Figure 3

Identification of Wscd1 and Wscd2 as SulT-Sia genes. (a) Schematic structure of Wscd1 and Wscd2. Both proteins share the two PAPS-binding motifs, 5′-PSB and 3′-PB, and a single membrane-spanning region, TM. Their amino acid sequences are shown. The schematic structure of the alanine mutants, mutWscd1 and mutWscd2, are presented in Supp_FigS4. (b, e) Western blotting of Mock-, Wscd1- Wscd2-, mutWscd1-, and mutWscd2-transfected CHO cells using antibodies against V5 and GAPDH (loading control). In (b), Lanes 1–3: Mock-, Wscd1-, and mutWscd1-transfected cells, respectively. In (e), Lanes 1–3: Mock-, Wscd2-, and mutWscd2-transfected cells, respectively. Wscd1, Wscd2, mutWscd1, and mutWscd2 were detected at 71 kDa (arrow). (c, f) Flow cytometry analysis (FCA) of 3G9 epitope expression on the transfected cells shown in (b) and (e), respectively. The samples include Mock, Wscd1-expressing, and mutWscd1-expressing cells in (c); Mock, Wscd2-expressing, and mutWscd2-expressing cells in (f). (d, g) The % proportion of 3G9-positive cell population of the histograms in (c) and (f), respectively. All the experiments were performed in triplicate. The error bars indicate the standard deviations; *p < 0.05 (Student t-test, n = 3).