Surface TLR 3 expression in metastatic IECs 15410

Toll-like receptors (TLRs) are innate immune receptors for sensing microbial molecules and damage-associated molecular patterns released from host cells. Double-stranded RNA and the synthetic analog polyinosinic:polycytidylic acid (poly(I:C)) bind and activate TLR3. This stimulation leads to recruitment of the adaptor molecule TRIF (Toll/IL-1 resistance (TIR) domain– containing adapter-inducing interferon ) and activation of the transcription factors nuclear factor B (NFB) and interferon regulatory factor 3 (IRF-3), classically inducing IFN production. Here we report that, unlike non-metastatic intestinal epithelial cells (IECs), metastatic IECs express TLR3 and that TLR3 promotes invasiveness of these cells. In response to poly(I:C) addition, the metastatic IECs also induced the chemokine CXCL10 in a TLR3-, TRIF-, and IRF3-dependent manner but failed to produce IFN . This was in contrast to healthy and nonmetastatic IECs, which did not respond to poly(I:C) stimulation. Endolysosomal acidification and the endosomal transporter protein UNC93B1 was required for poly(I:C)-induced CXCL10 production. However, TLR3-induced CXCL10 was triggered by immobilized poly(I:C), was only modestly affected by inhibition of endocytosis, and could be blocked with an anti-TLR3 antibody, indicating that TLR3 can still signal from the cell surface of these cells. Furthermore, plasma membrane fractions from metastatic IECs contained both full-length and cleaved TLR3, demonstrating surface expression of both forms of TLR3. Our results imply that metastatic IECs express surface TLR3, allowing it to sense extracellular stimuli that trigger chemokine responses and promote invasiveness in these cells. We conclude that altered TLR3 expression and localization may have implications for cancer progression. Toll-like receptors (TLRs), Nod-like receptors (NLRs), and retinoic acid–inducible gene 1 (RIG-I)–like receptors (RLRs) are innate signaling pattern recognition receptors (PRRs) that are important for mounting an inflammatory response against invading microorganisms (1). PRRs recognize conserved microbe-associated molecular patterns expressed by microorganisms, but also sense damage-associated molecular patterns released from damaged host cells during injury (2). Activation of PRRs results in the production of inflammatory cytokines, type I IFNs, and initiation of innate and adaptive immune responses. Toll-like receptor 3 (TLR3) recognizes doublestranded RNA (dsRNA), a viral replication intermediate associated with virus infection. It also recognizes the synthetic dsRNA mimic polyinosinic:polycytidylic acid (poly(I:C)) and RNA released from necrotic cells as ligands (3, 4). Viral dsRNA and poly(I:C) are also recognized by TLR-independent pathways and trigger activation of the cytoplasmic RNA helicases RIG-I and Mda-5 in the cytosol (5–7). PRR activation in cancer cells may contribute to the progression of cancer. Certain cancer cells up-regulate PRR expression and produce damage-associated molecular patterns that activate other cells in the tumor microenvironment, leading to further cytokine release and tumor-promoting inflammation (8). TLR3 activation induces inflammatory cytokines but also induces type I IFN production, which impairs proliferation and induces apoptosis in some cancer cells (9 –11). IFNs additionally activate natural killer cells, enhance CD8 T cell responses, and promote DC maturation (12). The ability to trigger type I IFNs has rendered TLR3 an attractive target in cancer therapy (12), but TLR3 has also been acknowledged as a mediator of pro-tumorigenic inflammation, which can drive cancer cell