Fig. 3

(A-E). IGF1-SRPK2 mediates FASN gene expression and de novo lipid synthesis. A MCF-7, MDA-MB-231, or MCF-10A cells were serum starved 16 hours followed by vehicle (DMSO) or SRPK2 inhibitor (SRPN-340) (3 μM) pretreatment for 1 hour followed by IGF-1 (100 ng/mL) exposure for 6 hours in full growth media. Graphs represent a relative fold change to vehicle control treatment. All values were standardized to actin and are represented as a mean of three independent trials (n = 3) ± SD. Students T test were used to calculate significance. “N.S.” = p-value ≥ .05 B MCF-7 and MDA-MB-231 cells were transfected with either control siRNA or SRPK2 siRNA prior to IGF-1 exposure (100 ng/mL) for 24 hours. SRPK2, FASN, and GAPDH protein expression were detected by western blot. GAPDH served as the loading control. Graphs to the right of the blots are representative of band quantifications from three independent trials (n = 3) ± SD. Students T test were performed as statistical analysis for determining significance. Blots have been cropped for data interpretation and full blots can be found in Fig. S1. C SUM-159, MDA-MB-453, BT-549, and SKBR3 cells were transfected with control SRPK2 siRNA with or without IGF-1 exposure. FASN, SRPK2, and GAPDH protein expression were detected by western blot. Graphs below represent an average of three independent trials (n = 3) ± SD and expressed as a fold change to control siRNA. Students T test were performed as statistical analysis. Blots have been cropped for data interpretation and full blots can be found in Fig. S1. D MDA-MB-231 and MCF-7 cells were transfected with control or SRPK2 siRNAs followed by exposure to media containing charcoal stripped FBS, ¹³c glucose (4.5 g/L), and IGF-1 (100 ng/mL) for 48 hours prior to GC-MS analysis. Graphs represent relative fold change of total intracellular palmitate. Graphs represent an average of 3 independent trials (n = 3) ± S.E.M. Unpaired two-way students t-test were performed for statistical analysis