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The lymphatic system actively regulates tissue immunity through LECs in peripheral tissues, tumors, and draining LNs. LECs regulate leukocyte trafficking and T cell responses and known to influence antitumor immunity. Transcriptomic analyses identified Sema3a as an inflammation-induced, LEC-enriched gene in murine and human tissues. We investigated whether LEC-derived Sema3a suppresses antitumor immunity by limiting CD8⁺ T cell priming, promoting regulatory T cell trafficking, and establishing an immunosuppressive microenvironment. We also assessed its clinical relevance and therapeutic targetability.

Sema3a expression was analyzed in murine and human tissues using RNA-sequencing datasets, quantitative PCR, reporter mice, and immunofluorescence. Inducible Prox1-CreERT2 Sema3afl/fl mice were used to investigate the effects of LEC-derived Sema3a on T cell activation in contact hypersensitivity and immunization studies, supported by in vitro LEC-T cell co-culture assays. The role of LEC-derived Sema3a in immune-cell migration was assessed using in vitro migration assays and photoconvertible reporter mice. Antitumor effects were evaluated in the MC38 colorectal cancer and YUMMER1.7 melanoma models, including treatment with a Sema3a-neutralizing antibody. Patient melanoma and tumor-draining LN biopsies were analyzed by immunofluorescence.

LECs were a major source of Sema3a in murine and human LN and peripheral tissues, and its expression increased during inflammation. LEC-specific deletion reduced tissue Sema3a without altering lymphatic architecture, drainage, or steady-state LN composition. Loss of LEC-derived Sema3a enhanced inflammatory and antigen-specific CD8⁺ T cell responses, increased IFN-γ production, and reduced regulatory T cell migration to draining LNs. In both tumor models, deletion of LEC-derived Sema3a delayed growth and enhanced intratumoral CD8⁺ T cell abundance or function. Systemic Sema3a blockade also reduced MC38 tumor growth. Sema3a protein localized to lymphatic vessels in melanoma and tumor-draining LN biopsies.

LEC-derived Sema3a acts as a lymphatic stromal checkpoint linking regulatory T cell trafficking with suppression of CD8⁺ T cell priming. Its inhibition strengthens antitumor immunity and limits tumor growth, identifying Sema3a as a clinically relevant and therapeutically targetable regulator of the cancer-immunity cycle.

Cell-autonomous immunity provides a critical first line of defense against viral infection, but how antiviral restriction factors are wired in a cell‑ and tissue‑specific manner remains incompletely defined. The vascular endothelium forms a key barrier limiting systemic viral dissemination, and its breach is closely linked to severe complications across diverse infections. Despite this central role, endothelial‑intrinsic antiviral mechanisms have only recently begun to be delineated.

Here, we combined mouse and human tissue vessel imaging, in vitro and ex vivo multi‑virus infection assays, proximity ligation screening and endolysosomal trafficking assays.

We found that IFITM3 is constitutively expressed at the protein level in human and mouse endothelial cells across multiple vascular beds, in contrast to non‑endothelial tissues where IFITM3 is primarily interferon‑induced, highlighting the endothelium as a uniquely pre‑primed antiviral compartment. Functionally, loss of IFITM3 in endothelial cells increases infection across multiple viral models, establishing IFITM3 as a key component of endothelial antiviral restriction. Strikingly, depletion of MYCT1 – a pan-endothelial interactor of IFITM3 – has the opposite effect, enhancing antiviral restriction despite its direct interaction with IFITM3. Mechanistically, MYCT1 loss reduces IFITM3 degradation and drives its accumulation in early endosomes, which limits viral escape from the endosomal compartment and promotes lysosomal destruction of incoming virions. These data identify MYCT1-IFITM3 as a cell‑type‑specific rheostat, preventing excessive antiviral restriction at the vascular barrier under homeostatic conditions while allowing rapid amplification upon infection. Our ongoing work aims to define the molecular determinants of the MYCT1–IFITM3 complex and to identify upstream regulators that tune this checkpoint.

Together, these findings establish the vascular endothelium as a distinct compartment endowed with a constitutively primed yet tightly controlled antiviral program and reveal a MYCT1–IFITM3 complex that shapes virus–host interactions at the vascular interface. More broadly, this work advances our understanding of tissue‑specific antiviral immunity and uncovers regulatory mechanisms that may be exploited by pathogens or targeted to fine‑tune endothelial barrier protection in infection and inflammation.