Expression of the transcription factor NFAT2 in human neutrophils: IgE-dependent, Ca2+- and calcineurin-mediated NFAT2 activation

Abstract

NFAT (nuclear factors of activated T cells) proteins constitute a family of transcription factors involved in mediating signal transduction. The presence of NFAT isoforms has been described in all cell types of the immune system, with the exception of neutrophils. In the present work we report for the first time the expression in human neutrophils of NFAT2 mRNA and protein. We also report that specific antigens were able to promote NFAT2 protein translocation to the nucleus, an effect that was mimicked by the treatment of neutrophils with anti-immunoglobulin E (anti-IgE) or anti-Fcϵ-receptor antibodies. Antigens, anti-IgE and anti-FcϵRs also increased Ca2+ release and the intracellular activity of calcineurin, which was able to interact physically with NFAT2, in parallel to eliciting an enhanced NFAT2 DNA-binding activity. In addition, specific chemical inhibitors of the NFAT pathway, such as cyclosporin A and VIVIT peptide, abolished antigen and anti-IgE-induced cyclooxygenase-2 (COX2) gene upregulation and prostaglandin (PGE2) release, suggesting that this process is through NFAT. Our results provide evidence that NFAT2 is constitutively expressed in human neutrophils, and after IgE-dependent activation operates as a transcription factor in the modulation of genes, such as COX2, during allergic inflammation.

Efficient Neutrophil Extracellular Trap Induction Requires Mobilization of Both Intracellular and Extracellular Calcium Pools and Is Modulated by Cyclosporine A

“…In this context active calcineurin has previously been detected in human neutrophils [36], [39], [55], as well as other members of the signalling cascade such as NFAT2 (nuclear factor of activated T cells 2) [37]. In addition a number of previous studies have indicated calcineurin inhibition significantly influenced neutrophil activity, inhibiting chemokinesis, adhesion and motility on vitronectin, response to angiotensin II, as well as a reduction in phagocytic activity [38]–[41], [43], [44].…”

“…This investigation was prompted by a number of reports indicating that human neutrophils possess a functional calcineurin system [36], [39], [55], including nuclear signalling components such as NFAT2 [37], and that its pharmacological modulation influences neutrophil behaviour [38]–[41], [43], [44].…”

NFAT control of innate immunity

“…77 NFAT signaling has now been demonstrated in several different types of granulocytes (Table 1). NFAT expression was first described as an important inducer of inflammatory responses in human neutrophils, 82 which express both NFAT2 and NFAT4. IgE-mediated neutrophil expression of cyclo-oxygenase and release of prostaglandins are regulated by NFAT translocation.…”

“…IgE-mediated neutrophil expression of cyclo-oxygenase and release of prostaglandins are regulated by NFAT translocation. 82 Expression of IL-10, Cox2, Egr1, and Egr2 genes is impaired in mice that conditionally lack calcineurin subunit B in neutrophils. Each of these genes is dependent on calcineurin/NFAT signaling and exerts an important role in innate immune responses.…”

Distinct and additive effects of sodium bicarbonate and continuous mild heat stress on fiber type shift via calcineurin/NFAT pathway in human skeletal myoblasts

“…Quantitative real-time RT-PCR (qPCR) was performed using the SYBR Premix Ex Taq (Perfect Real Time) Premix (TaKaRa Bio, Otsu, Japan) or the LightCycler FastStart DNA Master SYBR Green I kit (Roche, Mannheim, Germany) in a fluorescent temperature cycler (LightCycler; Roche). Primer sequences were obtained from previous publications (6,15,35,38) (Table 1). The qPCR protocol included initial denaturation at 95°C for 10 min (LightCycler FastStart) or 10 s (SYBR Premix), followed by 40 -50 PCR cycles of 95°C for 5 s and 60°C for 20 s (SYBR Premix), or 95°C for 10 s, 60°C for 10 s, and 72°C for 12 s (LightCycler FastStart).…”