Transforming growth factor-beta up-regulates the beta(5) integrin subunit expression via Sp1 and Smad signaling

Integrin-mediated cell-matrix interactions play important:roles in regulati ng cell function. Since transforming growth factor-beta (TGF-beta) modulate s many osteoblast activities, we hypothesized that the growth factor acts i n part by modulating integrin expression. TGF-beta increased cell adhesion to vitronectin and up-regulated the surface level of alpha (v)beta (5) via increasing beta (5) protein synthesis,by a transcriptional mechanism. Promo ter activity analysis demonstrated that a TGF-beta -responsive element resi des between nucleotides -63 and -44. Electrophoretic mobility shift assay a nd immunoprecipitation/Western studies indicated that the nuclear complex f ormed using the -66/-42 oligonucleotide contained both Sp1/Sp3 and Smad pro teins. Since nuclear Sp1/Sp3 levels were not altered, whereas Smad levels w ere increased by TGF-beta, we investigated the roles of Smad proteins in th e up-regulation of beta (5) gene activation. Co-transfection of cells with beta (5) promoter reporter construct and expression vectors for Smad3, Smad 4, and Spl increased the stimulatory effect of TGF-beta. Furthermore, expre ssion of dominant negative Smad3 or Smad4 in cells decreased or abolished t he stimulation of beta (5) promoter activity by TGF-beta. Smad4 mutant also inhibited the up-regulation of surface beta (5) level by TGF-beta. Thus, T GF-beta increases expression of the integrin beta (5) gene by mechanisms in volving Sp1/Sp3 and Smad transcription factors.