ATF4 licenses C/EBPβ activity in human mesenchymal stem cells primed for adipogenesis

Abstract

A well-established cascade of transcription factor (TF) activity orchestrates adipogenesis in response to chemical cues, yet how cell-intrinsic determinants of differentiation such as cell shape and/or seeding density inform this transcriptional program remain enigmatic. Here, we uncover a novel mechanism licensing transcription in human mesenchymal stem cells (hMSCs) adipogenically primed by confluence. Prior to adipogenesis, confluency promotes heterodimer recruitment of the bZip TFs C/EBPβ and ATF4 to a non-canonical C/EBP DNA sequence. ATF4 depletion decreases both cell-density-dependent transcription and adipocyte differentiation. Global profiling in hMSCs and a novel cell-free assay reveals that ATF4 requires C/EBPβ for genomic binding at a motif distinct from that bound by the C/EBPβ homodimer. Our observations demonstrate that C/EBPβ bridges the transcriptional programs in naïve, confluent cells and early differentiating pre-adipocytes. Moreover, they suggest that homo- and heterodimer formation poise C/EBPβ to execute diverse and stage-specific transcriptional programs by exploiting an expanded motif repertoire.DOI: http://dx.doi.org/10.7554/eLife.06821.001

Exploring the bZIP regulatory network in the sponge Amphimedon queenslandica: insights into the ancestral animal regulatory genome

“…AqATF4 transcripts appear to be localised around the nuclei of mesenchymal archeocytes, which are pluripotent stem cells (Funayama, 2013). Interestingly, the human ATF4:CEBP heterodimer is a key regulator of cell differentiation from mesenchymal stem cells (Cohen et al, 2015). However, since AqATF4 and AqCEBP do not form a strong dimer in A.…”

“…AqATF4 peaks were detected using MACS2 and a p-value threshold of 1e -05 (-p 0.00001, --call-summits, --nomodel, --extsize 146, -g 1.45e8). Choice of peak calling algorithm and parameters were based on published similar work (Cohen et al, 2015;Han et al, 2013;Huggins et al, 2016;Li et al, 2011;Sebe-Pedros et al, 2016). AqATF4 peaks were also detected using the Peakzilla software (Bardet et al, 2013) relative to input DNA (-c 2 -s 3).…”

“…Using the widely used peak-calling algorithm MACS2 (Zhang et al, 2008) with stringent parameters (Cohen et al, 2015;Han et al, 2013;Landt et al, 2012;Li et al, 2011;Sebe-Pedros et al, 2016), 1,756 discrete AqATF4 peaks were identified (Appendix 5.4). Another peak-caller, Peakzilla (Bardet et al, 2013), identified 814 peaks, 80% of which belonged to the MACS2 peak sets.…”

“…Other significantly enriched motifs included a Zinc finger and an IRX binding motifs ( Figure 5.8A). Since the motifs discovered by MEME were only present in a small, but significant, subset of AqATF4 peaks, I completed this analysis using HOMER (Heinz et al, 2010), another widely used motif-mining tool (Cohen et al, 2015;Heinz et al, 2010;Sebe-Pedros et al, 2016). HOMER identified a wider range of motifs, e.g., several e3 ubiquitin-protein ligases).…”

“…Interestingly, distal binding of three bZIPs -ATF4 in human mesenchymal stem cells (Cohen et al, 2015), JUN and CEBP in murine macrophages (Heinz et al, 2015) -to enhancer elements has been shown to be critical for terminal cell differentiation. bZIP general occupancy in extended TSS-proximal regions that also contain enhancer chromatin signatures in bilaterians (Peter and Davidson, 2015) may, thus, reflect ATF4 enrichment both in promoter and putative enhancer regions.…”