The TEAD family of transcription factors (TEAD1-4) bind the MCAT element in the regulatory elements of both growth promoting and myogenic differentiation genes. Defining TEAD transcription factor function in myogenesis has proved elusive due to overlapping expression of family members and their functional redundancy. We show that silencing of either Tead1, Tead2 or Tead4 did not effect primary myoblast (PM) differentiation, but that their simultaneous knockdown strongly impaired differentiation. In contrast, Tead1 or Tead4 silencing impaired C2C12 differentiation showing their different contributions in PMs and C2C12 cells. Chromatin immunoprecipitation identified enhancers associated with myogenic genes bound by combinations of Tead4, Myod1 or Myog. Tead4 regulated distinct gene sets in C2C12 cells and PMs involving both activation of the myogenic program and repression of growth and signaling pathways. ChIP-seq from mature mouse muscle fibres in vivo identified a set of highly transcribed muscle cell-identity genes and sites bound by Tead1 and Tead4. Although inactivation of Tead4 in mature muscle fibres caused no obvious phenotype under normal conditions, notexin-induced muscle regeneration was delayed in Tead4 mutants suggesting an important role in myogenic differentiation in vivo. By combining knockdown in cell models in vitro with Tead4 inactivation in muscle in vivo, we provide the first comprehensive description of the specific and redundant roles of Tead factors in myogenic differentiation.

Understanding tumor heterogeneity is an important challenge in current cancer research. Transcription and epigenetic profiling of cultured melanoma cells have defined at least two distinct cell phenotypes characterized by distinctive gene expression signatures associated with high or low/absent expression of microphthalmia-associated transcription factor (MITF). Nevertheless, heterogeneity of cell populations and gene expression in primary human tumors is much less well characterized. We performed single-cell gene expression analyses on 472 cells isolated from needle biopsies of 5 primary human melanomas, 4 superficial spreading, and one acral melanoma. The expression of -high and-low signature genes was assessed and compared to investigate intra- and intertumoral heterogeneity and correlated gene expression profiles. Single-cell gene expression analyses revealed varying degrees of intra- and intertumor heterogeneity conferred by the variable expression of distinct sets of genes in different tumors. Expression of partially correlated with that of its known target genes, while expression correlated best with and Nevertheless, cells simultaneously expressing -high and-low signature genes were observed both by single-cell analyses and RNAscope. Single-cell analyses can be performed on limiting numbers of cells from primary human melanomas revealing their heterogeneity. Although tumors comprised variable proportions of cells with the -high and-low gene expression signatures characteristic of melanoma cultures, primary tumors also comprised cells expressing markers of both signatures defining a novel cell state in tumors .