Objective      To screen and identify the proteins that can interact with human C-terminal binding protein 2 (CtBP2).      Methods       Yeast two-hybrid screening technique

with CtBP2 as bait protein

was used to screen out the genes of the candidate CtBP2-interacting proteins from human liver cDNA library. Then

yeast co-transformation assay and co-immunoprecipitation assay were performed to confirm the specificity of interaction. Fluorescent fusion proteins and a fluorescence microscope were used for subcellular co-localization of CtBP2 and its interacting proteins in HeLa cells.      Results      The yeast two-hybrid screening method yielded 41 positive clones that coded 10 CtBP2-interacting proteins. Among them

4 clones coded different fragments of ubiquitin-conjugating enzyme E2Ⅰ (UBE2Ⅰ). The full-length coding sequence of UBE2Ⅰ was amplified from human liver cDNA library. The specific interaction between CtBP2 and UBE2Ⅰ was confirmed by the yeast co-transformation assay and co-immunoprecipitation assay. The subcellular localization results showed that: (1) GFP-CtBP2 was dispersed in the nuclei of HeLa cells

(2) DsRed-UBE2Ⅰ was unevenly distributed in the nuclei of HeLa cells with some spotty accumulation

and (3) GFP-CtBP2 and DsRed-UBE2Ⅰ could be co-localized in nuclear speckles when the HeLa cells were transfected with both pEGFP-CtBP2 and pDsRed-UBE2Ⅰ.      Conclusion       Human CtBP2 can interact with UBE2Ⅰ specifically in the nucleus and is a target protein of UBE2Ⅰ.