The present paper describes cloning and sequencing of the mouse cDNA encoding dihydroxyacetonephosphate acyltransferase (DAPAT), the peroxisomal key enzyme of plasmalogen (PM) biosynthesis. Using monospecific antibodies, we localized DAPAT and alkyl dihydroxyacetonephosphate synthase to peroxisomes of mouse lens epithelial cells (LECs) and determined their enzymatic activity. By electrospray ionization mass spectrometry of mouse lens lipid extracts, we identified phosphatidyl ethanolamine including plasmenyl ethanolamine species as major constituents. Our data demonstrate the capacity of LECs to synthesize PMs and the high coincidence between deficiency of PM and early manifestation of cataract in patients with peroxisomal disorders suggests that ether-bonded lipids may play an important role in maintaining lens transparency.