Hematopoietic cell transplantation (HCT) is a curative therapy for blood cancers but is complicated by graft-versus-host disease (GVHD), where donor CD4+ T helper (Th) cells drive gut pathology. Defining factors that drive pathogenic Th cell differentiation in the gut promises to inform therapies that mitigate GVHD without impacting the beneficial anti-cancer activity of HCT therapy. In our work, we identified GATA3 as a key transcription factor that may facilitate the development of gut pathogenic Th cells after HCT. We hypothesized that Th cell deletion of GATA3 will impair pathogenic Th cell differentiation and improve clinical outcomes in a murine GVHD model.
Using a murine allo-HCT model, we transplanted T cell-depleted bone marrow from WT C57BL/6 donors and splenocytes from C57BL/6-Gata3fl/fl Rosa26ERT2-Cre donors into irradiated BALB/c recipients. Donor cell GATA3 deletion was induced via tamoxifen treatment of recipient mice. Disease severity, intestinal pathology, donor cell trafficking, cytokine/granzyme expression, and innate immune infiltration were assessed by clinical scoring, histology, flow cytometry, and ex vivo Th cell stimulation.
GATA3 deletion reduced intestinal GVHD pathology and impaired early donor T cell trafficking and persistence in the gut. While IFN-γ and IL-17A production were largely unaffected in early timepoints, GATA3 was required for heightened granzyme A expression, sustained GM-CSF production and chronic GM-CSF-driven intestinal eosinophilia at later timepoints. Analysis of human GVHD samples exhibited enrichment of eosinophil chemotactic pathways, mirroring murine findings.
GATA3 promotes intestinal GVHD by supporting donor T cell gut infiltration, granzyme A expression, and sustained GM-CSF-dependent eosinophilia. These findings identify GATA3 as a key regulator of pathogenic Th responses and a potential therapeutic target for mitigating intestinal GVHD.
Purdue Institute for Cancer Research
Transplantation Immunology (TRAN)
Franklin J Yeo, Mengbo Wang, Sungtae Park et al.· Journal of Immunology· 0 citations
Enhancer RNAs (eRNAs) are noncoding transcripts from active enhancers whose functions in adaptive immunity are poorly defined. Because small changes in signaling strength can alter T cell fate and B cell help, we hypothesized that eRNAs act as rheostats for key fate decisions and signaling modules shaping antigen-induced immune responses.
We integrated rRNA-depleted RNA-seq, ATAC-seq, and ChIP-seq to map transcribed enhancers in human B, CD4, and CD8 T cells. We then focused on a conserved eRNA ∼140 kb upstream of KRAS (eKRAS) and tested its function using si/shRNA, CRISPR perturbations, and phospho-signaling assays in human T cells, together with eKras—/— mice, mixed bone marrow chimeras, influenza infection, and SARS-CoV-2 mRNA vaccination with downstream cellular and serologic analyses.
We catalogued and characterized ∼2,000 eRNAs in human adaptive immune cells; eKRAS was among the most highly expressed and conserved and functioned as a cis-acting enhancer of KRAS. Disruption of eKRAS reduced KRAS mRNA and attenuated RAS-ERK activation. Although eKras—/— mice developed normally, immunized mixed chimeras revealed a cell-intrinsic defect in T follicular helper (Tfh) differentiation, with impaired germinal center formation, reduced Tfh effector programs, and defective neutralizing antibody responses to protein antigens and influenza. Following SARS-CoV-2 mRNA vaccination, eKras—/— mice showed reduced class-switched anti-spike antibodies. An eKRAS-dependent Tfh transcriptional program was conserved in human blood and associated with neutralizing antibody titers after COVID-19 vaccination.
We define a systematic catalog of eRNAs in human adaptive immune cells and identify a distal transcribed enhancer that fine-tunes KRAS signaling in Tfh cells to support effective vaccine responses. These findings establish eRNAs as noncoding regulators of T cell circuits controlling antibody production and nominate the eKRAS-KRAS axis as a target to optimize humoral immunity.
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Immune Response Regulation: Molecular Mechanisms (IRM)
Dhaneshwar Kumar, S. Sahoo, B. Yan et al.· Journal of Immunology· 0 citations
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