Intronic regulation of matrix metalloproteinase-2 revealed by in vivo transcriptional analysis in ischemia.
Matrix metalloproteinase-2 (MMP-2) plays an essential role in angiogenesis and arteriogenesis, two processes critical to restoration of tissue perfusion after ischemia. MMP-2 expression is increased in tissue ischemia, but the responsible mechanisms remain unknown. We studied the transcriptional activation of the MMP-2 gene in a model of hindlimb ischemia by using various MMP-2-lacZ reporter mice and chromatin immunoprecipitation. MMP-2 activity and mRNA were increased after hindlimb ischemia. Mice with targeted deletion of MMP-2 had impaired restoration of perfusion and a high incidence of limb gangrene, indicating that MMP-2 plays a critical role in ischemia-induced revascularization. Ischemia induced the expression and binding of c-Fos, c-Jun, JunB, FosB, and Fra2 to a noncanonical activating protein-1 (AP-1) site present in the MMP-2 promoter and decreased binding of the transcriptional repressor JunD. Ischemia also activated the expression and binding of p53 to an adjacent enhancer site (RE-1) and increased expression and binding of nuclear factor of activated T-cells-c2 to consensus sequences within the first intron. Deletion of either the 5' AP-1/RE-1 region of the promoter or substitution of the first intron abolished ischemia-induced MMP-2 transcription in vivo. Thus, AP-1 transcription factors and intronic activation by nuclear factor of activated T-cells-c2 act in concert to drive ischemia-induced MMP-2 transcription. These findings define a critical role for MMP-2 in ischemia-induced revascularization and identify both previously uncharacterized regulatory elements within the MMP-2 gene and the cognate transcription factors required for MMP-2 activation in vivo after tissue ischemia.
Lee
Jackie G
JG
Division of Vascular Surgery, San Francisco Veterans Affairs Medical Center, and Pacific Vascular Research Laboratory, University of California, San Francisco, CA 94121, USA.
Dahi
Sia
S
Mahimkar
Rajeev
R
Tulloch
Nathaniel L
NL
Alfonso-Jaume
Maria A
MA
Lovett
David H
DH
Sarkar
Rajabrata
R
D000818:Animals; D004742:Enhancer Elements, Genetic; D006614:Hindlimb / Q000098:blood supply; D007438:Introns; D007511:Ischemia / Q000201:enzymology*; D020778:Matrix Metalloproteinase 2 / Q000235:genetics*; D051379:Mice; D008810:Mice, Inbred C57BL; D018482:Muscle, Skeletal / Q000201:enzymology; D050778:NFATC Transcription Factors / Q000378:metabolism; D011401:Promoter Regions, Genetic; D018808:Transcription Factor AP-1 / Q000378:metabolism; D014158:Transcription, Genetic; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism
D050778:NFATC Transcription Factors; C494007:Nfatc2 protein, mouse; D018808:Transcription Factor AP-1; D016159:Tumor Suppressor Protein p53; D020778:Matrix Metalloproteinase 2