Selected article for: "cell isolation and flow cytometry"

Author: Djeghloul, Dounia; Patel, Bhavik; Kramer, Holger; Dimond, Andrew; Whilding, Chad; Brown, Karen; Kohler, Anne-Céline; Feytout, Amelie; Veland, Nicolas; Elliott, James; Bharat, Tanmay A M; Tarafder, Abul K; Löwe, Jan; Ng, Bee L; Guo, Ya; Guy, Jacky; Huseyin, Miles K; Klose, Robert J; Merkenschlager, Matthias; Fisher, Amanda G
Title: Identifying proteins bound to native mitotic ESC chromosomes reveals chromatin repressors are important for compaction.
  • Cord-id: ng9o0svz
  • Document date: 2020_8_17
  • ID: ng9o0svz
    Snippet: Epigenetic information is transmitted from mother to daughter cells through mitosis. Here, to identify factors that might play a role in conveying epigenetic memory through cell division, we report on the isolation of unfixed, native chromosomes from metaphase-arrested cells using flow cytometry and perform LC-MS/MS to identify chromosome-bound proteins. A quantitative proteomic comparison between metaphase-arrested cell lysates and chromosome-sorted samples reveals a cohort of proteins that wer
    Document: Epigenetic information is transmitted from mother to daughter cells through mitosis. Here, to identify factors that might play a role in conveying epigenetic memory through cell division, we report on the isolation of unfixed, native chromosomes from metaphase-arrested cells using flow cytometry and perform LC-MS/MS to identify chromosome-bound proteins. A quantitative proteomic comparison between metaphase-arrested cell lysates and chromosome-sorted samples reveals a cohort of proteins that were significantly enriched on mitotic ESC chromosomes. These include pluripotency-associated transcription factors, repressive chromatin-modifiers such as PRC2 and DNA methyl-transferases, and proteins governing chromosome architecture. Deletion of PRC2, Dnmt1/3a/3b or Mecp2 in ESCs leads to an increase in the size of individual mitotic chromosomes, consistent with de-condensation. Similar results were obtained by the experimental cleavage of cohesin. Thus, we identify chromosome-bound factors in pluripotent stem cells during mitosis and reveal that PRC2, DNA methylation and Mecp2 are required to maintain chromosome compaction.

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