TNFα-Signaling Modulates the Kinase Activity of Human Effector Treg and Regulates IL-17A Expression

Abstract

Maintenance of regulatory T cells CD4 + CD25 high FOXP3 + (Treg) stability is vital for proper Treg function and controlling the immune equilibrium. Treg cells are heterogeneous and can reveal plasticity, exemplified by their potential to express IL-17A. TNFα-TNFR2 signaling controls IL-17A expression in conventional T cells via the anti-inflammatory ubiquitin-editing and kinase activity regulating enzyme TNFAIP3/A20 (tumor necrosis factor-alpha-induced protein 3). To obtain a molecular understanding of TNFα signaling on IL-17 expression in the human effector (eff Treg, CD25 high CD45RA −) Treg subset, we here studied the kinome activity regulation by TNFα signaling. Using FACS-sorted naïve (naïve Treg, CD25 high CD45RA +) and eff Treg subsets, we demonstrated a reciprocal relationship between TNFα and IL-17A expression; eff Treg (TNFα low /IL-17A high) and naïve Treg (TNFα high /IL-17A low). In eff Treg, TNFα-TNFR2 signaling prevented IL-17A expression, whereas inhibition of TNFα signaling by clinically applied anti-TNF antibodies led to increased IL-17A expression. Inhibition of TNFα signaling led to reduced TNFAIP3 expression, which, by using siRNA inhibition of TNFAIP3, appeared causally linked to increased IL-17A expression in eff Treg. Kinome activity screening of CD3/CD28-activated eff Treg revealed that anti-TNF-mediated neutralization led to increased kinase activity. STRING association analysis revealed that the TNF suppression eff Treg kinase activity network was strongly associated with kinases involved in TCR, JAK, MAPK, and PKC pathway signaling. Small-molecule-based inhibition of TCR and JAK pathways prevented the IL-17 expression in eff Treg. Together, these findings stress the importance of TNF-TNFR2 in regulating the kinase architecture of antigen-activated eff Treg and controlling IL-17 expression of the human Treg. These findings might be relevant for optimizing anti-TNF-based therapy and may aid in preventing Treg plasticity in case of Treg-based cell therapy.

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“…Signaling pathways activated by TNFR2 include the NF-κB and JNK pathways, canonical and non-canonical ( Cabal-Hierro et al, 2014 ). TNFα/TNFR2 signaling can regulate the enzyme TNFAIP3/A20 to reduce IL-17A expression in conventional T cells, thereby exhibiting an anti-inflammatory effect ( Urbano et al, 2019 ). Therefore, the TNF signaling pathway is also a double-edged sword and has potential research value.…”

Targeting TNFR2 in Cancer: All Roads Lead to Rome

“…Tregs are the most extensively studied immunosuppressive cells, and they are defined as CD4+CD25+Foxp3+ or CD4+CD25+ CD127low T cells (33,34). Current research suggests that TNFR2 is highly expressed in Tregs, especially in effector Tregs, while TNFR1 is hardly detected (35)(36)(37). The presence of high Tregs, especially TNFR2+ Tregs in the TME, is associated with an unfavorable prognosis in various types of cancers (38)(39)(40).…”

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“…TNF also inhibits the differentiation of TH17 cells by increasing IL-2 production 69 , and decreases IL-17 production by conventional T cells and effector Treg cells via activation of TNFAIP3 70,71 . This mechanism might explain the increase in numbers of TH17 cells described in TNFR1-knockout mice or after treatment with TNF inhibitors in mouse models of RA and psoriasis [44][45][46]72,73 .…”

“…Thus, TNF might increase Treg cell suppression and stability by favouring both phosphorylation of STAT5 and FOXP3 expression, which are key determinants of these Treg cell features 139,140 . Finally, TNF limits IL-17 production by Treg cells by activating TNFAIP3 71 .…”