Abstract 14957: Tristetraprolin and Poly(ADP-Ribose)-Polymerase-14 Post-Transcriptionally Regulate Macrophage Tissue Factor Expression: Novel Mechanisms for Regulating Thrombosis
BACKGROUND Monocyte-derived tissue factor (TF) plays critical roles in atherothrombosis. Little is known about its post-transcriptional regulation. Tristetraprolin (TTP), the most widely studied mRNA-binding protein, binds to the 3’UTR of target mRNAs, promoting their degradation, with its function negatively regulated by p38 MAPK. Poly(ADP-ribose)-polymerase-14 (PARP-14), belongs to a family of ∼17 proteins with a PARP domain that, when catalytically active, generates negatively charged poly(ADP-ribose) adducts on intracellular proteins. We have recently identified PARP-14 as a novel RNA-binding protein. Whether TTP and PARP-14 regulate TF is unknown.
METHODS/RESULTS p38 inhibition reduced procoagulant activity; TF mRNA and protein expression; and TF mRNA stability in human macrophages (p<0.05). TF mRNA and protein expression were increased in TTP-/- vs TTP+/+ macrophages (p<0.05), with increased TF mRNA stability (t1/2=352±82min vs. 97±15min,p<0.001). Similarly procoagulant activity, TF mRNA and protein levels were increased in PARP14-/- vs. PARP-14+/+ macrophages (p<0.05), with increased TF mRNA stability (t1/2=423±66min vs. 63±5min,p<0.001). RNP immunoprecipitation and RNA biotin pulldown assays demonstrated that TTP and PARP-14 interact with TF 3’UTR. Furthermore, PARP-14 and TTP form a ternary complex with the same ARE segment of TF mRNA 3’UTR. Thus, TTP binding to TF 3’UTR was reduced in the absence of PARP-14, and vice-versa. TF mRNA and activity were increased in vivo (heart, lung, liver and aorta) in PARP-14-/- vs. PARP-14+/+ mice (p<0.05). Preliminary findings using intravital microscopy demonstrate a 57% reduction in median arteriolar occlusion time in PARP-14-/- vs. PARP-14+/+ mice (129s vs. 291s, n=5).
CONCLUSIONS These data define novel roles for TTP and PARP-14 regulating TF mRNA turnover. Specifically, PARP-14 post-transcriptionally regulates TF expression through a mechanism involving interaction with TTP. PARP-14 deficiency translates to increased TF expression in vivo, and preliminary results indicate increased thrombogenicity (ongoing work). A better understanding of the post-transcriptional regulation of TF expression provides novel insights into thrombosis, and holds therapeutic potential.
- © 2012 by American Heart Association, Inc.