Perlecan Domain V Induces VEGf Secretion in Brain Endothelial Cells through Integrin α5β1 and ERK-Dependent Signaling Pathways
Figure 1
DV binding to α5β1 is partially mediated through its DGR sequence.
(A) Sequence schematic of the second EGF repeat within DV demonstrating the exact location, highlighted in yellow, where D3904A DV was mutated. (B) Coomassie stain of SDS PAGE of D3904A DV and LG3 protein preparations (left image for each as labeled) and anti-his immunoblot (right image for each as labeled) to recognize the 6xHis-tag present in the recombinant purified D3904A DV and LG3 (DV C-terminal fragment). In both analyses, D3904A DV yields a single 82 kDa band as does wild-type DV (not shown), and LG3 yields a single 25 kDa band. (C) Optical biosensor traces showing the association and dissociation of D3904A DV with immobilized α5β1 integrin at the concentrations listed (RU = relative units). (D) Quantification of proliferation of BECs after 24 h ± wild-type DV or D3904A DV at 100 nM or 300 nM concentrations in serum starved media as measured via MTS assay. N = 3, *P<0.05 at 100 nM, P = 0.06 at 300 nM. (E) Optical biosensor traces showing the association and dissociation of LG3 with immobilized α5β1 integrin at the concentrations listed (RU = relative units). (F) LG3 (different concentrations of LG3 as labeled) BEC 24 h proliferation assay.