NTE is essential, but not sufficient for protein-protein interactions of HMT-1.

A. Full-length HMT-1 was fused at the C-terminus with CubPLV (HMT-Cub) or with NubG (HMT-NubG). HMT-1 lacking NTE was fused at the C terminus with NubG (ΔNTE-NubG). B. SDS-PAGE and immunoblot analysis of microsomal membranes prepared from THY.AP5 cells expressing the NubG-fused full-length HMT-1 (HMT-NubG), NubG-fused HMT-1 lacking NTE (ΔNTE-NubG), NTE-fused to NubG (NTE-NubG), or empty NubG vector (NubG). C. Interactions were manifested by the ability of cells to grow on SC media without adenine and histidine (SC), but not in SC media with methionine (SC + Met). Minimal growth relative to the negative controls (co-expression with vector alone, NubG) was observed when HMT-1-NubG prey construct lacking NTE (ΔNTE-NubG) was co-expressed with the full-length HMT-1-CubPLV bait (HMT-Cub). Diploid cells did not grow when the NTE domain fused to NubG (NTE-NubG) was co-expressed with HMT-1-CubPLV bait (HMT-Cub). Interactions were also visualized by the presence of β-galactosidase activity. Note the presence of β-galactosidase activity in cells co-expressing the full-length HMT-1-NubG and HMT-1-CubPLV, but not the full-length HMT-1-CubPLV with ΔNTE-NubG, or NTE-NubG or NubG only.