Eukaryotic initiation factor 3 (eIF3) is a multisubunit complex that is required for binding of mRNA to 40 S ribosomal subunits, stabilization of ternary complex binding to 40 S subunits, and dissociation of 40 and 60 S subunits. These functions and the complex nature of eIF3 suggest multiple interactions with many components of the translational machinery. Recently, the subunits of mammalian and Saccharomyces cerevisiae eIF3 were identified, and substantial differences in the subunit composition of mammalian and S. cerevisiae were observed. Mammalian eIF3 consists of 11 nonidentical subunits, whereas S. cerevisiae eIF3 consists of up to eight nonidentical subunits. Only five of the subunits of mammalian and S. cerevisiae are shared in common, and these five subunits comprise a "core" complex in S. cerevisiae. eIF3 from wheat consists of at least 10 subunits, but their relationship to either the mammalian or S. cerevisiae eIF3 subunits is unknown. Peptide sequences derived from purified wheat eIF3 subunits were used to correlate each subunit with mammalian and/or S. cerevisiae subunits. The peptide sequences were also used to identify Arabidopsis thaliana cDNAs for each of the eIF3 subunits. We report seven new cDNAs for A. thaliana eIF3 subunits. A. thaliana eIF3 was purified and characterized to confirm that the subunit composition and activity of wheat and A. thaliana eIF3 were similar. We report that plant eIF3 closely resembles the subunit composition of mammalian eIF3, having 10 out of 11 subunits in common. Further, we find a novel subunit in the plant eIF3 complex not present in either mammalian or S. cerevisiae eIF3. These results suggest that plant and mammalian eIF3 evolved similarly, whereas S. cerevisiae has diverged.