Screening The Interaction Proteins Of POT1 And Setting-up Network Of Protein-protein Interaction

Purpose: We use POT1 as bait in our large scale yeast two-hybrid screen to reveal new potential interacting proteins, to construct a protein-portein interaction network centered with POT1, and to accumulate original data for the construction of human liver protein-protein interaction network.Method: We use POT1 as the first bait for yeast two-hybrid screen, then the potential interacting proteins identified, were used as the second baits in the next screen. Part of the interacting proteins with important functions from the second screen, were used as the third baits for a third screen. Large scale yeast two-hybrid screen methods were used later to screen against human liver cDNA library for more interacting proteins. Finaly we use the biological information method to analysis the data and make an interaction network picture with Osprey software.Results: We successfully constructed 20 bait expression vectors.Five new POT1 interact proteins, HSCK2B,LAMC3,SERTAD1,MVP and GEF,were screened. Then they were used as the second baits for futher screening.TGM2, JAB1, HIVEP1, AGPAT2, CKAP4, HRSP12, CDC5L, RPS6, ATF4, NF- B1, SHANK3, MCPH1, NR4A1, and GSDML1 were found.Finally,using the total 20 proteins as the third bait, we screen against adult human liver cDNA library. As a result, 236 interactions were found in our last screen. 107 interactions with higher reliability were selected with transformation confirmation and bioinformatics.All interactions are novel except five pairs which have reported. And these interactions form a complicated protein-protein interaction network. Conclusions: The 236 interactions found in our screen form a complicated protein-protein interaction network. New interaction pairs are helpful for study the function and regulation of a single protein; provide clues for new protein functions. This network provides new clues for study of POT1 function and provides original data for the human liver protein-protein interaction network project.