Because we also observed a tendency for the down-regulation of NM23-H2 when Diva is present, the effects of Bcl2L10 on NM23-H2 protein levels were further investigated. Transfection of increasing amounts of HA-tagged NM23-H2 yielded higher Acetylsalicylic acid of NM23-H2 as evident by Western blot analysis using anti-HA antibody (Supplementary Fig. 1B; upper panel, lanes 2–5). However, in the presence of Bcl2L10, NM23-H2 protein levels were attenuated (Supplementary Fig. 1B; upper panel, lanes 2–5 vs. 7–10), and increasing expression of Diva also decreased the levels of both endogenous and overexpressed NM23-H2 (Supplementary Fig. 1A; middle panel, lanes 1–12). Together, these data suggest that Diva (Bcl2L10) and NM23-H2 reciprocally affect each other’s protein level; however, the inhibitory effect of NM23-H2 on Diva was more prominent.
Diva, Bcl2L10, and NM23-H2 induce apoptosis, and NM23-H2 silencing increases Diva-induced apoptosis
To assess the biological role of Bcl2L10 and NM23-H2 in apoptosis, we performed cell viability assays and FACS analysis. Transfection of increasing amounts Diva or Bcl2L10 expression plasmids into SK-OV3 cells induced concentration-dependent cell death with similar trends (Supplementary Fig. 2A and B). Overexpression of rat or human NM23-H2 protein also resulted in the reduced cell viability, but to a lesser extent than Diva or Bcl2L10 (Supplementary Fig. 2A and B).
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The tooth sections were examined
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