Scatter plots and bar charts show mean and s.e.m., giving significance by Student’s and was more highly expressed in Foxa1/2cKO than in control, but is not required for differentiation at this stage of development; was also more highly expressed, but is required for positive selection and upregulated by TCR signal transduction, rather than functioning as a negative regulator (Bending et al., 2018; Georgopoulos, 2017; Costello et al., 2004; Wang et al., 2010). Open in a separate window Fig. of CD4SP, CD8SP and peripheral na?ve CD4+ T cells. Foxa1 and Foxa2 regulated the expression of many genes encoding splicing factors and regulators, including and locus, and requires TCR signalling: positive selection results in appropriate MHC restriction of SP cells, and is followed by negative selection of potentially self-reactive clones and selection of regulatory T cells (Tregs) (Huynh et al., 2014; Littman, 2016; Starr et al., 2003). The strength and duration of the TCR signal that a developing cell receives broadly determine its fate, with the strongest signals leading to negative selection or CD4 Treg differentiation, usually at the SP stage in the medulla, intermediate signals leading to positive selection usually in the cortex, and weaker signals or lack of TCR signalling leading to death by neglect (Singer et al., 2008). For DP thymocytes undergoing positive selection, TCR signal strength and duration also influence CD4 and CD8 lineage choice. Those cells receiving stronger and longer TCR signals tend towards the CD4SP fate, whereas weaker/more transient signals favour the CD8SP fate, and fate Benzamide decisions are also influenced by the relative timing of cytokine and TCR signalling that a developing cell receives (Littman, 2016; Klein et al., 2014; Bosselut, 2004). Many models have been proposed to describe this process and to explain how positive selection ensures that CD4SP and CD8SP populations express TCR appropriately restricted by MHCII Benzamide and MHCI, respectively (Littman, 2016; Starr et al., 2003; Carpenter and Bosselut, 2010). Currently, the consensus favours the kinetic signalling model (Littman, 2016; Singer et al., 2008; Egawa, 2015), in which CD8 is downregulated first during positive selection, leading to a CD4+CD8lo intermediate, with continued CD4 co-receptor expression allowing for prolonged stronger MHCII-TCR signalling, leading to differentiation to CD4SP, whereas cytokine signalling through the common gamma chain activates Stat5a and Stat5b and rescues cells that have received an interrupted MHCI-TCR signal to induce differentiation to CD8SP (Park et al., 2010; Brugnera et al., 2000). The CD4/CD8 lineage decision is also influenced by factors from the stroma, such Rabbit Polyclonal to EPHA7 (phospho-Tyr791) as Notch and Hedgehog (Hh) signalling (Laky and Fowlkes, 2008; Solanki et al., 2018; Furmanski et al., 2012; Rowbotham et al., 2007). Many transcription factors contribute to regulation of the CD4/CD8 lineage decision (Littman, 2016; Carpenter and Bosselut, 2010; Taniuchi, 2016; Naito et al., 2011). Additionally, epigenetic processes, such as DNA methylation and histone modification, may be involved in locking-in the pattern of gene expression to generate stable CD4SP and CD8SP Benzamide lineages (Issuree et al., 2017), and potentially also in preparing for initiation of a particular programme of differentiation. Here, we investigate the role of the transcription factors Foxa1 and Foxa2 in T-cell development. The Foxa proteins are a highly conserved subfamily of forkhead box transcription factors, which contain unique wing-helix DNA-binding domains (Jackson et al., 2010). The Foxa proteins can function as pioneer transcription factors, which by binding silent (condensed) chromatin early in a developmental programme prior to target gene activation, can act either to open up local chromatin, imparting competence to other transcriptional activators to initiate a developmental lineage or to directly facilitate other factors binding to nucleosomal DNA (Iwafuchi-Doi et al., 2016; Zaret, 2020). Foxa2 has recently also been shown to demethylate tissue-specific regions of DNA, to generate stable lineage-specific DNA methylation patterns that enhance gene expression (Reizel et al., 2021). Foxa1 and Foxa2 proteins are closely related to each other and are widely co-expressed during embryogenesis and in several tissues postnatally, including lung, liver, intestines, pancreas and thymus (Solanki et al., 2018; Kaestner, 2010; Besnard et al., 2004; Kaestner et al., 1994; Lau et al., 2018; Rowbotham et al., 2009). Genetic ablation of Foxa1 or Foxa2 in mice showed that they are both required for normal development during embryogenesis. Foxa2-deficient embryos display severe defects in notochord, floorplate and endoderm and die at embryonic day.