Novel functional proteins interact with midkine in hepatic cancer cells.
Midkine is a heparin-binding growth factor that promotes the proliferation, survival, migration and differentiation of various target cells. Midkine plays an important role in tumorigenesis and tumor progression, and is overexpressed in many human malignant tumors. Patients with high tumor midkine expression frequently have a worse prognosis than those with low expression. The present study was designed to investigate the interaction network of midkine in hepatic cancer cells, and to elucidate its role in hepatocellular carcinoma.
DNA encoding full-length midkine was cloned into pDBLeu vector to serve as bait in yeast two-hybrid screening to identify interacting proteins. Candidate proteins were examined on SC-Leu-Trp-His+3-AT (20 mmol/L) plates and assayed for X-gal activity, then sequenced and classified according to the GenBank. Finally, identified proteins were expressed by the in vitro expression system pCMVTnT, and protein interactions were confirmed by co-immunoprecipitation.
Using the yeast two-hybrid system, we found 6 proteins that interacted with midkine: NK-kappa-B inhibitor alpha (I-κ-B-alpha), Dvl-binding protein naked cuticle 2, granulin, latent active TGF-beta binding protein 3, latent active TGF-beta binding protein 4, and phospholipid scramblase 1. In vitro co-immunoprecipitation demonstrated that all identified proteins directly interacted with midkine.
The identification of midkine-interacting proteins in hepatic cancer cells indicates that midkine is a multifunctional factor that may participate in cell migration, differentiation, and proliferation, and is also associated with the multicellular response feedback during the development of hepatocellular carcinoma.
Zhejiang University School of Medicine, Hangzhou, China.
SourceHepatobiliary & pancreatic diseases international : HBPD INT 11:3 2012 Jun pg 272-7
I-kappa B Proteins
Intercellular Signaling Peptides and Proteins
Latent TGF-beta Binding Proteins
Nerve Growth Factors
Phospholipid Transfer Proteins
Two-Hybrid System Techniques
Pub Type(s)Journal Article
Research Support, Non-U.S. Gov't