Cancer du sein et micro-environnement tumoral: rôle de la protéase ...

Figure 1. Cath-D interacts with the b chain of LRP1 in vitro(A) Sequence of the LRP1b chain interacting with cath-D. Panel a shows aschematic representation of human cath-D. The locations of 4-kDa cath-D profragment,14-kDa light and 34-kDa heavy mature chains are indicated. Theintermediate 48-kDa form (not shown) corresponds to non-cleaved 14 + 34-kDachains. Number 1 corresponds to the first amino acid of the mature cath-D. Theposition of the catalytic aspartic acid is shown.The sequence of the LRP1 b chain is shown in panel b. LRP1 is synthesized as a4544-amino acid precursor that is cleaved into a a chain and a b chain. Thesequence of the b chain is shown, and the amino acids are numbered starting atthe first residue of the b chain. Residues 307-479, which form the cath-D bindingsite, are shown in bold. The trans-membrane sequence is underlined, and thetwo NPXY motifs are in italics.(B) Binding of the full-length extracellular domain of LRP1b to cath-D. The radiolabeled,full-length extracellular domain of LRP1b (1-476) was incubated withglutathione-Sepharose beads containing GST-cath-D/52K, GST-cath-D/34K, GSTcath-D/14K,GST-cath-D/4K, GST-RAP and GST. GST proteins were stained byCoomassie (panels a, c and e). Bound LRP1b (1-476) was detected byautoradiography (panels b, d and f). The input corresponds to the lysate used forthe binding reaction.(C) Binding of pro-cath-D to LRP1b (307-479). Radio-labeled pro-cath-D wasincubated with beads containing GST-LRP1b (307-479) or GST. GST proteinsstained by Coomassie are shown in panels a and c. Bound pro-cath-D (panel b)was detected by autoradiography. APP was used a negative control (panel d).