I-Ak- and I-Ed-transfected L fibroblasts were supertransfected with cDNA coding for hen egg lysozyme (HEL) or measles virus hemagglutinin (HA). Well-defined cell culture conditions allowed us to obtain L cells with either no detectable endogenous Ii mRNA or a high level of endogenous Ii mRNA induced by serum starvation. Cells supertransfected with mouse Ii chain gene stably expressing a high level of Ii chain were also used as APC in parallel experiments. Class II presentation of endogenously secreted HEL or an ER-retained form of HEL to the HEL-specific I-Ak-restricted 3A9 T cell hybridoma was found to be strongly enhanced in cell transfectants expressing Ii chain. Similar results were obtained with the presentation of transmembrane HA to the HA-specific I-Ed-restricted TH5.143 T cell hybridoma. These findings correlate with those obtained with the presentation of exogenous HEL and HA. In addition, as reported to be the case for exogenous antigen, expression of a large amount of endogenous HA by the APC supplants the requirement for Ii chain expression in the enhancement of antigen presentation. These data demonstrate that presentation by MHC class II molecules of a given antigen, either exogenously provided or endogenously synthesized, is controlled in a similar manner by the Ii chain.