IGF1-mediated human embryonic stem cell self-renewal recapitulates the embryonic niche
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PROVIDER: PRJNA522065 | ENA |
REPOSITORIES: ENA
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Project description:One of the most striking epigenetic alterations that occurs at the level of the nucleosome is the complete exchange of the canonical H2A histones for the macroH2A variant. Here, we provide insight in the function of this unique histone variant in embryonic and adult stem cells. Knockdown of macroH2A1 in mouse embryonic stem (mES) cells limited their capacity to differentiate but not their self-renewal. The loss of macroH2A1 interfered with the proper activation of differentiation genes, most of which are direct target genes of macroH2A. Additionally, macroH2A1-deficient mES cells displayed incomplete inactivation of pluripotency genes and formed defective embryoid bodies. In vivo, macroH2A1-deficient teratomas contained a massive expansion of malignant, undifferentiated carcinoma tissue. In the heterogeneous culture of primary human keratinocytes, macroH2A1 levels negatively correlated with the self-renewal capacity of the pluripotent compartment. Together these results establish macroH2A1 as a critical chromatin component that regulates the delicate balance between self-renewal and differentiation of embryonic and adult stem cells. Examination of the histone variant macroH2A1 in mouse Embryonic stem cells
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Project description:Recent studies have shown that the RNA binding protein Musashi 2 (Msi2) plays prominent roles during development and leukemia. Additionally, in embryonic stem cells (ESC) undergoing the early stages of differentiation, Msi2 has been shown to associate with Sox2, which is required for the self-renewal of ESC. These findings led us to examine the effects of Msi2 on the behavior of ESC. Using an shRNA sequence that targets Msi2 and a scrambled shRNA sequence, we determined that knockdown of Msi2 disrupts the self-renewal of ESC and promotes their differentiation. Collectively, our findings argue that Msi2 is required to support the self-renewal and pluripotency of ESC. We used microarrays to better understand global changes in ESC gene expression following the knockdown of the RNA-binding protein Msi2 as compared to control ESC expressing a scrambled shRNA. Mouse embryonic stem cells (D3) were treated with lentivirus engineered for the expression of a Msi2 targeting shRNA sequence or scrambled (control) shRNA sequence. Following selection for infected cells with puromycin, cells were subcultured at low density and allowed to grow for 4 days (see treatment protocol) before RNA was extracted. RNA was collected and analyzed one time for each of the two samples.
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