Altered lipid raft-associated proximal signaling and translocation of CD45 tyrosine phosphatase in B lymphocytes from patients with systemic lupus erythematosus.
OBJECTIVE
B lymphocytes from patients with systemic lupus erythematosus (SLE) exhibit defective intracellular signaling, hyperactivity, and autoantibody production. Recent evidence indicates a reduced expression of Lyn kinase, a negative regulator of B cell signaling, and reduced translocation of Lyn into membrane signaling domains in SLE. The present study was undertaken to investigate the causes of this altered regulation of Lyn by assessing the expression levels of regulatory molecules and their translocation into the signaling domains of SLE B lymphocytes.


METHODS
Blood was obtained from 48 patients with SLE and 28 healthy controls for the assessment of B lymphocytes. Levels and intracellular distribution of Lyn, CD45, COOH-terminal Src kinase (Csk), and c-Cbl were studied by Western blotting, confocal microscopy, and flow cytometry. The kinetics of signaling molecule translocation to the B cell receptor (BCR)-antigen synapse were investigated by confocal microscopy.


RESULTS
A profound alteration in the expression and translocation of regulatory signaling molecules in membrane domains of B cells from patients with SLE was observed. B lymphocytes from SLE patients, but not those from healthy controls, expressed a low molecular weight isoform of CD45 in lipid raft signaling microdomains. Kinetic studies revealed that translocation of Lyn, CD45, Csk, and c-Cbl led to increased recruitment and retention of Lyn and CD45 in the BCR-antigen synapse in SLE B cells.


CONCLUSIONS
The results provide evidence of altered expression and translocation/interaction of kinases and phosphatases in membrane signaling microdomains of B cells from patients with SLE. Altered translocation of CD45 correlated with reduced expression of Lyn, indicating that Lyn is a key molecule in the regulation of BCR-mediated signaling.
Flores-Borja
Fabian
F
William Harvey Institute, Queen Mary School of Medicine and Dentistry, London, UK.
Kabouridis
Panagiotis S
PS
Jury
Elizabeth C
EC
Isenberg
David A
DA
Mageed
Rizgar A
RA
D000328:Adult; D000368:Aged; D001402:B-Lymphocytes / Q000201:enzymology* / Q000473:pathology; D002478:Cells, Cultured; D005260:Female; D006801:Humans; D017493:Leukocyte Common Antigens / Q000378:metabolism*; D008180:Lupus Erythematosus, Systemic / Q000201:enzymology* / Q000473:pathology; D008297:Male; D021962:Membrane Microdomains / Q000276:immunology / Q000378:metabolism*; D008875:Middle Aged; D021381:Protein Transport; D017027:Protein Tyrosine Phosphatases / Q000096:biosynthesis*; D050721:Proto-Oncogene Proteins c-cbl / Q000378:metabolism; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000378:metabolism; D015398:Signal Transduction / Q000276:immunology; D019061:src-Family Kinases / Q000378:metabolism
D050721:Proto-Oncogene Proteins c-cbl; C085377:MATK protein, human; D016392:Proto-Oncogene Proteins pp60(c-src); C060902:lyn protein-tyrosine kinase; D019061:src-Family Kinases; D017493:Leukocyte Common Antigens; D017027:Protein Tyrosine Phosphatases; C067419:CBL protein, human