PRMT4 interacts with Mi2α and Mi2β.

<p>A, B: Co-immunoprecipitation of PRMT4 with Mi2α. HEK293 cells were transfected with Flag-Mi2α construct (+) or empty vector (−). After 48 hours, cells were lysed and 1 mg protein extract was subjected to IP of (A) endogenous PRMT4 (α-PRMT4) or (B) overexpressed Flag-Mi2α (α-Flag). IPs using isotype control IgG were performed in parallel (α-ctrl). Input (1%) and immunoprecipitates were stained by Western Blot analysis using anti-Flag and anti-PRMT4 antibodies. C: Among the PRMT family, PRMT4 preferentially interacts with Mi2α. Protein lysates of Flag-Mi2α-overexpressing HEK293 cells (as in A, B) were subjected to GST-pulldown experiments. Equal amounts of recombinant proteins GST alone and GST-PRMT1, -3, -4 and -6 (<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003343#pgen.1003343.s001" target="_blank">Figure S1</a>) coupled to glutathione beads were incubated with 250 µg of HEK293 extracts. Input (1%) and bound Flag-Mi2α protein were visualised by Western Blot analysis using anti-Flag antibodies. D: Co-immunoprecipitation of PRMT4 with Mi2β. HEK293 cells were transfected with HA-Mi2β construct (+) or empty vector (−). Protein extracts were subjected to IP using anti-PRMT4 (α-PRMT4) antibodies or isotype control IgG (α-ctrl). Input (1%) and precipitates were stained by Western Blot analysis using anti-HA and anti-PRMT4 antibodies.