in the Rabl configuration, which may contribute to the self-replication and haploidy of the chromosome in vivo. The data-carrying chromosome displayed highly pervasive transcriptional ability and transcribed hundreds of non-coding RNAs. In summary, this work explores the chromatin epigenetic state, chromatin structure and transcriptional landscape of an exogenous artificial chromosome. The results demonstrated that the exogenous artificial chromosome did form a chromatin structure and was not a naked and incompact plasmid, which strengthen our understanding of artificial chromosomes.