FIGURE 2.

Effect of siRNA-mediated knockdown of EXTL2 on HS chain length. A and B, ³H-labeled (A) or ³⁵S-labeled (B) HS was purified from the cell surface/extracellular matrix of siRNA (siL2M, siL2A, siL2C) and the non-targeting control siRNA-transfected (siNC) and mock-transfected (Mock) HEK293 cells 48 h after transfection as described under “Experimental Procedures.” Isolated glycosaminoglycans were digested with chondroitinase ABC and subjected to gel chromatography on a Superose 6 column. The retarded components eluting at ∼19–23 ml correspond to material that was degraded by chondroitinase ABC. Similar results were obtained from three independent transfections. The samples in A and B were run on two different Superose 6 columns calibrated using size-defined fragments of heparin and hyaluronan. The elution positions of molecular mass standards derived from heparin (8.6 kDa) and hyaluronan (19, 30, 43 and 210 kDa) are indicated by arrows.