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.Human body fat consists mostly of fat-storing cells called adipocytes. These cells develop in two main steps. First, mesenchymal stem cells—which can potentially become one of many different types of cell—commit to becoming pre-adipocyte cells. These pre-adipocytes then develop into mature adipocytes. Proteins called transcription factors control both steps of this process by binding to particular sites in the DNA of the cell and activating certain genes that control the cell's identity and activity.Various different transcription factors are known to stimulate the development of mesenchymal stem cells into adipocytes. Experiments performed on cells that have been grown in the laboratory suggest that the cells must also be tightly packed together to become adipocytes.Cohen et al. have now investigated the role a protein called C/EBPβ plays in the development of adipocytes, and have found that it plays different roles at different stages of development. When mesenchymal stem cells become tightly packed, more of another protein called ATF4 is produced. This protein binds to C/EBPβ, and the resulting two-protein complex then binds to sites on the DNA of the mesenchymal stem cell to activate genes that turn the stem cell into a pre-adipocyte. Reducing the amount of ATF4 in mesenchymal stem cells reduces the number of pre-adipocytes that develop.When not bound to ATF4, and in response to certain cell signals, C/EBPβ binds to different DNA sites and helps pre-adipocytes develop into mature adipocytes. Whether transcription factors other than ATF4 partner with C/EBPβ to change DNA-binding-site preferences remains unknown. Future studies will search for these, with the aim of providing a clearer molecular understanding for how C/EBPβ acts as a ‘bridge’ that links together the two stages of adipocyte development.