Figure 1. Extracellular 20S standard proteasomes cleave OPN molecules.

(a) Representative in vitro digestion (n = 3) of recombinant OPN-FL, OPN-N, OPN-C by different amount of human erythrocyte 20S standard proteasome. (b) Degradation kinetics of OPN-FL, OPN-N and OPN-C by T2 20S standard proteasomes are shown by representative Western Blot assay (left panel; the proteasome α4 subunit is used as control marker since it is incorporated in each proteasome). The density of the Western Blot bands is measured and the corresponding relative OPNs’ degradation computed (right panel; mean and SD of 4–5 independent experiments measured in duplicate). The relevant bands of the Western Blot assays shown here are cropped from the full blots shown in Supplementary Fig. 1. No significant differences between the degradation rate of the three OPNs are observed by Kruskal-Wallis test. In (b) we use a ratio OPNs/proteasome that roughly resembles that observed in the peripheral blood of healthy and MS donors (Fig. 5) and that used in the cell migration experiments (Fig. 2).