Actin-dependent regulation of connective tissue growth factor.
Expression of connective tissue growth factor (CTGF) in endothelial cells is modulated by shear stress affecting the organization of the cytoskeleton. The molecular connection between alterations of actin and CTGF expression was investigated in human umbilical vein endothelial cells (HUVEC) and a microvascular endothelial cell line. Overexpression of nonpolymerizable monomeric actin R62D interfered with stress fiber formation in HUVEC and concomitantly reduced immunoreactive CTGF. In microvascular endothelial cells, flow-dependent upregulation of CTGF was prevented by this actin mutant. In contrast, overexpression of actin S14C strengthened filamentous actin and increased CTGF expression. These data indicated an inverse relationship between CTGF expression and monomeric actin. Coexpression of the mutant actins and different CTGF promoter constructs revealed an actin-sensitive site between 3 and 4.5 kb of the CTGF promoter. A CArG-like box at -3791 bp was responsible for actin-dependent CTGF induction as shown by mutagenesis. Overexpression of actin S14C activated the nonmutated promoter significantly more strongly than the mutated promoter. Actin polymerization is regulated by the small GTPase RhoA and activation of serum response factor (SRF). Overexpression of constitutively active RhoA or SRF significantly increased CTGF protein synthesis. The 4.5-kb promoter construct, but not the construct with a mutation in the CArG box, was activated by SRF or RhoA, providing evidence for a functional role of this site in CTGF induction. These findings provide novel evidence that monomeric actin is the connecting link between alterations in the cytoskeleton and CTGF gene expression and demonstrate the importance of SRF in regulating CTGF transcription.
Muehlich
Susanne
S
Department of Nephrology and Hypertension, University of Erlangen-Nuremberg, Loschgestrasse 8, D-91054 Erlangen, Germany.
Cicha
Iwona
I
Garlichs
Christoph D
CD
Krueger
Bettina
B
Posern
Guido
G
Goppelt-Struebe
Margarete
M
D000199:Actins / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D002460:Cell Line; D055513:Connective Tissue Growth Factor; D042783:Endothelial Cells / Q000378:metabolism*; D006801:Humans; D017874:Immediate-Early Proteins / Q000235:genetics / Q000378:metabolism*; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D007668:Kidney / Q000098:blood supply; D051379:Mice; D008833:Microcirculation / Q000166:cytology / Q000378:metabolism; D009154:Mutation; D011401:Promoter Regions, Genetic; D027602:Serum Response Element; D026362:Serum Response Factor / Q000235:genetics / Q000378:metabolism; D022502:Stress Fibers / Q000378:metabolism*; D013314:Stress, Mechanical; D014158:Transcription, Genetic; D014162:Transfection; D014471:Umbilical Veins / Q000166:cytology / Q000378:metabolism; D020742:rhoA GTP-Binding Protein / Q000235:genetics / Q000378:metabolism
D000199:Actins; C526317:CCN2 protein, human; C526318:CCN2 protein, mouse; D017874:Immediate-Early Proteins; D036341:Intercellular Signaling Peptides and Proteins; D026362:Serum Response Factor; D055513:Connective Tissue Growth Factor; D020742:rhoA GTP-Binding Protein