Despite the important role played by the nonmuscle
isoform of
myosin light chain kinase (nmMLCK) in vascular barrier regulation and the implication of both nmMLCK and
vascular endothelial growth factor (
VEGF) in the pathogenesis of
acute respiratory distress syndrome (ARDS), the role played by nmMLCK in
VEGF-induced vascular permeability is poorly understood. In this study, the role played by nmMLCK in
VEGF-induced vascular hyperpermeability was investigated. Human lung endothelial cell barrier integrity in response to
VEGF is examined in both the absence and the presence of nmMLCK small interfering RNAs. Levels of nmMLCK
messenger RNA (
mRNA),
protein, and promoter activity expression were monitored after
VEGF stimulation in lung endothelial cells. nmMYLK promoter activity was assessed using nmMYLK promoter
luciferase reporter constructs with a series of nested deletions. nmMYLK transcriptional regulation was further characterized by examination of a key transcriptional factor. nmMLCK plays an important role in
VEGF-induced permeability. We found that activation of the
VEGF signaling pathway in lung endothelial cells increases MYLK gene product at both
mRNA and
protein levels. Increased nmMLCK
mRNA and
protein expression is a result of increased nmMYLK promoter activity, regulated in part by binding of the
Sp1 transcription factor on triggering by the
VEGF signaling pathway. Taken together, these findings suggest that MYLK is an important ARDS candidate gene and a therapeutic target that is highly influenced by excessive
VEGF concentrations in the inflamed lung.