The neonatal microenvironment programs conventional and intestinal Tbet⁺γδT17 cells through the transcription factor STAT5

Abstract

15Interleukin(IL)-17-producing RORγt + γδ T (γδT17) cells develop in the embryonic thymus and 16 participate in type 3 immune responses. Herein we show that γδT17 cells rapidly proliferate within 17 neonatal lymph nodes and gut, where upon entry they uniquely upregulate Tbet and co-express IL-18 17, IL-22 and interferon(IFN) γ in a STAT3 and retinoic acid dependent manner. Neonatal 19 expansion was halted in mice conditionally deficient in STAT5 and its loss resulted in γδT17 cell 20 depletion from all adult organs. Hyperactive STAT5 mutant mice showed that the STAT5A 21 homologue had a dominant role over STAT5B in promoting γδT17 cell expansion and 22 downregulating gut-associated Tbet. In contrast, STAT5B preferentially expanded IFNγ-producing 23 γδ populations. Importantly, mice lacking γδT17 cells due to STAT5 deficiency displayed a 24 profound resistance to experimental autoimmune encephalomyelitis. Our data identify for the first 25 time STAT5 as a key molecular checkpoint allowing γδT17 cells to pass through a critical neonatal 26 developmental window to acquire tissue-specific characteristics essential for infection and 27 autoimmunity.28 subtypes, such as hepatosplenic T cell lymphoma (Nicolae et al., 2014), monomorphic 81 epitheliotropic intestinal T cell lymphoma (Kucuk et al., 2015; Nairismagi et al., 2016) and primary 82 cutaneous γδ T cell lymphoma (Kucuk et al., 2015). Notably, approximately 20% of identified 83 N642H mutations occur in γδ T cell derived lymphomas (de Araujo ED, 2019).84 4Herein, we show that STAT5 is critically required for the progression and expansion of γδT17 85 cells through neonatal life in the intestine and periphery. We provide evidence that intestinal γδT17 86 cells upregulate Tbet upon entry into the lamina propria after birth and co-express the cytokines IL-87 17, IL-22 and IFNγ in a mechanism dependent on STAT3 and retinoic acid. Furthermore, loss of 88 γδT17 cells due to STAT5 deficiency results in resistance to experimental autoimmune 89 encephalomyelitis in adult mice. Importantly, we show that STAT5A promotes γδT17 cell 90 expansion and downregulates intestinal Tbet favoring a type 17 program, whereas STAT5B favors 91 IFNγ-producing γδ populations and increases intestinal Tbet expression. Collectively, our data 92 suggest that neonatal life is a critical window of development and tissue specification for γδT17 93 cells, and that this process is tightly regulated by STAT5. 94 95 Results 96 STAT5 regulates the neonatal expansion of γδT17 cells 97 In order to test the importance of STAT5 in RORγt expressing γδ T cells, we crossed RORγt-Cre 98 mice (Eberl and Littman, 2004) with mice floxed for both STAT5a and STAT5b (RORγt CRE -99

The cIAP ubiquitin ligases sustain type 3 γδ T and innate lymphoid cells during aging to allow normal cutaneous and mucosal responses

“…We have previously shown that antagonization of cIAP1 and cIAP2 in Th17 cells downregulates Il17a and Maf through the non-canonical NF-κB transcription factors RelB and p52 (Rizk et al, 2019). Thus, it is plausible that a similar mechanism exists in γδT17 cells; as such, the lack of cIAP1 and cIAP2 in γδT17 activates the non-canonical NF-κB, which downregulates Maf expression.…”

“…We have previously shown that the non-canonical NF-κB transcription factor RelB downregulates the expression of the transcription factor cMAF in Th17 cells; this occurs following the activation of the non-canonical NF-κB pathway through depletion of cIAP1 and 2 by SMAC mimetic compounds (Rizk et al, 2019). Thus, we hypothesized that the lack of cIAP1 and 2 expression in γδT17 activates the non-canonical NF-κB pathway and mobilizes RelB to downregulate cMAF.…”

“…We have previously characterized a population of RORγt + Tbet + γδ T cells in the small intestinal (siLP) and colonic lamina propria (cLP) (Kadekar et al, 2019). Thus, we investigated the impact of cIAP1 and cIAP2 deficiency on the different RORγt + γδ T cells in gut lamina propria.…”

“…Lymphocytes were isolated from peripheral lymph nodes (axial, brachial and inguinal), thymus, ear skin, small intestinal and colonic lamina propria following the previously described protocols (Kadekar et al, 2019). Lymphocytes were isolated from cervical and auricular lymph nodes in case of IMQ-induced psoriasis.…”

“…Surface antigens, intracellular cytokines and transcription factors were stained for flow cytometry as previously described (Rizk et al, 2019). The following antibodies were used herein at 1:200 dilution unless otherwise indicated:…”