Modulation of nuclear receptor-binding factor 2 on the activity of autophagy initiation complex PI3KC3-C1 / 上海交通大学学报（医学版）

ABSTRACT

Objective • To study the effects of different phosphorylation levels of nuclear receptor-binding factor 2 (NRBF2) on the activity of class III phosphatidylinositol 3-kinase complex (PI3KC3-C1), and find a type of PI3KC3-C1 with the highest kinase activity as a target for the screening of autophagy-specific inhibitors. Methods • First, in vitro protein purification system was established to obtain quaternary NRBF2-free PI3KC3-C1 and quinary PI3KC3-C1 containing different phosphorylation levels of NRBF2. The integrity of the complex was determined by gel filtration chromatography. The different kinase activity of purified PI3KC3-C1 was detected in vitro, and thus the effect of phosphorylation of NRBF2 on PI3KC3-C1 activity was observed. Results • The purified PI3KC3-C1 had good purity and yield. The five-membered PI3KC3-C1 containing NRBF2 showed higher kinase activity than the quaternary protein complex, and PI3KC3-C1 containing the continuous dephosphorylated NBRF2 had the highest kinase activity. Conclusion • NRBF2 does exist as one of the components of PI3KC3-C1. The presence of NRBF2 promotes the kinase activity of PI3KC3-C1, and PI3KC3-C1 containing continuous dephosphorylation of NRBF2 may serve as a new regulatory target for the screening of autophagy-specific inhibitor.