Signalling by means of toll- like receptors ( TLRs) is essential for the development of innate and adaptive immune responses(1-3). UNC93B1, essential for signalling of TLR3, TLR7 and TLR9 in both humans and mice, physically interacts with these TLRs in the endoplasmic reticulum ( ER)(4-6). Here we show that the function of the polytopic membrane protein UNC93B1 is to deliver the nucleotide- sensing receptors TLR7 and TLR9 from the ER to endolysosomes. In dendritic cells of 3d mice, which express an UNC93B1 missense mutant ( H412R) incapable of TLR binding, neither TLR7 nor TLR9 exits the ER. Furthermore, the trafficking and signalling defects of the nucleotide- sensing TLRs in 3d dendritic cells are corrected by expression of wild- type UNC93B1. However, UNC93B1 is dispensable for ligand recognition and signal initiation by TLRs. To our knowledge, UNC93B1 is the first protein to be identified as a molecule specifically involved in trafficking of nucleotide- sensing TLRs. By inhibiting the interaction between UNC93B1 and TLRs it should be possible to achieve specific regulation of the nucleotide- sensing TLRs without compromising signalling via the cell- surface- disposed TLRs.