Project description:Phase separation, a biophysical process that segregates subcellular environments into condensates, is recognized for its role in regulating transcription. However, the extent of its influence on cellular senescence processes remains to be fully elucidated. Here, we have established that the depletion of MRG15, a conserved chromodomainprotein, leads to cellular senescence in human mesenchymal stem cells (hMSCs). MRG15 is capable of forming phase-separated liquid condensates via its intrinsically disordered region (IDR). Through the utilization of IDR deletion and replacement assays, we have shed light on the pivotal role that MRG15 condensation plays in the senescence of hMSCs. Through epigenomic and transcriptomic analysis, the depletion of MRG15 impacts pathways that are integral to the cell cycle and the senescence process, as evidenced by the diminished binding and the modified expression of key genes, including CDKN1A, LMNB1, CCNB1, NPM1, MYC, and HMGB2. Within a bleomycin-induced model of idiopathic pulmonary fibrosis in mice, hMSCs of the wild-type variety were able to ameliorate inflammation and fibrosis, a stark contrast to the outcomes observed in MSCs where MRG15 was knocked down or its IDR was deleted. Our findings establishes a link between phase separation and senescence regulation and could present a promising new therapeutic target for the alleviation of age-related diseases and the potential extension of lifespan.
Project description:Phase separation, a biophysical process that segregates subcellular environments into condensates, is recognized for its role in regulating transcription. However, the extent of its influence on cellular senescence processes remains to be fully elucidated. Here, we have established that the depletion of MRG15, a conserved chromodomainprotein, leads to cellular senescence in human mesenchymal stem cells (hMSCs). MRG15 is capable of forming phase-separated liquid condensates via its intrinsically disordered region (IDR). Through the utilization of IDR deletion and replacement assays, we have shed light on the pivotal role that MRG15 condensation plays in the senescence of hMSCs. Through epigenomic and transcriptomic analysis, the depletion of MRG15 impacts pathways that are integral to the cell cycle and the senescence process, as evidenced by the diminished binding and the modified expression of key genes, including CDKN1A, LMNB1, CCNB1, NPM1, MYC, and HMGB2. Within a bleomycin-induced model of idiopathic pulmonary fibrosis in mice, hMSCs of the wild-type variety were able to ameliorate inflammation and fibrosis, a stark contrast to the outcomes observed in MSCs where MRG15 was knocked down or its IDR was deleted. Our findings establishes a link between phase separation and senescence regulation and could present a promising new therapeutic target for the alleviation of age-related diseases and the potential extension of lifespan.
Project description:The aim of this analysis is to identify the difference between interaction partners of MRG15 wild type and Chromodomain and MRG domain mutants upon UV irradiation. Therefore, MRG15 wild type and mutants (N-terminal FLAG tag) were overexpressed in U2OS cells. After UV irradiation, immunoprecipitation of the constructs was performed using FLAG M2 affinity gel. The constructs were eluted from the beads with 3xFLAG peptide.
Project description:SPARK-OFF tag does not perturb the core transcriptional function of YAP-MAML2.~2260 DEGs with ≥ 1.5-fold change in transcript level, including ~1270 up-regulated genes and ~990 down-regulated genes can be identified from YAP-MAML2 overexpression HEK293T cells.We identified 88 DEGs (p-value < 0.01, |Log2FC| ≥ 0.58, FDR < 0.1) that are regulated by YAP-MAML2 phase separation with ≥ 1.5-fold change in transcript levels, including 44 up-regulated genes and 44 down-regulated genes
Project description:Ash1 is a classic Trithorax group protein possessing an H3K36-specific histone methyltransferase activity. Ash1 plays a critical role in antagonizing Polycomb silencing and its loss-of-function mutations lead to inactivation of certain Hox genes and homeotic transformation. Here, we report the purification of Ash1 complex with the identification of two novel subunits, Mrg15 and Nurf55. Interestingly, Mrg15 stimulates the enzymatic activity of Ash1 in vitro, and such stimulation is independent of the chromo domain of Mrg15. In vivo, Mrg15 is recruited by Ash1 to their common target genes and Mrg15 is essential for the proper deposition of H3K36me2 at these regions.
Project description:Epigenetic reprogramming requires depletion of 4 factors ; Oct4, Sox2, Klf-4 and c-Myc. We have 4 groups of samples. 2 group is collected as fibroblasts and it is referred as Day Zero. The other groups are collected at the sixth day of epigenetic reprogramming referred as Day Six. Day 18 of epigenetic reprogramming, %5- 15 of cell population transformed into pluripotent stem cells. At day six this percentage is lower.Our previous studies have showed that inhibition of H3K36me3 mark thusleading to H3K36me2 mark on chromatin) significantly increased reprogramming efficiency. A CRISPR Knock-Out Library was built to identify which of the H3K36me3 mark reader protein can phenocopy absence of K36me3 loss in terms of reprogramming efficiency. MRG15 Knock Out has showed the most similar increase in reprogramming as loss of K36me3 mark. It has been reported that MRG15 is responsible from alternative splicing of some genes. Our hypothesis is; in the presence of H3K36me3 mark MRG15 can bind this mark on chromatin and recruit proteins related to alternative splicing ( eg. PTBP1)and regulate alternative splicing.