A post-transcriptional pathway represses monocyte VEGF-A expression and angiogenic activity

Ray PS, Fox PL
Source: EMBO J
Publication Date: (2007)
Issue: 26(14): 3360-72
Research Area:
Cancer Research/Cell Biology
Cells used in publication:
Species: human
Tissue Origin: blood
Nucleofector® I/II/2b
(1) U937 cells were nucleofected with eukaryotic, CMV-driven expression vectors containing the FLuc gene upstream of either wild-type (CMV-FLuc-VEGF-A GAIT-A30) or mutant VEGF-A GAIT element (CMV-FLuc-VEGF-A GAITmut-A30) or lacking any GAITelement (CMV-FLuc). Cells were co-transfected with a vector containing RLuc gene under the SV40 promoter. (2) U937 cells were subjected to nucleofection with either pSUPER encoding a small hairpin RNA targeted to human L13a or empty pSUPER vector. Puromycin resistant clones of transfected cells were selected and expanded to form stably transfected cell lines showing knockdown of L13a expression
Monocyte-macrophage activation by interferon (IFN)-gamma is a key initiating event in inflammation. Usually, the macrophage response is self-limiting and inflammation resolves. Here, we describe a mechanism by which IFN-gamma contributes to inflammation resolution by suppressing expression of vascular endothelial growth factor-A (VEGF-A), a macrophage product that stimulates angiogenesis during chronic inflammation and tumorigenesis. VEGF-A was identified as a candidate target of the IFN-gamma-activated inhibitor of translation (GAIT) complex by bioinformatic analysis, and experimentally validated by messenger RNA-protein interaction studies. Although IFN-gamma induced persistent VEGF-A mRNA expression, translation was suppressed by delayed binding of the GAIT complex to a specific element delineated in the 3'UTR. Translational silencing resulted in decreased VEGF-A synthesis and angiogenic activity. Our results describe a unique anti-inflammatory pathway in which IFN-gamma-dependent induction of VEGF-A mRNA is translationally silenced by the same stimulus, and they suggest the GAIT system directs a post-transcriptional operon that contributes to inflammation resolution.