摘要
We previously reported that v-Fos transformation effector (Fte-1) is a novel CD2 binding protein identified by yeast two hybrid system. In the present study, we further characterize the molecular properties and biological activity of the Fte-1. In vitro interaction analysis by an IAsys Resonant Mirror Bio- sensor further demonstrated that Fte-1 interacts with CD2 specifically, and the dissociation constant (Kd) of Fte-1-CD2 is 10?7 mol. The molecular section analysis showed that protein kinase C (PKC) phos- phorylation site at Ser238 of Fte-1, as confirmed by in vitro phosphorylation, is essential for the specific in- teraction with CD2. Jurkat T cells transfected with expression vector encoding for EGFP-Fte-1 fusion protein showed that Fte-1 displays a clustering dis- tribution in the cells. Upon stimulation by CD2 mono- clonal antibody T11, Fte-1 lost the character of clus- tering distribution and translocation to the plasma membrane, which were disrupted by mutation of Fte-1 (Ser238Gly), suggesting that Fte-1 could be co-localized with CD2 on the membrane, and Fte-1 phosphorylation at Ser238 is a crucial factor in CD2-mediated interaction with Fte-1. Suppression of Fte-1 expression by small interference RNA (siRNA) diminished the susceptibility of Jurkat T cells to apoptosis triggered by phorbol 12-myristate 13-ace- tate (PMA) and ionomycin, indicating that the bio- logical function of Fte-1 may be involved in the regu- lation of activation and apoptosis of T cells. These results provide further evidence that Fte-1 is a novel binding protein of CD2, and may function as a down- stream molecule in the CD2-mediated events.
We previously reported that v-Fos transformation effector （Fte-1） is a novel CD2 binding protein identified by yeast two hybrid system. In the present study, we further characterize the molecular properties and biological activity of the Fte-1. In vitro interaction analysis by an IAsys Resonant Mirror Bio- sensor further demonstrated that Fte-1 interacts with CD2 specifically, and the dissociation constant （Kd） of Fte-1-CD2 is 10-7 mol. The molecular section analysis showed that protein kinase C （PKC） phosphorylation site at Ser238 of Fte-1, as confirmed by in vitro phosphorylation, is essential for the specific interaction with CD2. Jurkat T cells transfected with expression vector encoding for EGFP-Fte-1 fusion protein showed that Fte-1 displays a clustering distribution in the cells. Upon stimulation by CD2 monoclonal antibody T11, Fte-1 lost the character of clustering distribution and translocation to the plasma membrane, which were disrupted by mutation of Fte-1 （Ser238Gly）, suggesting that Fte-1 could be co-localized with CD2 on the membrane, and Fte-1 phosphorylation at Ser238 is a crucial factor in CD2-mediated interaction with Fte-1. Suppression of Fte-1 expression by small interference RNA （siRNA） diminished the susceptibility of Jurkat T cells to apoptosis triggered by phorbol 12-myristate 13-acetate （PMA） and ionomycin, indicating that the biological function of Fte-1 may be involved in the regulation of activation and apoptosis of T cells. These results provide further evidence that Fte-1 is a novel binding protein of CD2, and may function as a downstream molecule in the CD2-mediated events.
基金
partially supported by State Key Basic Research Program of China(No.G2000057002)
the National Natural Science Foundation of China(Grants Nos.30271207&30210103903).
