Gelatinase A expression in endothelial cells is regulated by at least two cis-acting promoter elements.
Increased expression of gelatinase A is associated with both angiogenesis and alterations in blood vessel structure. Heart-derived endothelial cells derived from spontaneously hypertensive rats (SHR) were found to express significantly more gelatinase A in culture, both at the protein and mRNA level, than endothelial cells from normotensive Wistar-Kyoto (WKY) rats. Other matrix metalloproteinases, as well as their tissue inhibitors, were not differentially regulated. A 1683 bp gelatinase A promoter fragment linked to a luciferase reporter demonstrated up to 40-fold more activity when transfected into SHR-derived cells versus WKY-derived cells. The promoter region between -1324 and -1272, previously termed RE1, contributed up to a five-fold increase in basal promoter activity in both cells, but contributed only 12% of the promoter activity in SHR-derived cells compared to 85% in WKY-derived cells. In SHR-derived cells, but not in WKY-derived cells, a second region between -1435 and -1375, termed RE2, contributed 60% of the total activity of the 1683 bp promoter fragment. Both electrophoretic mobility shift assays and Southwestern blots demonstrated differences in RE2-specific binding factors in nuclear extracts derived from the two cell types. SHR-derived endothelial cells thus represent a new model system to study the regulation of gelatinase A expression, which itself may contribute to the abnormal vascular structure seen in the SHR.
Bottles
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KD
Department of Pathology, BMSB 434, University of Oklahoma Health Sciences Center, 940 Stanton L. Young Blvd., Oklahoma City, OK 73104, USA.
Bullen
E C
EC
Updike
D L
DL
Vu
T K
TK
Phelps
E
E
Grammas
P
P
Howard
E W
EW
D000818:Animals; D001483:Base Sequence; D002467:Cell Nucleus / Q000201:enzymology; D002478:Cells, Cultured; D017077:Culture Media, Conditioned; D004699:Endocardium / Q000201:enzymology*; D018093:Gelatinases / Q000096:biosynthesis / Q000235:genetics*; D005786:Gene Expression Regulation; D008297:Male; D020778:Matrix Metalloproteinase 2; D008666:Metalloendopeptidases / Q000096:biosynthesis / Q000235:genetics*; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D011918:Rats, Inbred SHR; D011921:Rats, Inbred WKY
D017077:Culture Media, Conditioned; D012333:RNA, Messenger; D018093:Gelatinases; D008666:Metalloendopeptidases; D020778:Matrix Metalloproteinase 2