Identification of PlexinD1 and AHDC1 as a putative interactors for Tip-1 protein

The class 1 PDZ domain Tip-1 protein was first identified as a binding partner for the Human T-Cell Leukaemia Virus type 1 (HTLV-1) Tax oncoprotein. It was later shown to interact with: the RhoA signaling effector Rhotekin, the Wnt signaling effector beta-catenin and the E6 oncoprotein from high-risk HPV16 but not low-risk HPV6. These observations suggested that Tip-1 may be an important hub protein that is involved in pathways with a proven link to carcinogenesis. Based on these findings, it was decided to further characterize the cellular role of Tip-1 by carrying out a yeast two-hybrid screen to identify new binding partners in order to uncover potentially novel functions of this protein. This identified an intracellular fragment of the trans-membrane receptor plexin D1 and a C-terminal fragment of the AT hook DNA binding containing 1 (AHDC1) protein which had a carboxyl terminal PDZ binding domain consensus sequence. Both of these interactions were confirmed by yeast mating assay which was also used to show that mutant constructs of AHDC1 lacking the carboxyl PDZ binding site did not bind Tip-1. Immunofluorescent imaging of these proteins in HPV16 E6 expressing human C33A cervical carcinoma cells suggested they may co-localize.