Assessing Regulation of ERG and its Target Proteins by ERG Isoform 8

Ets-related gene (ERG) is a key protein in vascular homeostasis. It has multiple isoforms. These isoforms have predominant nuclear localization except ERG isoform 8. This isoform can modify the nuclear localization of other isoforms and can down-regulate other isoforms. However, the mechanism of this regulation and its impact on ERG target proteins is still unclear. This study is designed to analyze the effects of ERG8 on functional aspects of other isoforms. For this purpose, the expressions of ERG3 target genes; VE-cad, sox4, cxcr4 were analyzed after ERG8 and ERG3 co-transfection. Increasing ERG8 to ERG3 ratios were transfected in 293-HEK cells. qPCR analysis was performed on total RNA extracted from transfected cells to evaluate the changes in expression of target genes. Results showed that ERG3 alone significantly increased the expression of these target genes (p˂0.05). However, the increasing concentrations of ERG8 progressively decreased the expression of the target genes significantly (p˂0.05). For in silico prediction of the nature of ERG3-ERG8 interaction, their three-dimensional models were built using I-TASSER and interaction was predicted using HADDOCK 2.4. HADDOCK scores show that ERG8 via its pointed (PNT) domain can interact with Ets domain of ERG3 (HADDOCK score; -28.956 +/- 15.881) as well as PNT domain (HADDOCK score; -4.426 +/- 7.770). Our results indicate that ERG8 affect the transcription factor activity of ERG3 by hindering its binding with the DNA either through PNT or Ets domain