Glypican 4, a membrane binding protein for bactericidal/permeability-increasing protein signaling pathways in retinal pigment epithelial cells.
OBJECTIVE
Originally identified as a lipopolysaccharide binding protein with Gram-negative bactericidal activity in the leukocytes, bactericidal/permeability-increasing protein (BPI) has been shown to induce various effects in retinal cells in vivo and in vitro.


METHODS
The authors recently reported that BPI can induce ERK1/2 and Akt activity and that it increases DNA synthesis in the bovine retinal pigment epithelial (RPE) and pericyte cells. The authors have extended the characterization of BPI interaction with membrane proteins from bovine RPE. Crude membrane pools from RPE were isolated, solubilized, and bound to rBPI(21) affinity column. Bound proteins were separated by SDS-PAGE and stained with Coomassie blue, which showed an intense band at 36 kDa consistently displaced by rBPI(21).


RESULTS
Tandem mass spectrometry of the 36-kDa band suggested that cell surface protein glypican 4 (GPC4) serves as a putative BPI-binding protein. Heparitinase, phosphatidylinositol-specific phospholipase C, and anti-GPC4 antibody suppressed BPI-induced ERK and Akt phosphorylation in bovine RPE. Moreover, heparitinase also inhibited BPI actions on VEGF and PDGF-B mRNA expression induced by H(2)O(2).


CONCLUSIONS
These new findings suggest that GPC4 is a specific binding protein for BPI on RPE to mediate the activation of ERK1/2, Akt, and the mRNA expressions of PDGF-B and VEGF.
Research Division, Joslin Diabetes Center, Boston, MA 02215, USA.
Pedro
P
Geraldes
Michiko
M
Yamagata
Susan L
SL
Rook
Yukio
Y
Sassa
Ronald C
RC
Ma
Allen
A
Clermont
Benbo
B
Gao
Lloyd Paul
LP
Aiello
Edward P
EP
Feener
George L
GL
King
D000818:Animals; D023181:Antimicrobial Cationic Peptides / Q000737:chemistry / Q000378:metabolism*; D001798:Blood Proteins / Q000737:chemistry / Q000378:metabolism*; D002417:Cattle; D002478:Cells, Cultured; D004591:Electrophoresis, Polyacrylamide Gel; D053673:Glypicans / Q000737:chemistry / Q000378:metabolism*; D015151:Immunoblotting; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D010766:Phosphorylation; D010857:Pigment Epithelium of Eye / Q000187:drug effects / Q000378:metabolism*; D011133:Polysaccharide-Lyases / Q000494:pharmacology; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D020574:Proto-Oncogene Proteins c-sis / Q000235:genetics; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000502:physiology*; D053719:Tandem Mass Spectrometry; D010738:Type C Phospholipases / Q000494:pharmacology; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics
D023181:Antimicrobial Cationic Peptides; D001798:Blood Proteins; D053673:Glypicans; D008565:Membrane Proteins; D020574:Proto-Oncogene Proteins c-sis; D012333:RNA, Messenger; D042461:Vascular Endothelial Growth Factor A; C021623:bactericidal permeability increasing protein; D051057:Proto-Oncogene Proteins c-akt; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D010738:Type C Phospholipases; D011133:Polysaccharide-Lyases; C021349:heparitinsulfate lyase