This review discusses the assembly and signaling functions of lipopolysaccharides (LPS) from Gram-negative bacteria, focusing on the recent advancements in understanding their structure and function. LPS consists of a hydrophobic domain called lipid A, a non-repeating "core" oligosaccharide, and a distal polysaccharide (O-antigen). Recent genomic data have enabled the study of LPS assembly in diverse Gram-negative bacteria, including human and plant pathogens, and the creation of mutants or hybrid constructs with novel properties. Key genes for lipid A biosynthesis have also been identified in higher plants, suggesting the existence of eukaryotic lipid A-like molecules. The carbohydrate diversity of LPS is better understood, but more research is needed to elucidate its functions. Sequence comparisons indicate extensive lateral transfer of genes for O-antigen assembly among bacteria. The most significant finding since 1990 has been the identification of the plasma membrane protein TLR4 as the lipid A signaling receptor in animal cells, which belongs to a family of innate immunity receptors. The expanding knowledge of TLR4 specificity and its downstream signaling pathways offers new opportunities for blocking the inflammatory side effects of sepsis. Future research will require insights into atomic-level recognition of LPS-protein interactions, better understanding of intra- and inter-cellular LPS trafficking, and the development of biological approaches combining bacterial and animal genetics.This review discusses the assembly and signaling functions of lipopolysaccharides (LPS) from Gram-negative bacteria, focusing on the recent advancements in understanding their structure and function. LPS consists of a hydrophobic domain called lipid A, a non-repeating "core" oligosaccharide, and a distal polysaccharide (O-antigen). Recent genomic data have enabled the study of LPS assembly in diverse Gram-negative bacteria, including human and plant pathogens, and the creation of mutants or hybrid constructs with novel properties. Key genes for lipid A biosynthesis have also been identified in higher plants, suggesting the existence of eukaryotic lipid A-like molecules. The carbohydrate diversity of LPS is better understood, but more research is needed to elucidate its functions. Sequence comparisons indicate extensive lateral transfer of genes for O-antigen assembly among bacteria. The most significant finding since 1990 has been the identification of the plasma membrane protein TLR4 as the lipid A signaling receptor in animal cells, which belongs to a family of innate immunity receptors. The expanding knowledge of TLR4 specificity and its downstream signaling pathways offers new opportunities for blocking the inflammatory side effects of sepsis. Future research will require insights into atomic-level recognition of LPS-protein interactions, better understanding of intra- and inter-cellular LPS trafficking, and the development of biological approaches combining bacterial and animal genetics.