Recognition of the microbial world is mediated chiefly by a small group of immune receptors that activate a characteristic host inflammatory response, the innate immune response. Known as the Toll-like receptors (TLRs), these molecules are represented among most metazoans. In mammals, forward genetic analysis of the lipopolysaccharide (LPS) response led to the identification of TLR4 as the LPS receptor. Through a combination of forward and reverse genetic studies, a relatively detailed understanding of the functions of mammalian TLRs has now been achieved. As discussed here, mutagenesis has revealed proteins that participate in TLR signaling pathways, and informed our understanding of the subtleties of these molecules' structure and function.