Abstract:
Urokinase-type plasminogen activator (uPA) is
expressed by lung epithelial cells and regulates fibrin turnover
and epithelial cell viability. PMA, LPS, and TNF-alpha, as well
as uPA itself, induce uPA expression in lung epithelial cells.
PMA, LPS, and TNF-alpha induce uPA expression through
increased synthesis as well as stabilization of uPA mRNA,
while uPA increases its own expression solely through uPA
mRNA stabilization. The mechanism by which lung epithelial
cells regulate uPA expression at the level of mRNA stability is
unclear. To elucidate this process, we sought to characterize
protein−uPA mRNA interactions that regulate uPA expres-
sion. Regulation of uPA at the level of mRNA stability involves the interaction of a ∼40 kDa cytoplasmic−nuclear shuttling
protein with a 66 nt uPA mRNA 3′UTR sequence. We purified the uPA mRNA 3′UTR binding protein and identified it as
ribonucleotide reductase M2 (RRM2). We expressed recombinant RRM2 and confirmed its interaction with a specific 66 nt uPA
3′UTR sequence. Immunoprecipitation of cell lysates with anti-RRM2 antibody and RT-PCR for uPA mRNA confirmed that
RRM2 binds to uPA mRNA. Treatment of Beas2B cells with uPA or LPS attenuated RRM2−endogenous uPA mRNA
interactions, while overexpression of RRM2 inhibited uPA protein and mRNA expression through destabilization of uPA mRNA.
LPS exposure of lung epithelial cells translocates RRM2 from the cytoplasm to the nucleus in a time-dependent manner, leading
to stabilization of uPA mRNA. This newly recognized pathway could influence uPA expression and a broad range of uPA-
dependent functions in lung epithelial cells in the context of lung inflammation and repair.
