Regulation of the stepwise proteolytic cleavage and secretion of PDGF-B by the proprotein convertases.
Platelet-derived growth factor-B (PDGF-B) is important for normal tissue growth and maintenance and its overexpression has been linked to several diseases, including cancer, fibrotic disease and atherosclerosis. Here, we show that synthesized as a precursor, proPDGF-B is converted to a mature form by proteolytic cleavage at two sites and its N-terminal cleavage is a prerequisite for processing at its C-terminus. The first cleavage occurs at residues RGRR81/, and the second cleavage close to residues ARPVT190, just before the C-terminal amino-acid sequence crucial for PDGF-B retention to cell surface. Cotransfection of a Furin-deficient cell line LoVo-C5 with proPDGF-B and different PC members revealed that Furin, PACE4, PC5, and PC7 are candidate proPDGF-B convertases. This finding is consistent with the in vitro digestions of a synthetic peptide mimicking the cleavage site of proPDGF-B. The processing of proPDGF-B is blocked by site-directed mutagenesis of the RGRR81/ sequence and by various PC inhibitors. Mutation of the PDGF-A and/or PDGF-B convertase sites, revealed that processing of both A and B chains is required for the formation of mature PDGF-B dimers and that the processing of the B chain controls the level of secreted and matrix-bound PDGF-BB forms. Our findings emphasize the importance of the convertase-directed processing of proPDGF-B at the RGRR81/ sequence for PDGF-B maturation and secretion.
Siegfried
Geraldine
G
Laboratory of Biochemical Neuroendocrinology, Clinical Research Institute of Montreal, 110 Pine Ave West, Montreal, Quebec, Canada H2W 1R7.
Basak
Ajoy
A
Prichett-Pejic
Wendy
W
Scamuffa
Nathalie
N
Ma
Liming
L
Benjannet
Suzanne
S
Veinot
John P
JP
Calvo
Fabien
F
Seidah
Nabil
N
Khatib
Abdel-Majid
AM
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D017931:DNA Primers; D019281:Dimerization; D045683:Furin / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D007150:Immunohistochemistry; D051379:Mice; D008969:Molecular Sequence Data; D043484:Proprotein Convertases / Q000378:metabolism*; D011498:Protein Precursors / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D020574:Proto-Oncogene Proteins c-sis / Q000737:chemistry / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D017931:DNA Primers; D011498:Protein Precursors; D020574:Proto-Oncogene Proteins c-sis; D043484:Proprotein Convertases; D045683:Furin