Phospholipase A2 activity-dependent stimulation of Ca2+ entry by human parvovirus B19 capsid protein VP1.
Recent reports demonstrated an association of human parvovirus B19 with inflammatory cardiomyopathy (iCMP), which is accompanied by endothelial dysfunction. As intracellular Ca(2+) activity is a key regulator of cell function and participates in mechanisms leading to endothelial dysfunction, the present experiments explored the effects of the B19 capsid proteins VP1 and VP2. A secreted phospholipase A2 (PLA2)-like activity has been located in the VP1 unique region of the B19 minor capsid protein. As PLA2 has recently been shown to activate the store-operated or capacitative Ca(2+) channel I(CRAC), we analyzed the impact of the viral PLA2 motif on Ca(2+) entry. We cloned the VP1 and VP2 genes isolated from a patient suffering from fatal B19 iCMP into eukaryotic expression vectors. We also generated a B19 replication-competent plasmid to demonstrate PLA2 activity under the control of the complete B19 genome. After the transfection of human endothelial cells (HMEC-1), cytosolic Ca(2+) activity was determined by utilizing Fura-2 fluorescence. VP1 and VP2 expression did not significantly modify basal cytosolic Ca(2+) activity or the decline of cytosolic Ca(2+) activity following the removal of extracellular Ca(2+). However, expression of VP1 and of the full-length B19 clone, but not of VP2, significantly accelerated the increase of cytosolic Ca(2+) activity following the readdition of extracellular Ca(2+) in the presence of thapsigargin, indicating an activation of I(CRAC.) The effect of VP1 was mimicked by the PLA2 product lysophosphatidylcholine and abolished by an inactivating mutation of the PLA2-encoding region of the VP1 gene. Our observations point to the activation of Ca(2+) entry by VP1 PLA2 activity, an effect likely participating in the pathophysiology of B19 infection.
Lupescu
Adrian
A
Physiologisches Institut, der Universität Tübingen, Gmelinstr. 5, D-72076 Tübingen, Germany.
Bock
C-Thomas
CT
Lang
Philipp A
PA
Aberle
Susanne
S
Kaiser
Heike
H
Kandolf
Reinhard
R
Lang
Florian
F
D002118:Calcium / Q000378:metabolism*; D036022:Capsid Proteins / Q000235:genetics / Q000502:physiology*; D002460:Cell Line; D045744:Cell Line, Tumor; D003593:Cytoplasm / Q000737:chemistry; D042783:Endothelial Cells / Q000821:virology; D005453:Fluorescence; D016257:Fura-2 / Q000378:metabolism; D006801:Humans; D016732:Parvovirus B19, Human / Q000235:genetics / Q000502:physiology*; D010741:Phospholipases A / Q000235:genetics / Q000378:metabolism*; D054467:Phospholipases A2; D013194:Staining and Labeling; D014162:Transfection
D036022:Capsid Proteins; C095547:capsid protein VP1, parvovirus B19; C095548:capsid protein VP2, parvovirus B19; D010741:Phospholipases A; D054467:Phospholipases A2; D002118:Calcium; D016257:Fura-2