IL-2-STAT5 signaling is vital for CD4+ T cell function, whereas BACH2 restrains activation. How these transcription factors co-operate at the genome remains unclear. Intracellular C3 supports T cell survival and effector programming, but how it is transcriptionally controlled and whether it matches serum C3 is unknown. We hypothesized that a BACH2-STAT5 module coordinates C3 expression and licenses a distinct C3 proteoform in activated CD4+ T cells.
We profiled wild-type and Bach2—/— CD4+ T cells by RNA-seq and mapped STAT5 and BACH2 binding using ChIP-seq and H3K27ac Hi-ChIP. STAT5 and BACH2 were re-expressed in Stat5—/— T cells, and shared elements at the C3 locus were deleted by CRISPR. IL-2/STAT5 inhibition, C3 overexpression or knockdown, and complement activation assays defined C3 expression and function. AlphaFold2 modeling and domain analyses characterized novel C3 proteoforms.
IL-2/STAT5 target genes were enriched in Bach2—/— CD4+ T cells, with overlapping STAT5 and BACH2 binding at differentially expressed loci. Combined activating or repressive control was confirmed by re-expression in Stat5—/— cells. At the C3 locus, BACH2-STAT5 co-occupancy and IL-2/STAT5 signaling were required for C3 transcription; C3 was lost in Bach2—/— cells, after IL-2/STAT5 blockade, or upon CRISPR deletion of shared elements. Resting CD4+ T cells expressed canonical, signal peptide-containing C3, whereas activation induced a shorter, signal peptide-deficient transcript predicted to encode an intracellular proteoform. Purified short C3 failed to seed classical C3 convertase or support hemolysis, indicating altered function.
We identify a BACH2—STAT5 transcriptional module that is essential for C3 expression in activated CD4+ T cells and redirects C3 output toward a short, intracellular proteoform with noncanonical complement activity. Proteoform-specific, T cell-intrinsic complement suggests new opportunities to target BACH2-STAT5 and C3 in autoimmunity and immunotherapy.
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Immune Response Regulation: Molecular Mechanisms (IRM)
Jorge Trujillo, D. Chauss, Luopin Wang et al.· Journal of Immunology· 0 citations
Systemic complement protects the vascular compartment, while cell-intrinsic complement components shape tissue immunity by regulating normal cell physiology and metabolism. In human CD4 T helper cells, intracellular C5a generation engages the intracellular receptor C5aR1, promoting Th1 differentiation. Here, we identify a counter-regulatory role for the alternative C5a receptor, C5aR2, in restraining T cell effector responses by studying the first reported family with a heterozygous C5aR2 mutation. The affected individuals present with an autoinflammatory syndrome, and disease penetrance tracks with the mutation: the affected mother and child carry the variant, whereas the unaffected father and siblings do not.
To define the impact of the C5aR2 mutation, we performed single-cell RNA sequencing, in vitro stimulation assays, and flow cytometric profiling of patient and control PBMCs. CPM inhibition or deletion, C5aR2 agonism, and transcriptional profiling of Cpm- or C5ar2-deficient mouse CD4 T cells, along with a T cell transfer colitis model, were used to define mechanistic consequences.
Patient samples exhibited a profound loss of naïve and central memory CD4 and CD8 T cells, accompanied by expansion of IFN-γ—producing effector memory populations. The C5aR2 mutation abolishes C5aR2 β-arrestin signaling. Additionally, we identified carboxypeptidase M (CPM) as a T cell—intrinsic enzyme generating C5a-desArg, a potent ligand for C5aR2. Loss or inhibition of CPM heightened inflammatory T cell responses, which were normalized by C5aR2 agonism. Mouse Cpm- or C5ar2-deficient CD4 T cells displayed overlapping transcriptional perturbations, and Cpm-knockout CD4 T cells induced more severe colitis.
Collectively, our data uncover a previously unrecognized CPM-dependent mechanism that balances C5aR1 and C5aR2 signaling to limit pathological T cell activation, revealing an intrinsic complement-driven checkpoint that constrains effector T cell immunity.
NHLBI/NIH intramural program
Immune Mechanisms of Human Disease (HUM)
Erin E. West, Nicolas S. Merle, Ayden Case et al.· Journal of Immunology· 0 citations
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