FIGURE 1.

Identification of IQGAP1 as a pendrin binding partner. (A) The C-terminus of pendrin was used as bait in Y2H in order to identify the protein(s) that interact with pendrin. The C-terminus of mouse pendrin protein spanning amino acids 508–780 was used in Y2H studies in order to identify the proteins that interact with pendrin. This region was used because it contains the STAS domain. The missense mutations (highlighted, bolded and underlined) correspond to those of that affect the function of the human pendrin protein and are associated with Pendred Syndrome and DFNB4. (B) Our studies identified a total of 33 interactions between pendrin and prey sequences. Of these only 22 interactions were shown to be specific while others we excluded due to out of frame translation, reversed direction of the cloned sequence, or low affinity of the bait/prey interaction. The interaction with IQGAP1 represented 14 of the 22 significant interactions. Based on the Global PBS score, IQGAP/pendrin interaction had a very high degree of confidence. The Global PBS score is automatically computed through algorithms with the following scoring system: A) Very high confidence in the interaction; B) High confidence in the interaction; C) Good confidence in the interaction; D) Moderate confidence in the interaction; E) Interactions highly connected prey domains, warning of non-specific interaction; and F) Experimentally proven technical artifacts. In addition to IQGAP1, the Y2H system also identified CD97 as a protein with a “high confidence of interaction” giving it a Global PBS score of B. The relationship between pendrin and CD97 as a binding partner needs further examination. The % Id 5p/3p indicates the degree of identity to the putative binding protein.