Efficient uptake and channeling of long-chain fatty acids (LCFAs) are critical cellular functions. Although spontaneous flip-flop of nonionized LCFAs from one leaflet of a bilayer to the other is rapid, evidence is emerging that proteins are important mediators and/or regulators of trafficking of LCFAs into and within cells. Genetic screens have led to the identification of proteins that are required for fatty acid import and utilization in prokaryotic organisms. In addition, functional screens have elucidated proteins that facilitate fatty acid import into mammalian cells. Although the mechanisms by which these proteins mediate LCFA import are not well understood, studies in both prokaryotic and eukaryotic organisms provide compelling evidence that uptake of LCFAs across cellular membranes is coupled to esterification by acyl-CoA synthetases. This review will summarize results of studies of non-protein-mediated and protein-mediated LCFA transport and discuss how these different mechanisms may contribute to cellular metabolism.