FIG. 1.

An epitope tag can be added to the C terminus of MT without disrupting transformation. (A) Amino acid sequences of the MT C terminus and three mutants tagged with the HA epitope. The amino acid sequence of wild-type MT is shown at the top, together with the sequence of three mutants where the HA epitope (shown on the right) is attached to MT. The designation of each MT species is listed on the left. The hydrophobic region of MT is identified with a space on either side, and the HA tag is also marked by a space between it and MT. (B) Transformation assays using the mutants shown in panel A. Plasmid DNA encoding each of the mutants shown in panel A was transfected into Rat2 fibroblasts and stained for focus formation after 14 days. The MT species used is indicated above or below each image. (C) MT polypeptides expressed by the mutants shown in panel A. Cell lines expressing each of the mutant MTs shown in panel A were isolated, and lysates were prepared. An equal volume of cell lysate from each line was separated by SDS-gel electrophoresis, Western blotted, and then probed with anti-MT monoclonal antibody PAb762. The migration position of MT is shown on the right, and the mutant used is indicated above each lane.