The Transcriptional Co-activator Protein P100 Regulate The Transcriptional Activity Of STAT1 And STAT6

Objective:The mechanism of signal transduction leading to transcription is to activate transcriptional factors,which makes corresponding gene expression and change the biological responses.JAK-STAT is one of these signal pathway.It is important to lead to proliferation,differentiation,apoptosis,with other transcriptional activated factors.The p100 was first identified as a homologous protein acting as a co-activator of EBNA..Studies indicate that p100 participated in the gene transcriptional regulation of STAT6 as a Co-activator,and increase its transcriptional activity.STAT6 and STAT1 are the members of STATs,p100 also can increase the transcriptional activity of STAT1.Furthermore,we observe the intracellular localization of STAT6 and p100,which provide evidence for the function of p100.Methods:The experiment is divided into three parts:the first part is to study the interaction of p100 and STAT1 or STAT6,this part is divided into two terms:the interacton between p100 and STAT1 or STAT6 in vitro by GST pull down assay;the interaction between them in vivo by CO-IP.the second part is to study p100 regulate the transcriptional activation of STAT1 or STAT6 with luciferase assay.observe.the third part is to observe the intracellular localization of p100 and STAT6.At the all,to construct pEGFP-CI-STAT6 and pERFP-CI-STAT6,which can express GFP or RFP fusion protein.Then transfect them to HeLa cell,with stimulation,to observe the change localization of fusion protein.Result:â‘ p100,p100-SN can interact with STAT6,but p100-TSN can not interact with STAT6;p100 can not interact with STAT1.â‘¡with IL-4 stimulation, STAT6-mediated stronger than without IL-4 stimulation.When add to p100,or p100-SN,this result stronger than without them.Otherwise,p100-TSN do not affect this.With IFN-r stimulation,STAT1-mediated more stronger than without IFN-r stimulation.But when add to p100,there is no stronger effectâ‘¢.construct the plamid pEGFP-CI-STAT6 and pERFP-CI-STAT6 successfully,and can express in HeLa cell.With IL-4 stimulation,we can observe p100,p100-SN can get into nucleus with STAT6.Otherwise,p100-TSN can not.Conclusion:â‘ the result indicate p100,p100-SN can interact with STAT6,and inhance STAT6-dependment transcriptional activation,p100-TSN can not interact with STAT6,and do not effect STAT6-dependment transcriptional activation,p100 do not interact with STAT1,and do not effect STAT1-dependment transcriptional activation.â‘¡successfully construct pEGFP-CI-STAT6 and pERFP-CI-STAT6,and can express in HeLa cell.With IL-4 stimulation,p100 or p100-SN can locate in nuclear with STAT6,which indicate that they are co-location in the nuclus.However, p100-TSN do not.â‘¢with longer IL-4 stimulation,STAT6 can focus granulo-material in nucleus.The reason and mechanism should be further study.