Fig. 4

Knockdown of BMPR2 but not BMP type 1 receptors decreases expression of XIAP and increases cytosolic Smac/DIABLO and cytochrome c. a Quantitative PCR demonstrating a decrease in expression of ALK3 and ALK6 following siRNA knockdown in H1299. Data represents the mean of 2 independent experiments done in duplicate reported as the percent of siRNA control. b Western blot analysis of cells transfected with siRNA targeting alk3 or alk6 alone or in combination for 24 h showing no change in XIAP expression. NS is a nonspecific band used as a loading control. c Western blot analysis of H1299 cells that were transfected with constitutively active ALK3 or ALK6 showing activation of Smad-1/5 but no change in XIAP expression. d Western blot analysis showing that knockdown of ALK3 or ALK6 alone or in combination does not increase cytosolic expression of Smac/DIABLO or cytochrome c. e Quantitative PCR using two different siRNA for controls and BMPR2 demonstrating decreased BMPR2 RNA expression in H1299 cells. Data represents the mean of 2 independent experiments done in duplicate reported as the percent of the siRNA control. f Western blot analysis of H1299 cells transfected with control or BMPR2 siRNA showing a decrease in XIAP expression. g The actin and BMPR2 bands were quantified using Image J and reported as the ratio of BMPR2/actin. h Western blot analysis of A549 cells transfected with control or BMPR2 siRNA showing a decrease in XIAP expression. i The GAPH and BMPR2 bands were quantified using Image J and reported as the ratio of BMPR2/GAPDH. j Western blot analysis of cytosol of A549 cells following knockdown of BMPR2 using two different siRNA demonstrating an increase in Smac/DIABLO and cytochrome c expression. k Western blot analysis showing knockdown of BMPR2 decreases BMP signaling. l Western blot of H1299 cells following BMPR2 knockdown demonstrating an increase in cytosolic Smac/DIABLO. b-l Studies were done at least 3 times with similar results