This review highlights the emerging roles of the Pellino family of E3 ubiquitin ligases as upstream mediators in Toll-like receptor (TLR) pathways that lead to activation of MAP kinases and transcription factors. The functional importance of the Pellino family as RING-like-domain-containing proteins with intrinsic ubiquitin E3 ligase activity that can catalyse polyubiquitylation of the key TLR signalling molecule IRAK1 is discussed in detail. The importance of Pellino proteins as novel targets for mediating negative regulation of TLR signalling is also explored. This new knowledge and understanding of Pellino biology begins to fill some long-standing voids in our understanding of TLR signalling.