Figure 2.

Co-transfection/co-IP experiments clarify the physical interaction of OGT1 and NSL3. A, clarification of physical interaction of OGT1 and NSL3 in 293T cells. Cells were co-transfected with the indicated FLAG- or Myc-tagged plasmids. 48 h after transfection, the prepared whole-cell lysate was subjected to co-IP experiments. Anti-FLAG- or anti-Myc-agarose eluates were subjected to SDS-PAGE, and bound proteins were detected by Western blotting with anti-FLAG or anti-Myc antibody. Fl:NSL3, FLAG-NSL3; Fl:PHF20, FLAG-PHF20. B and C, detection of endogenous OGT1 or NSL3 using IP approaches. 293T cells were transfected with Myc-OGT1 or FLAG-NSL3 to perform IP experiments. Immunoprecipitated endogenous NSL3 or OGT1 was then detected by Western blotting using specific antibodies. D and E, a schematic of the deletion mutants of OGT1 (D) and NSL3 (E). F and G, mapping the binding region between OGT1 and NSL3. 293T cells were co-transfected with full-length NSL3 and the indicated deletion mutants of OGT1 (F) or full-length OGT1 and the indicated deletion mutants of NSL3 (G). 48 h later, co-IP experiments were performed with prepared whole-cell lysate. Bound proteins were detected using Western blot analysis with the indicated antibodies. IB, immunoblot.