Abstract
The regulation of gene transcription requires posttranslational modifications of histones that, in concert with chromatin remodeling factors, shape the structure of chromatin. It is currently under intense investigation how this structure is modulated, in particular in the context of proliferation and differentiation. Compelling evidence suggests that the transcription factor NF-Y acts as a master regulator of cell cycle progression, activating the transcription of many cell cycle regulatory genes. However, the underlying molecular mechanisms are not yet completely understood. Here we show that NF-Y exerts its effect on transcription through the modulation of the histone “code”. NF-Y colocalizes with nascent RNA, while RNA polymerase II is I phosphorylated on serine 2 of the YSPTSPS repeats within its carboxyterminal domain and histones are carrying modifications that represent activation signals of gene expression (H3K9ac and PAN-H4ac). Comparing postmitotic muscle tissue from normal mice and proliferating muscles from mdx mice, we demonstrate by chromatin immunoprecipitation (ChIP) that NF-Y DNA binding activity correlates with the accumulation of acetylated histones H3 and H4 on promoters of key cell cycle regulatory genes, and with their active transcription. Accordingly, p300 is recruited onto the chromatin of NF-Y target genes in a NF-Y-dependent manner, as demonstrated by Re-ChIP. Conversely, the loss of NF-Y binding correlates with a decrease of acetylated histones, the recruitment of HDAC1, and a repressed heterochromatic state with enrichment of histones carrying modifications known to mediate silencing of gene expression (H3K9me3, H3K27me2 and H4K20me3). As a consequence, NF-Y target genes are downregulated in this context. In conclusion, our data indicate a role of NF-Y in modulating the structure and transcriptional competence of chromatin in vivo and support a model in which NF-Y-dependent histone “code” changes contribute to the proper discrimination between proliferating and postmitotic cells in vivo and in vitro.
Highlights
A major question in biology today is how the chromatin environments are interrelated with the machinery that drives transcription, and how this impacts on biological processes
The pattern of acetylated and methylated histones is different if we analyze the promoters of two muscle genes that are not regulated by NF-Y, Myogenin and Myosin Light Chain (MLC) (Figure 2C). These results suggest that, both in culture and in primary muscle cells, the association of NF-Y with CCAAT boxes correlates with hyperacetylation of histones H3 and H4; in contrast, the absence of NF-Y binding to promoters of cell cycle regulatory genes correlates with a local recruitment of heterochromatic markers, such as H3K9me3, H4K20me3
As cells progress through the cell cycle and differentiation, they must ensure that the right sections of their genomes are activated and repressed at the appropriate times
Summary
A major question in biology today is how the chromatin environments are interrelated with the machinery that drives transcription, and how this impacts on biological processes. Two major players of this code consist in acetylation and methylation of specific lysine residues within histones H3 and H4 [1,2,3,4]. Hyperacetylation correlates with transcriptional activation, while methylation of specific lysines is associated either with transcriptional repression or with activation [1,5,6,7]. It seems clear that transcription factors, chromatin remodellers and histone modification enzymes work together to guide the cell through cell cycle and differentiation as well as cell transformation [1,9,10,11,12]. Many transcriptional co-activators and co-repressor complexes with histone-modifying activities bind transcription factors in vivo, suggesting that targeted chromatin remodeling may be important for the function of transcription factors [13,14]. Transcription factors like Myc, E2F, p53 and Rb are implicated in chromatin remodeling [15,16,17,18,19,20]
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