Project description:Heterochronic grafts (between different embryonic stages) have been described as having more posterior axis contributions than homochronic (stage-matched) ones. Here, we performed homochronic (HH4 to HH4) and heterochronic (HH8 to HH4) grafts from GFP expressing transgenic donor embryos to wild-type Bovans brown embryos and carried out single cell RNA-sequencing to understand the transcriptomic cell states occupied by homochronic and heterochronic grafts early after grafting.
Project description:Non-coding RNA plays an important regulatory role in the occurrence and development of hypoxic pulmonary hypertension (HPH). Therefore, we use high-throughput RNA sequence and bioinformatics methods to analyze the whole transcriptome HPH rats in lung tissue.
Project description:The family of Heterochromatin Protein 1 (HP1) consists of highly conserved proteins, which have important functions in the nucleus of eukaryotic cells. In mammals there are three HP1 paralogs: HP1(alpha), Hp1(beta), and Hp1(gamma)They are encoded by the Cbx5, Cbx1, and Cbx3 genes, respectively. Hp1 and Hp1 stably interact with Chd4 and Adnp to form the ChAHP complex. In this project, Chd4, Adnp, and the three Cbx genes were endogenously tagged with a FLAG-Avi tag in mouse embryonic stem cells. The tagged proteins were subjected to tandem-affinity purification and analysis by mass spectrometry.
Project description:HP1 alpha, beta, and gamma play an evolutionarily conserved role in the formation and maintenance of heterochromatin. In addition, some HP1 family members may also participate in transcriptional regulation of genes. Recently, HP1 gamma binding to the bodies of a subset of genes has been observed in human and murine cells. However, the generality of this phenomenon and the role HP1 gamma may play in this context are unknown. Genome-wide localization analysis reveals broad and general HP1 gamma binding at actively transcribed genes, which strongly correlates with gene activity across multiple cell types. Unexpectedly, we find HP1 gamma binding is necessary for efficient RNA processing of its target genes. Loss of HP1 gamma results in defective recruitment of splicing factors including U1-70K, and leads to accumulation of unspliced nascent transcripts genome-wide. Therefore, our data argue that the primary role of HP1 gamma is to ensure efficient RNA processing in mammalian cells. Genome-wide localization and function of HP1 gamma in multiple cell types using ChIP-chip, ChIP-seq, and RNA-seq
Project description:17β-estradiol (E2) exerts complex and context-dependent effects in pulmonary hypertension. In hypoxia-induced pulmonary hypertension (HPH), E2 attenuates lung vascular remodeling through estrogen receptor (ER)-dependent effects; however, ER target genes in the hypoxic lung remain unknown. In order to identify the genome regulated by the E2-ER axis in the hypoxic lung, we performed a microarray analysis in lungs from HPH rats treated with E2 (75 mcg/kg/d) ± ER-antagonist ICI182,780 (3 mg/kg/d). Untreated HPH rats and normoxic rats served as controls. Using a false discovery rate of 10%, we identified a significantly differentially regulated genome in E2-treated vs. untreated hypoxia rats. Genes most up-regulated by E2 encoded matrix metalloproteinase 8, S100 calcium binding protein A8, and IgA Fc receptor; genes most down-regulated by E2 encoded olfactory receptor 63, secreted frizzled-related protein 2, and thrombospondin 2. Several genes affected by E2 changed in the opposite direction after ICI182,780 co-treatment, indicating an ER-regulated genome in HPH lungs. The bone morphogenetic protein antagonist Grem1 (gremlin 1) was up-regulated by hypoxia, but found to be among the most down-regulated genes after E2 treatment. Gremlin 1 protein was reduced in E2-treated vs. untreated hypoxic animals, and ER-blockade abolished the inhibitory effect of E2 on Grem1 mRNA and protein. In conclusion, E2 ER-dependently regulates several genes involved in proliferative and inflammatory processes during hypoxia. Gremlin 1 is a novel target of the E2-ER axis in HPH. Understanding the mechanisms of E2 gene regulation in HPH may allow for selectively harnessing beneficial transcriptional activities of E2 for therapeutic purposes.
Project description:Heterochromatin protein 1 (HP1) is commonly seen as a key factor of repressive heterochromatin, even though a few genes are known to require HP1-chromatin for their expression. In order to obtain insight into the targeting of HP1 and its interplay with other chromatin components, we have mapped HP1 binding sites on chromosome 2 and 4 in Drosophila Kc cells using high-density oligonucleotide arrays and the DamID technique. The resulting high-resolution maps show that HP1 forms large domains in pericentric regions, but is targeted to single genes on chromosome arms. Intriguingly, HP1 shows a striking preference for exon-dense genes on chromosome arms. Furthermore, HP1 binds along entire transcription units, except for 5’ regions. Comparison with expression data shows that most of these genes are actively transcribed. HP1 target genes are also marked by the histone variant H3.3 and dimethylated histone 3 lysine 4 (H3K4me2), which are both typical of active chromatin. Interestingly, H3.3 deposition, which is usually observed along entire transcription units, is limited to the 5’ ends of HP1-bound genes. Thus, H3.3 and HP1 are mutually exclusive marks on active chromatin. Additionally, we observed that HP1-chromatin and Polycomb-chromatin are non-overlapping, but often closely juxtaposed, suggesting an interplay between both types of chromatin. These results demonstrate that HP1-chromatin is transcriptionally active and has extensive links with several other chromatin components. Keywords: DamID
Project description:HP1 alpha, beta, and gamma play an evolutionarily conserved role in the formation and maintenance of heterochromatin. In addition, some HP1 family members may also participate in transcriptional regulation of genes. Recently, HP1 gamma binding to the bodies of a subset of genes has been observed in human and murine cells. However, the generality of this phenomenon and the role HP1 gamma may play in this context are unknown. Genome-wide localization analysis reveals broad and general HP1 gamma binding at actively transcribed genes, which strongly correlates with gene activity across multiple cell types. Unexpectedly, we find HP1 gamma binding is necessary for efficient RNA processing of its target genes. Loss of HP1 gamma results in defective recruitment of splicing factors including U1-70K, and leads to accumulation of unspliced nascent transcripts genome-wide. Therefore, our data argue that the primary role of HP1 gamma is to ensure efficient RNA processing in mammalian cells.
Project description:lncRNA sequencing for 6 samples of the HPH (10% fractional inspired oxygen) and the control(N, 21% fractional inspired oxygen) groups.
Project description:m7G-MeRIP-sequencing for 6 samples of the HPH (10% fractional inspired oxygen) and the control(N, 21% fractional inspired oxygen) groups.