Elevated DS-epi1 levels in human ESCC prompted studies on the functional effects of DS-epi1 in vitro. To evaluate the role of IdoA in tumorigenesis, we made use of TE-1 cells, a well established cellular model of ESCC (15, 23, 24). TE-1 cells displayed the highest DS epimerase activity among the TE-1, -2, -4, -5, -8 cell lines tested, (Supplementary Table II). TE-1 cells were infected by lentiviruses containing two DS-epi1 shRNA sequences (shRNA-a and shRNA-b) and one control shRNA sequence. Two clones containing shRNA-a or shRNA-b were studied. Epimerase activity was downregulated by approx. 90% in shRNA-a transduced cells and approx. 82% in shRNA-b transduced cells, as compared to control non-target shRNA transduced clones (Supplementary Table II). Accordingly, DS-epi1 protein was substantially reduced in shRNA-a and shRNA-b cells (Fig. 2A). Additionally, confocal microscopy analysis demonstrated that DS-epi1 co-localized with the cis Golgi marker GM130 in control cells, while virtually below detection level in shRNA-a cells (Fig. 2B). To study the effect of DS-epi1 down regulation on DS structure, cells were labeled with [35S]-sulfate and PGs derived either from the cell
