Toll-like receptor (TLR) signalling pathways constitute an evolutionarily conserved component of the host immune response to pathogenic infection. Here, we describe the ability of a virally encoded form of the Pellino protein to inhibit Toll- and TLR-mediated activation of downstream Rel family transcription factors. In addition to inhibiting drosomycin promoter activation by Spa¨ tzle in Drosophila melanogaster cells, viral Pellino attenuates the activation of NF-jB by TLR signalling components and by the TLR4 ligand, LPS, in human cells. We propose that viral Pellino, like mammalian Pellinos, contains a forkhead-associated domain but differs from the mammalian forms in that it lacks a complete and functional RING-like domain. We produce a homology model and present experimental data to support this model by demonstrating that, like mammalian Pellinos, viral Pellino can interact with IRAK-1 via its forkhead-associated domain, whereas unlike its mammalian counterparts, it fails to post-translationally modify IRAK-1. Furthermore, we demonstrate that viral Pellino can functionally antagonise the activity of human Pellino3S. Thus, our findings identify potential immunoevasive capabilities possessed by a poxviral homolog of the Pellino protein and add growing evidence for a likely role for Pellino proteins in Toll and TLR signalling.