Toll-like receptors (TLRs) are responsible for the detection of invading microbes. The transport of TLRs after ligand stimulation to endosomes is essential for their signalling function. The proteins reggie1 and 2 which form heterodimers and are also known as flotilin2 and 1, are markers of lipid raft fraction but it is unknown whether they contribute to raft formation or endosome internalization. We studied whether reggie2 is required for the agonist-induced trafficking of TLR4 and TLR3 in human umbilical vein endothelial cells (HUVECs). Knockdown of reggie2 by shRNA leads to a decreased internalization of TLR3 upon polyIC-rhodamine stimulation. Similarly, reggie2 shRNA attenuated the TLR4- and TLR3-but not TNFa-dependent induction of the adhesion molecules VCAM and ICAM-1 in response to agonist-stimulation. Importantly, overexpression of reggie2 in cells exposed to reggie2 shRNA restored TLR-dependent VCAM and ICAM-1 expression excluding non-specific effects of the shRNA. Reggie2 silencing also disrupted the TLR-TRIF-dependent signaling pathway as demonstrated by a loss of the phosphorylation of the TLR-downstream target interferon regulatory factor-3 (IRF3). Surprisingly, knockdown of reggie1 or combined knockdown of both, reggie1 and 2 was without effect on adhesion molecule expression. This may suggest that not the lack of reggie2 but an aberrant action of reggie1 in the absence of its binding partner reggie2 mediates the anti-inflammatory effect observed.

Conclusion: 

Down-regulation of reggie2 elicits an inhibitor effect on TLR signaling which is mediated by reggie1.