Histidine Triad Nucleotide-binding Protein 1 Up-regulates Cellular Levels of p27KIP1 by Targeting ScfSKP2 Ubiquitin Ligase and Src

The one or more underlying mechanisms of the tumor suppressing activity of the histidine triad nucleotide-binding protein 1 (HINT1) are not well defined. In this study we found that HINT1 regulates cellular levels of the cyclin-dependent kinase inhibitor p27(KIP1) through multiple mechanisms. Increased expression of HINT1 increases cellular levels of p27(KIP1), and HINT1 knockdown with small hairpin RNA leads to decreased cellular levels of p27(KIP1). HINT1 does not affect the transcription of p27(KIP1), but it does inhibit proteasomal degradation of the p27(KIP1) protein. HINT1 directly interacts with the SCFSKP2 ubiquitin ligase complex and inhibits the ubiquitylation of p27(KIP1). Src has been shown to phosphorylate p27(KIP1) and thus decrease its stability. We found that HINT1 is a negative regulator of Src transcription apparently by forming a complex with the transcription factor Sp1 on the promoter of Src. Taken together, our findings indicate that HINT1 up-regulates cellular levels of p27(KIP1) by two mechanisms: 1) it inhibits its ubiquitylation by targeting the SCFSKP2 ubiquitin ligase complex, and 2) it inhibits the phosphorylation of p27(KIP1) by Src via inhibiting Src expression.