The Arabidopsis genome contains many gene families that are not found in the animal kingdom. One of these is the multidrug and toxic compound extrusion (MATE) family, which has homology with bacterial efflux transporters. Arabidopsis has at least 54 members of this family, which often are found in tandem repeats. Analysis of ALF5, one member of this Arabidopsis family, suggests that its function is required for protection of the roots from inhibitory compounds. Loss of ALF5 function results in the sensitivity of the root to a number of compounds, including a contaminant of commercial agar. Moreover, expression of the Arabidopsis ALF5 cDNA in yeast confers resistance to tetramethylammonium. These phenotypes are consistent with a role for ALF5 as an efflux transporter. Both transcriptional and translational fusions of ALF5 to the beta-glucuronidase reporter gene show that ALF5 is expressed strongly in the root epidermis, a tissue in direct contact with the external environment. The distinct requirement for ALF5 function is remarkable because of the large number of MATE gene family members in Arabidopsis, one of which is adjacent to ALF5 and 83% identical to ALF5 at the amino acid level.