KAR9 and BIM1 interact with SUMO (SMT3) by two-hybrid analysis, but not with ubiquitin (UBI4) and another ubiquitin-related protein (URM1). Diploid two-hybrid reporter strains were generated by crossing yRM1757/PJ69-4A containing KAR9-BD (pRM1493), BIM1-BD (pRM3145), KIP2-BD (pRM3595), STU2-BD (pRM1916), or empty BD (pRM1154) with yRM1756/PJ69-4α containing AD-UBI4 (pRM5880), AD-URM1 (pRM5829), AD-SMT3 (pRM4920), or empty AD (pRM1151). Diploids were selected on SD −ura −leu media and tested for interaction by growth on SD −his media at 30° for 2–3 days. Two independent diploid colonies were tested. These results indicate that although UBI4, URM1, and SMT3 encode related molecules, only SMT3 interacts with KAR9 and BIM1.