Abstract Despite the significant improvements in short-term survival of organ transplants, rejection remains a leading cause of long-term transplant loss. Innate immune activation potentiates the adaptive immunity and is a crucial player in precipitating acute rejection and preventing transplant tolerance. However, most immunosuppressive drugs used in the clinic primarily target T cells and not innate immune cells. Therefore, there is an unmet need to develop effective therapies to modulate innate immunity in transplantation. Siglec (sialic acid-binding immunoglobulin-like lectin)-E, or SigE, is an innate receptor that down-modulates inflammation. SigE is expressed by dendritic cells (DCs) and inhibits TLRs-triggered inflammatory responses. Engagement of SigE is a promising strategy to promote immune regulation. However, the role of SigE in transplantation has not been investigated. We have found that allografts lacking SigE have an accelerated rejection, and kidney transplant patients have decreased expression of human SigE counterpart (Siglec-9) during rejection. Moreover, we have recently discovered that a mycobacterial protein DnaK can bind to SigE with potent immunomodulatory effects in alloimmunity by decreasing DCs activation through downregulation of MHC II and CD86. In situ targeting of donor DCs with DnaK is capable of prolonging skin allograft survival in the absence of systemic immunosuppression and DnaK-effect was dependent on SigE. Thus, targeting SigE represents a novel potential therapeutic target to enhance the modulation of the immune response in transplantation. Based on our preliminary data, we hypothesize that SigE is a critical regulator of the immune response following transplantation. To address this hypothesis, we propose three specific aims: 1) To define the role of SigE in regulating DC maturation and antigen processing; 2) To determine the role of SigE in alloimmunity and in the generation of antigen-specific T cells; and 3) To investigate a novel immunomodulatory molecule designed based on the interaction between Siglec-9 and DnaK. To accomplish these aims, we will utilize heart and humanized mouse transplantation models, a novel synthetic agonist peptide to SigE, SigE-deficient mice, tracking of antigen-specific T cells using adoptive transferred cells and endogenous staining using tetramers. The proposed studies are innovative and significant because we will explore the biology of an important regulatory innate immune receptor, SigE, in transplantation and we will investigate a novel SigE targeting molecule to inhibit alloimmunity,