Validation of NRP1 and NRP2 knockdown effects on SC cluster formation.

(A) mRNA levels of NRP1 were decreased in SCs after transfection with the siRNA pool or either of the 4 single siRNAs. Expression levels were ranging from 9 to 66% of endogenous expression measured in the siGlo condition. Knockdown was significant for all conditions (p<0.001) except for siNRP1 single 1 (p<0.12). The value presented for the siNRP1 pool is the result of 4 different experiments. The values for the siNRP1 singles are the result of 1 experiment with 4 technical replicates. The siGlo condition presented in the graph is the control for the single siRNA conditions. (B) Three single siRNAs confirm the significant decrease in the number of clusters which is observed with the siNRP1 pool (p<0.01). (C) Two of the single siRNAs confirm the increase of cluster size observed with the siNRP1 pool (p<0.001). All values are normalized to the siGlo condition. The presented values represent the average of 3 separate experiments. The differential effects of siNRP1 single 2 and 3 may be the result of off-target effects (D) mRNA levels of NRP2 were decreased in SCs after transfection with the siRNA pool or either of the 4 single siRNAs. Expression levels were ranging from 18 to 64% of endogenous expression measured in the siGlo condition. Knockdown was significant for all conditions (p<0.05). The value presented for the siNRP2 pool is the result of 4 different experiments. The values for the siNRP1 singles are the result of 1 experiment with 4 technical replicates. The siGlo condition presented in the graph is the control for the single siRNA conditions. (E) All 4 single siRNAs confirm the significant decrease in the number of clusters which is observed with the siNRP2 pool (p<0.01). The presented values represent the average of 3 separate experiments. Error bars indicate the SEM.