Fig 2: SETD3 interacts with and methylates FoxM1 in cells.(A) FoxM1 was immunoprecipitated from cells using an anti-FoxM1 antibody or control beads followed by western blot analysis with the indicated antibodies. Input is shown in the bottom panel. (B,C) HEK-293T cells were transfected with the indicated plasmids or with an empty plasmid. The chromatin fraction was then subjected to immunoprecipitation with FLAG M2 beads followed by western blot with the indicated antibodies. (D) Cells were transfected without or with HA-SETD3 and whole cell lysate was subjected to immunoprecipitation with a pan-methyl antibody followed by western blot with the indicated antibodies. (E) Chromatin isolated fraction of the transfected cells were subjected to immunoprecipitation with FoxM1 antibody followed by immunoprecipitation with pan-methyl antibody.

Fig 3: SETD3 and FoxM1 are enriched at the VEGF promoter.(A,B) ChIP-qPCR analysis of the occupancy of FoxM1 (A) and SETD3 (B) at the VEGF promoter in U87 and HeLa cells. Data are shown as % of input, from at least two independent experiments (error bars, S.E.M).

Fig 4: SETD3 dissociates from the VEGF promoter under hypoxic conditions.(A) HEK-293T cells transfected with FLAG-SETD3 were subjected to normoxic (N) or hypoxic (H) conditions. The chromatin fraction was then subjected to immunoprecipitation with FLAG M2 beads followed by western blot with the indicated antibodies. (B) Chromatin immunoprecipitation followed by real-time qPCR analysis of the occupancy of FoxM1 (left) or SETD3 (right) at the VEGF promoter in control and CoCl2-treated HeLa cells. Data are shown as % of input, from at least three independent experiments (error bars, S.E.M). (C) Suggested model for the interplay between SETD3 and FoxM1 at chromatin under normoxic and hypoxic conditions (see text).