Prediction of probability of oncogenic pathway activation or chemotherapy resistance in individual samples We used publicly available and validated gene expression read outs of oncogenic pathway activation previously generated by experimentally controlled activation of these pathways in vitro

April 7, 2017

he N-terminal portion of NS3 and its protease-dead mutant activated the Notch1 IC induced transcription of the Hes-1 promoter. These data suggest that the protease activity of NS3 is not required for the activation of Notch-mediated transcription. Finally, to investigate whether the NS3 function for activation of Notch-signaling pathway is also functional under the condition of which the other HCV proteins exist, we performed the reporter gene assay using 6-Methoxy-2-benzoxazolinone expression vector for full-length HCV polyprotein. The results show that Notch1 IC-mediated activation of June 2011 | Volume 6 | Issue 6 | e20718 HCV NS3 Can Activate Notch-Signaling Pathway Hes-1 promoter was enhanced by the expression of the full-length HCV polyprotein. NS3 also binds to the fragment of p400 which is homologous to the NS3-binding region of SRCAP Sequence analysis showed that the positive clone obtained by the yeast two-hybrid screening encodes the 1639990 amino-acid region of SRCAP. To investigate whether this region of SRCAP is responsible for the binding to NS3, we constructed a plasmid to express this region of SRCAP and performed a co-immunoprecipitation assay. The results show that HA-tagged SRCAP 1639990 was co-immunoprecipitated with FLAGtagged-NS3, suggesting that this region which almost fully overlaps the previously reported CBP binding site of SRCAP is responsible for binding to NS3. There is a significant sequence similarity of SRCAP with p400, which is known as a transcriptional factor binding to the June 2011 | Volume 6 | Issue 6 | e20718 HCV NS3 Can Activate Notch-Signaling Pathway adenovirus E1A oncoprotein, and sequence analysis showed that the region homologous to the p400 for the NS3 binding region of SRCAP can be assigned to amino acid position 1450808. We constructed a HA-tagged expression vector for this region by cloning, and performed a coimmunoprecipitation assay to investigate the binding activity of NS3 to this region. The results showed that NS3 is also able to bind to this region of p400. The data suggest that p400 must be considered a candidate for host-cellular molecules targeted by NS3 as well as by 18288792 SRCAP. Gene specific silencing of SRCAP and p400 mRNAs by short hairpin RNAs inhibits the NS3-mediated activation of Hes-1 promoter To investigate the effect of SRCAP knockdown in relation to the enhancement of Notch-mediated activation of Hes-1 promoter by June 2011 | Volume 6 | Issue 6 | e20718 HCV NS3 Can Activate Notch-Signaling Pathway 6 June 2011 | Volume 6 | Issue 6 | e20718 HCV NS3 Can Activate Notch-Signaling Pathway NS3, we initially performed the reporter gene assay using short hairpin RNA to specifically silence the SRCAP mRNA expression via RNA interference. However, the knockdown of SRCAP mRNA did not significantly affect the activation of the Hes-1 promoter. As shown in Fig. 4D, the results of the co-immunoprecipitation assay showed that NS3 also exhibits binding activity to a partial coding region of p400 which is a protein closely related to SRCAP. This finding suggested the possibility that both SRCAP and p400 are involved in the enhancement of Notch-mediated Hes-1 promoter activation by NS3. To evaluate this, we constructed a plasmid vector to express shRNA which specifically silences the gene expression of p400 in addition to the shRNA expression vector for SRCAP. These shRNA expression vectors were transfected into HEK293 cells and the expression levels of SRCAP and p400 mRNAs were quantified by real-time RT-PCR to conf