Transforming growth factor 13 (TGF-3) is the name of a group of closely related polypeptides characterized by a multiplicity of effects, including regulation of extracellular proteolysis and turnover of the extracellular matrix. Its cellular mechanism of action is largely unknown. TGF-131 is a strong and fast inducer of type 1 plasminogen activator inhibitor gene transcription. We have identified a TGF-131-responsive element in the 5'-flanking region of the human type 1 plasminogen activator inhibitor gene and shown that it is functional both in its natural context and when fused to a heterologous nonresponsive promoter. Footprinting and gel retardation experiments showed that two different nuclear factors, present in extracts from both TGF-131-treated and nontreated cells, bind to adjacent sequences contained in the responsive unit. A palindromic sequence binds a trans-acting factor(s) of the CCAAT-binding transcription factor-nuclear factor I family. A partially overlapping dyad symmetry interacts with a second protein that much evidence indicates to be USF. USF is a transactivator belonging to the basic helix-loop-helix family of transcription factors. Mutations which abolish the binding of either CCAAT-binding transcription factor-nuclear factor I or USF result in reduction of transcriptional activation upon exposure to TGF-131, thus showing that both elements of the unit are necessary for the TGF-131 response. We discuss the possible relationship of these findings to the complexity of the TGF-13 action.Transforming growth factor I0 (TGF-3) is a general regulator of cellular activities with a multiplicity of effects in the normal organism. TGF-ps include a group of chemically closely related 25,000-Mr dimeric polypeptides, TGF-pl, , with largely identical effects. (Unless otherwise specified, "TGF-P3" will refer to TGF-fi1 heterodimers throughout the rest of the text.) TGF-,B can be synthesized by most cells of the organism and is stored in large quantities in blood platelets. It acts presumably in an autocrine and paracrine manner. Besides affecting cellular proliferation, TGF-3 influences the differentiation of several cell types; for instance, it inhibits myogenesis and adipogenesis and stimulates chondrogenesis. TGF-, regulates the turnover of the extracellular matrix. It stimulates production by fibroblasts of extracellular matrix components, such as type I, II, and III procollagen, fibronectin, and proteoglycans, and of cellular receptors for such proteins. TGF-1 inhibits production of enzymes catalyzing degradation of the extracellular matrix and stimulates production of inhibitors of such enzymes (22,27,31,41,47).It has previously been demonstrated that TGF-p induces type 1 inhibitor of plasminogen activators (PAI-1) in cultured human fibroblasts (24,26). PAI-1 is an important regulator of plasminogen activation and thus of extracellular proteolytic events, including fibrinolysis and degradation of the extracellular matrix (1, 6). The induction of PAI-1 protein by * Corresponding author. TGF-0 was la...

We have characterized regulation of type-1 plasminogen activator inhibitor (PAI-1) gene expression by phorbol 12-myristate 13-acetate (PMA) and the CAMP-inducing agent forskolin in the human breast carcinoma cell line MCF-7. PMA caused a strong induction of PAI-1, while forskolin suppressed the PMA response. Transfection experiments with fusion genes showed that sequences mediating PMA induction as well as forskolin suppression were present between base pairs -100 and -30 of the 5'-flanking region of the PAI-1 gene. The region was found to contain two Spl binding sites. A proximal sequence in the region, TGAGTTCA (P box), with sequence similarity to phorbol ester response elements (TRE) as well as to cAMP response elements (CRE), bound a lowabundance, as yet unidentified nuclear protein in MCF-7 cells. This sequence had a higher affinity to purified c-jun homodimer than to c-junk-fos heterodimer in MCF-7 nuclear extracts; it had no affinity to the proteins binding to CRE consensus sequences in these extracts. A distal TRE-like sequence, TGAGTGG (D box), had a weak affinity to c-junk-fos heterodimer and c-jun homodimer ; binding of proteins to this sequence was facilitated by binding of proteins to the P box. Both the P box and the D box were necessary for PMA responsiveness, suggesting a cooperativity between the two binding sites. A mutation of the P box removing the CRE similarity abolished the forskolin suppression of the PMA response. We propose that the protein kinase C and the protein kinase A signal-transduction pathways, with opposite effects on PAI-1 gene expression, converge by modulating differently P-box-binding proteins.The plasminogen activation system includes the urokinase-type plasminogen activator (u-PA), the tissue-type plasminogen activator (t-PA), the specific and fast acting inhibitors of plasminogen, PAI-1 and PAI-2, and the urokinase receptor. The plasminogen activation system has many physiological functions, both under normal and pathological conditions, including fibrinolysis and the turn-over of the extracellular matrix (for reviews, see Dan@ et al., 1985;Andreasen et al., 1990;Vassalli et al., 1991 ;Pollanen et al., 1991).Correspondence to P.

The structure of the exon-intron boundary was compared for an intron within 23S ribosomal RNA of Desulfurococcus mobilis and a newly discovered intron in tRNA(Met) from the same organism. The occurrence of a putative common structural feature suggests that intron excision occurs by the same mechanism. The possible recognition of this structural feature by the cleavage enzyme was investigated for the ribosomal RNA intron using RNA substrates exhibiting various exon and intron deletions. The results support the involvement of the structural features in the cleavage process. The evolutionary implications of these results are considered.

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Transforming growth factor beta (TGF-beta) is the name of a group of closely related polypeptides characterized by a multiplicity of effects, including regulation of extracellular proteolysis and turnover of the extracellular matrix. Its cellular mechanism of action is largely unknown. TGF-beta 1 is a strong and fast inducer of type 1 plasminogen activator inhibitor gene transcription. We have identified a TGF-beta 1-responsive element in the 5'-flanking region of the human type 1 plasminogen activator inhibitor gene and shown that it is functional both in its natural context and when fused to a heterologous nonresponsive promoter. Footprinting and gel retardation experiments showed that two different nuclear factors, present in extracts from both TGF-beta 1-treated and nontreated cells, bind to adjacent sequences contained in the responsive unit. A palindromic sequence binds a trans-acting factor(s) of the CCAAT-binding transcription factor-nuclear factor I family. A partially overlapping dyad symmetry interacts with a second protein that much evidence indicates to be USF. USF is a transactivator belonging to the basic helix-loop-helix family of transcription factors. Mutations which abolish the binding of either CCAAT-binding transcription factor-nuclear factor I or USF result in reduction of transcriptional activation upon exposure to TGF-beta 1, thus showing that both elements of the unit are necessary for the TGF-beta 1 response. We discuss the possible relationship of these findings to the complexity of the TGF-beta action.