Growth Arrest-Specific 6-Mediated Efferocytosis Reveals Bidirectional Tolerogenic Crosstalk Between DCs and T Cells 2309074

Abstract

Abstract Introduction Efferocytosis is a key driver of immune tolerance, yet the mechanisms linking apoptotic cell clearance to tolerogenic antigen presentation remain unclear. Existing models suggest that tolerance relies on immunological silence through reduced antigen presentation on MHC. Recognizing the role of GAS6 in facilitating efferocytic antigen uptake, we engineered GAS6-OVA (GO), fusing bridge protein GAS6 to ovalbumin to dissect antigen-specific tolerance mechanisms in both APCs and T cells. Methods Variants of GO were expressed in HEK293F cells and purified by chromatography. BMDCs were antigen-pulsed ±LPS and co-cultured with OT-I or OT-II T cells. Flow cytometry assessed phenotypes. In vivo, adoptively transferred OT cells were tracked following GO/OVA treatment and CFA-OVA challenge. Wild-type mice were pretreated with GO or controls prior to induction of allergic airway inflammation. Results GO exhibited PS-specific binding and enhanced DC uptake. Instead of dampening presentation, GO-pulsed BMDCs upregulated MHC-I and -II, driving robust proliferation of OT-I and -II cells. CD8+ T cells displayed a phenotype resembling terminal exhaustion (PD-1+TIM-3+TOX+), while CD4+ T cells were enriched for Tregs displaying phenotypic signs of robustness (Helios+CD73+FR4hi). Crucially, the tolerogenic phenotype of DCs did not emerge immediately; DC expression of PD-L1 and TAM receptors increased only after engagement with antigen-specific T cells. These signatures of tolerance persisted under LPS stimulation. In vivo, GO reduced antigen-specific T cell recovery, enriched Tregs, and prevented allergic airway pathology including IgE and mucus hypersecretion. Conclusion GAS6-mediated efferocytosis drives bidirectional tolerogenic programming where APCs and T cells reciprocally reinforce tolerance. Enhanced, not reduced, antigen presentation accompanies tolerance-associated phenotypes. This self-reinforcing circuit may enable durable antigen-specific therapies for allergies, autoimmunity, and transplant rejection. Funding Source n/a Topic Categories Immune Response Regulation: Cellular Mechanisms (IRC)