Noer Agate, Lindeman Leif C, Collas Philippe
Department of Biochemistry, Institute of Basic Medical Sciences, Faculty of Medicine, University of Oslo, Oslo, Norway.
Stem Cells Dev. 2009 Jun;18(5):725-36. doi: 10.1089/scd.2008.0189.
Long-term culture of mesenchymal stem cells leads to a loss of differentiation capacity, the molecular mechanism of which remains not understood. We show here that expansion of adipose stem cells (ASCs) to late passage (replicative senescence) is associated with promoter-specific and global changes in epigenetic histone modifications. In undifferentiated ASCs, inactive adipogenic and myogenic promoters are enriched in a repressive combination of trimethylated H3K4 (H3K4m3) and H3K27m3 in the absence of H3K9m3, a heterochromatin mark. Sequential chromatin immunoprecipitation assays indicate that H3K4m3 and H3K27m3 co-occupy a fraction of nucleosomes on some but not all lineage-specific promoters examined. However in cultured primary keratinocytes, adipogenic and myogenic promoters are enriched in trimethylated H3K4, K27, and K9, illustrating two distinct epigenetic states of inactive promoters related to potential for activation. H3K4m3 and H3K27m3 stably mark promoters during long-term ASC culture indicating that loss of differentiation capacity is not due to alterations in these histone modifications on these loci. Adipogenic differentiation in early passage results in H3K27 demethylation and H3K9 acetylation specifically on adipogenic promoters. On induction of differentiation in late passage, however, transcriptional upregulation is impaired, H3K27 trimethylation is maintained and H3K9 acetylation is inhibited on promoters. In addition, the polycomb proteins Ezh2 and Bmi1 are targeted to promoters. This correlates with global cellular Ezh2 increase and H3K9 deacetylation. Promoter targeting by Ezh2 and Bmi1 in late passage ASCs suggests the establishment of a polycomb-mediated epigenetic program aiming at repressing transcription.
间充质干细胞的长期培养会导致分化能力丧失，其分子机制尚不清楚。我们在此表明，脂肪干细胞（ASC）扩增至后期传代（复制性衰老）与表观遗传组蛋白修饰的启动子特异性和全局变化相关。在未分化的ASC中，无活性的成脂和生肌启动子在缺乏异染色质标记H3K9m3的情况下，富含三甲基化H3K4（H3K4m3）和H3K27m3的抑制性组合。连续染色质免疫沉淀分析表明，H3K4m3和H3K27m3共同占据了一些但并非所有检测的谱系特异性启动子上的一部分核小体。然而，在培养的原代角质形成细胞中，成脂和生肌启动子富含三甲基化H3K4、K27和K9，说明了与激活潜力相关的无活性启动子的两种不同表观遗传状态。在ASC长期培养过程中，H3K4m3和H3K27m3稳定地标记载体，表明分化能力丧失并非由于这些基因座上这些组蛋白修饰的改变。早期传代的成脂分化导致H3K27去甲基化和H3K9乙酰化，特别是在成脂启动子上。然而，在后期传代诱导分化时，转录上调受损，启动子上H3K27三甲基化得以维持，H3K9乙酰化受到抑制。此外，多梳蛋白Ezh2和Bmi1靶向启动子。这与全局细胞Ezh2增加和H3K9去乙酰化相关。后期传代ASC中Ezh2和Bmi1对启动子的靶向作用表明建立了一个旨在抑制转录的多梳介导的表观遗传程序。
