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. 2011 Jun 15;6(6):e21023. doi: 10.1371/journal.pone.0021023

Figure 1. Progranulin interacts with the beta-propeller region of sortilin.

Figure 1

(A) A schematic drawing of sortilin fragments cloned in the pDisplay vector (full length sortilin ecto domain (residues 81–757), sortilin 10CC region (residues 612–757) or sortilin beta propeller domain (residues 81–611)) and western blot to confirm their expression levels in COS-7 cells using anti-myc antibodies. (B) Binding of 100 nM alkaline phosphatase tagged progranulin (AP-PGRN) to COS-7 cells expressing various sortilin construct as indicated. Expression of these constructs was verified by anti-myc staining. (C) S283E mutation of sortilin abolished progranulin binding. Cell lysates from HEK293T cells transfected with vector control, wild type sortilin or the S283E mutant of sortilin were incubated with anti-Flag beads bound to Flag-PGRN. The presence of sortilin in the 3% input and anti-Flag IP was detected using anti-sortilin antidodies. (D) COS-7 cells expressing wild type sortilin or sortilin S283E mutant were allowed to bind to 100 nM AP-PGRN. The expression of S283E mutant was confirmed with western blot using anti-sortilin antibodies. Scale bar = 100 µm.