Abstract
Nuclear factor of activated T cells (NFAT) comprises a family of transcription factors that regulate T cell development, activation and differentiation. NFAT signalling can also mediate granulocyte and dendritic cell (DC) activation, but it is unknown whether NFAT influences their development from progenitors. Here, we report a novel role for calcineurin/NFAT signalling as a negative regulator of myeloid haematopoiesis. Reconstituting lethally irradiated mice with haematopoietic stem cells expressing an NFAT-inhibitory peptide resulted in enhanced development of the myeloid compartment. Culturing bone marrow cells in media supplemented with Flt3-L in the presence of the calcineurin/NFAT inhibitor Cyclosporin A increased numbers of differentiated DC. Global gene expression analysis of untreated DC and NFAT-inhibited DC revealed differential expression of transcripts that regulate cell cycle and apoptosis. In conclusion, these results provide evidence that calcineurin/NFAT signalling negatively regulates myeloid lineage development. The finding that inhibition of NFAT enhances myeloid development provides a novel insight into understanding how the treatment with drugs targeting calcineurin/NFAT signalling influence the homeostasis of the innate immune system.
Highlights
Nuclear factor of activated T cells (NFAT) comprises a family of transcription factors originally described as regulators of interleukin-2 (IL-2) expression in T cells (Shaw et al, 1988)
NFAT1 expression is up-regulated upon fms-like tyrosine kinase receptor-3 ligand (Flt3-L) stimulation of haematopoietic stem cells lines In order to achieve long-term, stable inhibition of calcineurin/ NFAT signalling, we introduced the VIVIT peptide (Aramburu et al, 1999) into established, long-term haematopoietic ‘stem cell-like’ cell lines
haematopoietic stem cells (HSCs) lines have all of the key characteristics of their in vivo HSC counterparts; they reconstitute haematopoiesis when transplanted into an irradiated host, they can be serially transplanted for many generations, they support selfrenewal and generation of progenitor cells, and they can differentiate into dendritic cell (DC) in vitro in the presence of GM-CSF or Flt3-L (Ruedl et al, 2008) In the current study, HSC lines were transduced with constructs expressing either VIVIT-IRES-eGFP or control-IRES-tdTomato, and fluorescent cells were sorted for further analyses
Summary
Nuclear factor of activated T cells (NFAT) comprises a family of transcription factors originally described as regulators of interleukin-2 (IL-2) expression in T cells (Shaw et al, 1988). Hyperactivation of NFAT1 is deleterious in embryogenesis and restricts the development of lymphocytes and their progenitors (Muller et al, 2009). Calcineurin/NFAT signalling is an important mediator of T cell selection in the thymus (Cante-Barrett et al, 2007; Gallo et al, 2008; Muller et al, 2009). While NFAT expression in haematopoietic progenitor cells has been reported previously (Kiani et al, 2004, 2007) and appears to play a significant role in lymphopoiesis (Muller et al, 2009), it is unclear how far NFAT signalling regulates the development of other leukocyte populations during myeloid differentiation
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