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M B Sporn

Publications and source records attributed to M B Sporn.

At least 91 records · Page 5Linked to original sources
PubMed1993

An element of the transforming growth factor-beta 1 5'-untranslated region represses translation and specifically binds a cytosolic factor.

In many cell types, there is a discrepancy between transforming growth factor-beta 1 (TGF-beta 1) mRNA and TGF-beta 1 protein, suggesting that expression of TGF-beta 1 is regulated posttranscriptionally. We have previously shown that a 137-nucleotide (nt) region of the TGF-beta 1 5'-untranslated region (UTR) potently inhibits the expression of a heterologous reporter gene, suggesting a role for this region in the posttranscriptional inhibition of TGF-beta 1 expression. To study the mechanism of inhibition, a chimeric plasmid containing this region of the TGF-beta 1 5'-UTR and the reading frame of the human GH gene was stably transfected into C2C12 myoblastic cells. Our results show that the TGF-beta 1 5'-UTR inhibits GH expression by inhibiting GH mRNA translation. In vitro gel retardation and cross-linking assays using a radiolabelled RNA probe transcribed from this region of the TGF-beta 1 5'-UTR demonstrate the specific binding of a cytosolic factor. Deletion of a potential stem-loop-forming region abolishes binding of this factor and partially restores GH production. These results suggest that posttranscriptional inhibition of TGF-beta 1 expression is at the level of mRNA translation and that a cytosolic factor may regulate TGF-beta 1 mRNA translation.

Animals↗
PubMed1993

Synergistic increase of phorbol ester-induced c-fos mRNA expression by retinoic acid through stabilization of the c-fos message.

Retinoic acid (RA) has been shown to be able to antagonize or synergize with phorbol 12-myristate 13-acetate (PMA). In contrast to its antagonistic effects on PMA-dependent gene expression, no molecular target or mechanism of synergism has been characterized yet. We now report, that RA synergistically enhances the induction of c-fos, but not c-jun mRNA by PMA in cells whose growth was stimulated by RA alone. The responding cells were hybrids of tumor cell lines whose growth and PMA-dependent c-fos mRNA expression remained unaffected by RA. The increase in PMA-dependent c-fos expression required pretreatment of cells with RA for at least 2-4 h and was achieved at doses as low as 10(-10) M. Nuclear run-on experiments and transient transfection assays using a chimeric reporter gene construct with sequences from the c-fos promoter indicated that RA did not affect PMA-dependent c-fos transcription. Instead, RA stabilized the c-fos message after induction by PMA as assessed by measuring the half-life of c-fos mRNA in actinomycin D-treated cells. This post-transcriptional regulation provides a mechanism whereby RA can synergistically enhance gene expression by PMA.

Dose-Response Relationship, Drug↗
PubMed1993

Expression of transforming growth factor-beta isoforms in small round cell tumors of childhood. An immunohistochemical study.

The transforming growth factor (TGF)-betas are a highly conserved group of potent multifunctional cell regulatory proteins with variable effects on cell growth and differentiation. Most of the small round cell group of childhood tumors are thought to arise from either primitive mesenchyme or neuroectoderm and show evidence of skeletal muscle or neural differentiation, and rarely both. To investigate the possibility that the TGF-betas have a role in the growth or differentiation of these neoplasms, we used antibodies specific for peptide sequences of the three known mammalian TGF-beta isoforms (TGF-betas 1, 2, and 3) to probe for TGF-beta protein expression in a total of 49 cases. TGF-beta 1 immunoreactivity was present in 16/17 (94%) of rhabdomyosarcomas, and the staining intensity was usually strong. TGF-beta 1 was also present in three of three ectomesenchymomas. In contrast, TGF-beta 1 was absent in all but one out of nine poorly differentiated neuroblastomas. Differentiating neuronal cells of ganglioneuroblastomas, however, were strongly positive for TGF-beta 1. Ewing's sarcomas and peripheral primitive neuroectodermal tumors had a less consistent, but usually positive, staining pattern. TGF-beta 3 staining patterns were very similar to those of TGF-beta 1. TGF-beta 2 immunoreactivity was only rarely detected in this group of tumors. The results suggest different roles for TGF-betas 1 and 3 in neuroblastoma and rhabdomyosarcoma. Expression of TGF-betas 1 and 3 is associated with neuronal differentiation of neuroblastoma. In contrast, these proteins may promote the growth of rhabdomyosarcoma by suppressing differentiation.

Adolescent↗
PubMed1992

Identification of an activating transcription factor (ATF) binding site in the human transforming growth factor-beta 2 promoter.

Transforming growth factor TGF-beta 2 is encoded by multiple mRNA transcripts of 5.8, 5.1, 4.0, 3.8, and 2.8 kilobase pairs (kb) that are expressed in various human and monkey cells. Northern blot analysis using genomic fragments of DNA was used to demonstrate that some of this size heterogeneity is due to differences in the length of the 5'-untranslated region. Probes that were colinear with the first 600 nucleotides of the 5'-untranslated region detected only the 5.8-, 4.0-, and 3.8-kb transcripts. In order to identify DNA elements that regulate the transcription of these mRNA transcripts, deletion constructs of 5'-flanking DNA were ligated to the coding region for chloramphenicol acetyltransferase (CAT) and analyzed for promoter activity in several cell lines. Sequences responsible for putative enhancer and silencer regions were identified between -778 and -40 relative to the transcription initiation site. Addition of a cyclic AMP-responsive element/activating transcription factor-like element at -74 resulted in a 5-10-fold increase in CAT activity over that expressed with a construct that contained only the TATA box. This increase in CAT activity was suppressed by the addition of DNA sequences between -257 and -187, whereas sequences between -778 and -257 stimulated CAT activity. Point mutations within the ATF binding site at -74 resulted in a marked decrease in CAT expression. Cotransfection with ATF-1 or ATF-2 expression plasmids resulted in both dose-dependent stimulatory and inhibitory activities that were cell type-dependent. These studies identify multiple transcription initiation sites for TGF-beta 2 and demonstrate that transcription from one of these promoters is dependent upon an ATF binding site located 5' of the TATA box.

Activating Transcription Factors↗
PubMed1992

Inhibition of mitogen-induced c-fos expression in melanoma cells by retinoic acid involves the serum response element.

To investigate the mechanism(s) by which all-transretinoic acid (RA) inhibits cell growth, we studied its effect on the expression of c-fos and c-jun in B16 melanoma cells. RA differentially inhibited proto-oncogene induction by mitogens, such as phorbol 12-myristate 13-acetate and serum. Suppression of c-fos was achieved with doses of RA as low as 10(-10) M and required pretreatment of cells with RA for a minimum of 2 h. In contrast, inhibition of c-jun required pretreatment for greater than 16 h with at least 10(-8) M RA and coincided with the observed decrease in cell growth. RA blocked c-fos induction by inhibiting transcription. This inhibition of transcription occurs through the serum response element (SRE), since the SRE alone was sufficient to confer down-regulation by RA to a minimal c-fos promoter construct. Thus, the SRE plays a critical role in the suppression of c-fos transcription by RA.

Animals↗
PubMed1992

Autocrine secretion--10 years later.

The concept of autocrine secretion, its subsequent modifications, its application for understanding pathogenesis of disease, and its potential for developing new approaches to prevention and treatment are reviewed. Peptide growth factors (cytokines) act as local autocrine and paracrine mediators of tissue homeostasis. Many diseases, including cancer, atherosclerosis, rheumatoid arthritis, and other fibrotic diseases characterized by chronic inflammation, are associated with aberrant expression and cellular coordination of the homeostatic action of these regulatory molecules. Modern biotechnology and pharmacology offer unique opportunities for the therapeutic prevention and treatment of these molecular and cellular lesions, using either cytokines or other agents that modify their synthesis and activity.

Amino Acid Sequence↗
PubMed1992

Enhanced expression of transforming growth factor beta 1 in the rat brain after a localized cerebral injury.

It is becoming clear that transforming growth factor beta (TGF beta) may be a key factor regulating inflammatory and tissue specific wound responses. Because the formation of a glial-collagen scar at CNS lesion sites is thought to contribute to the pathology associated with penetrating CNS injuries, and because in the periphery TGF beta 1 stimulates fibroblast deposition of scar tissue, we used in situ hybridization and immunohistochemistry to investigate the effect of a defined cerebral lesion on the local expression of TGF beta 1. Induction of TGF beta 1 mRNA and protein is relatively diffuse in the neuropile around the margins of the lesion at 1, 2 and 3 days, but becomes localized to the region of the glial scar at 7 and 14 days. The signal intensity for TGF beta 1 mRNA and protein is maximal between 2 and 3 days and decreases between 7 and 14 days after lesion. The predominant cell types in the neuropile localizing TGF beta 1 mRNA and protein have the morphological characteristics of astrocytes, although macrophages are also detected. An induction of TGF beta 1 mRNA was also observed in endothelial cells of the meninges, hippocampal fissure and choroid plexus, at 2 and 3 days. However, this is dramatically reduced by 7 days and has disappeared by 14 days. These results suggest a role for TGF beta 1, not only in inflammation, but also in the tissue-specific glial scar formation that occurs in the CNS. Furthermore, they suggest a potential therapeutic use of TGF beta 1 antagonists in the CNS to help limit the pathogenesis associated with matrix deposition in the wound.

Animals↗
PubMed1992

Induction of transforming growth factor beta 1 in human breast cancer in vivo following tamoxifen treatment.

We have investigated the ability of tamoxifen to regulate members of the transforming growth factor beta (TGF-beta) family in human breast cancers in vivo. Using immunohistochemical techniques, we find that 3 months of tamoxifen treatment causes a consistent induction of extracellular TGF-beta 1 in breast cancer biopsies, compared with matched pretreatment samples from the same patient. The induced TGF-beta is localized between and around stromal fibroblasts and appears to be derived from these cells. Lower levels of TGF-beta 1,-beta 2, and -beta 3 seen in epithelial cells were not altered by tamoxifen treatment. The increased stromal staining of TGF-beta 1 occurred in estrogen receptor-negative as well as estrogen receptor-positive tumors. These results provide in vivo evidence for a novel, estrogen receptor-independent mechanism of action for tamoxifen, involving the stromal induction of a potent growth inhibitor for epithelial cells.

Biomarkers, Tumor↗
PubMed1992

Identification of a structural domain that distinguishes the actions of the type 1 and 2 isoforms of transforming growth factor beta on endothelial cells.

A chimeric transforming growth factor beta (TGF-beta) molecule has been synthesized to map the amino acids responsible for the substantially greater activity of TGF-beta 1 than TGF-beta 2 on growth and migration of endothelial cells. This chimera consists of a dimer of a monomeric unit composed of amino acids 1-39 of TGF-beta 2, 40-82 of TGF-beta 1, and 83-112 of TGF-beta 2. Structural identity of the purified recombinant protein has been confirmed by immunoblotting and NH2-terminal sequencing. The biological potency of the TGF-beta 2-1-2 chimera was equal to that of TGF-beta 1 in inhibition of growth of both fetal bovine heart endothelial cells and rat epididymal fat pad microvascular endothelial cells. Similarly, the TGF-beta 2-1-2 chimera was nearly equivalent to TGF-beta 1 and at least 10-fold more active than TGF-beta 2 in inhibiting migration of bovine aortic endothelial cells. These results identify the sequence between amino acids 40-82 as an important region within TGF-beta that functions to specify a TGF-beta 1- or TGF-beta 2-like activity.

Animals↗
PubMed1992

Post-transcriptional regulation of the human transforming growth factor-beta 1 gene.

Since many lines of evidence suggest that expression of the transforming growth factor-beta 1 (TGF-beta 1) gene may be regulated post-transcriptionally, we examined the effect of the 5'-untranslated region (UTR) of this gene on TGF-beta 1 expression. For this purpose, fragments of the 840-nucleotide highly GC-rich TGF-beta 1 5'-UTR were inserted into the 5'-UTR of the structural gene for human growth hormone driven by the simian virus 40 early promoter. A portion of the 5'-UTR of TGF-beta 1 mRNA spanning the sequences from +11 to +147 was shown to inhibit growth hormone expression by as much as 22-fold. This effect was cell-specific; growth hormone production was inhibited in PC-3 human prostate adenocarcinoma and A-549 human lung adenocarcinoma cells, while no effect was seen in rat pheochromocytoma PC12 cells, which show efficient translation of endogenous TGF-beta 1 mRNA. Computer analysis showed that this region of the 5'-UTR contained a stable secondary stem-loop structure spanning sequences +49 to +76. This stem-loop region alone is sufficient to inhibit expression of the growth hormone gene, suggesting that it plays an important role in post-transcriptional regulation of TGF-beta 1 gene expression.

Animals↗
PubMed1992

Inhibition of growth by transforming growth factor-beta following fusion of two nonresponsive human carcinoma cell lines. Implication of the type II receptor in growth inhibitory responses.

Loss of growth regulation by transforming growth factor-beta (TGF-beta) may be an important step in carcinogenesis. We have used a cell fusion system to show that inhibition of growth by TGF-beta can be restored to carcinoma cell lines that are unresponsive to the inhibitory effects of TGF-beta. In a previous study, the EJ bladder carcinoma line was fused to the SW480 colon adenocarcinoma line and found to produce nontumorigenic hybrid cells along with one hybrid cell clone of low tumorigenicity. Here we show that the capacity of the nontumorigenic hybrid cells to respond to either TGF-beta 1 or TGF-beta 2 has been restored, while the parental or tumorigenic hybrid cells show little or no inhibition of growth following TGF-beta treatment. Cross-linking analyses with labeled TGF-beta 1 demonstrated much higher levels of the type II (85 kDa) receptor in the hybrid cells compared with the parental tumor lines. Both the parental and tumorigenic hybrid cell lines were capable of responding to TGF-beta as evidenced by increased levels of mRNA for fibronectin, type IV collagenase, and plasminogen activator inhibitor after treatment with TGF-beta 1. These results suggest that the type II receptor is necessary for mediating the effects of TGF-beta on inhibition of growth but not on gene activation of the hybrid cells.

Blotting, Northern↗
PubMed1992

Suppressor T cells generated by oral tolerization to myelin basic protein suppress both in vitro and in vivo immune responses by the release of transforming growth factor beta after antigen-specific triggering.

Oral administration of myelin basic protein (MBP) is an effective way of suppressing experimental autoimmune encephalomyelitis (EAE). We have previously shown that such suppression is mediated by CD8+ T cells, which adoptively transfer protection and suppress immune responses in vitro. In the present study we have found that modulator cells from animals orally tolerized to MBP produce a suppressor factor upon stimulation with MBP in vitro that is specifically inhibited by anti-transforming growth factor beta (TGF-beta) neutralizing antibodies. No effect was observed with antibodies to gamma interferon, tumor necrosis factor alpha/beta, or indomethacin. In addition, the active form of the type 1 isoform of TGF-beta 1 (TGF-beta 1) can be directly demonstrated in the supernatants of cells from animals orally tolerized to MBP or ovalbumin after antigen stimulation in vitro. Antiserum specific for TGF-beta 1 administered in vivo abrogated the protective effect of oral tolerization to MBP in EAE. Furthermore, injection of anti-TGF-beta 1 serum to nontolerized EAE animals resulted in an increase in severity and duration of disease. These results suggest that immunomodulation of EAE induced by oral tolerization to MBP and natural recovery mechanisms use a common immunoregulatory pathway that is dependent on TGF-beta 1. Implications of such an association are of therapeutic relevance to human autoimmune diseases and may help to explain one of the mechanisms involved in the mediation of active suppression by T cells.

Administration, Oral↗
PubMed1992

Pattern of expression of transforming growth factor-beta 4 mRNA and protein in the developing chicken embryo.

Expression of TGF-beta 4 mRNA and protein was studied in the developing chicken embryo using specific cDNA probes and antibodies for chicken TGF-beta 4. Expression of TGF-beta 4 mRNA was detected by day 4 of incubation (Hamburger and Hamilton stage 22, E4) by RNA Northern blot analysis and increased with developmental age until day 12 of incubation (stage 38, E12) where it was detected in every embryonic tissue examined, with expression being highest in smooth muscle and lowest in the kidney. The steady-state level of expression of TGF-beta 4 mRNA remained relatively constant in most embryonic tissues through day 19 (stage 45, E19). In situ hybridization analysis detected TGF-beta 4 mRNA as early as the "definitive primitive streak" stage (stage 4); during neurulation (stage 10), TGF-beta 4 mRNA was detected in all three germ layers, including neuroectoderm. Following neurulation, TGF-beta 4 mRNA was detected in the neural tube, notochord, ectoderm, endoderm, sclerotome, and myotome, but not dermotome at stage 16. By day 6 of incubation (stage 29, E6), TGF-beta 4 mRNA was localized in several tissues including heart, lung, and gizzard. Immunohistochemical staining analysis also showed expression of TGF-beta 4 protein in all three germ layers as early as stage 4 in various cell types in qualitatively similar locations as TGF-beta 4 mRNA. These results suggest that TGF-beta 4 may play an important role in the development of many tissues in the chicken.

Animals↗
PubMed1992

Differential regulation of the expression of transforming growth factor-beta mRNAs by growth factors and retinoic acid in chicken embryo chondrocytes, myocytes, and fibroblasts.

Transforming growth factor-beta (TGF-beta) autoregulates its expression in several mammalian cell types. We now report that addition of TGF-beta s 1, 2, and 3 to primary chicken embryo cells differentially affects expression of the messenger RNAs for the different TGF-beta isoforms depending on the cell type. In cultured sternal chondrocytes, addition of TGF-beta s 1, 2, or 3 results in an increase in the steady-state levels of the messenger RNAs for TGF-beta s 2 and 3, but does not change expression of TGF-beta 4 mRNA. In contrast, in cultured cardiac myocytes, addition of TGF-beta s 1, 2, or 3 results in an increase in expression of TGF-beta s 3 and 4 mRNAs, but does not change expression of TGF-beta 2 mRNA. Moreover, expression of TGF-beta s 2, 3, and 4 mRNAs is not affected by addition of any of the TGF-beta s to fibroblasts. Addition of platelet-derived growth factor (PDGF), epidermal growth factor (EGF), or interleukin-1 (IL-1) to these chicken cells also has differential effects on expression of the different TGF-beta mRNAs depending on the cell type. Retinoic acid also has contrasting effects on chondrocytes and myocytes either increasing or decreasing, respectively, expression of TGF-beta s 2 and 3 mRNAs and TGF-beta 2 protein. Our results indicate a complex pattern of regulation of the different TGF-beta genes by themselves as well as by PDGF, EGF, IL-1, dexamethasone, TPA, and retinoic acid in chicken embryo cells.

Animals↗
PubMed1992

Differential expression of the TGF-beta isoforms in embryogenesis suggests specific roles in developing and adult tissues.

The TGF-beta's are multifunctional, pleiotropic molecules with major effects in control of cellular migration, cellular proliferation, and elaboration of extracellular matrix. Thus far, five distinct isoforms of TGF-beta have been described, each approximately 65-85% homologous and containing the characteristic 9 positionally conserved cysteine residues. Although the actions of the activated mature forms of the different isoforms on cells are qualitatively similar in most cases, there are a few examples of distinct activities. For example, TGF-beta's 1 and 3, but not TGF-beta 2, inhibit the growth of large vessel endothelial cells, and TGF-beta's 2 and 3, but not TGF-beta 1, inhibit the survival of cultured embryonic chick ciliary ganglionic neurons. In addition, selective targeting of the latent forms of the TGF-beta's is suggested by the observation that latent TGF-beta 2 is the prominent isoform found in body fluids such as amniotic fluid, breast milk, and the aqueous and vitreous humor of the eye; it is noteworthy in this regard that TGF-beta 2 is unique among various isoforms in that it lacks a RGD integrin-binding sequence in its precursor. The most dramatic differences in the TGF-beta isoforms are seen at the level of expression, where there is now a wealth of data demonstrating both spatially and temporally distinct expression of both the mRNAs and proteins in developing tissues, regenerating tissues, and in pathologic responses.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Localization of transforming growth factor-beta isotypes in lesions of the human breast.

Transforming growth factor-beta s (TGF-beta) comprise a highly conserved family of multifunctional cell regulatory peptides that may play a role in a variety of pathologic processes. To date, five TGF-beta isotypes have been identified, three of these in mammalian systems. A number of cultured human breast carcinoma cell lines produce biologically inactive latent TGF-beta and are growth inhibited by activated TGF-beta; TGF-beta production is estrogen-influenced in some of these cell lines. To investigate the potential role of the TGF-beta isotypes in human breast disease, we localized TGF-beta s 1, 2, and 3 immunohistochemically in normal breast, fibrocystic change, epithelial hyperplasia, sclerosing adenosis, fibroadenoma, cystosarcoma phyllodes, and several carcinoma variants. Transforming growth factor-beta s 1, 2, and 3 were identified intracellularly in most active mammary epithelia, regardless of the lesion, including carcinoma; the associated stroma contained little or no intracellular TGF-beta. An antibody that recognizes an extracellular conformation of TGF-beta stained normal intralobular stroma and, more extensively, the stroma of active fibroadenomas and low-grade phyllodes tumors and the desmoplastic stroma of carcinomas. The results indicate the potential for paracrine and autocrine regulation of the mammary gland by TGF-beta and suggest an association between TGF-beta and abnormal stromal proliferations. Altered expression of TGF-beta s 1, 2, and 3 at the protein level in mammary epithelia appears not to be a major feature of most breast lesions, raising the possibility that altered cellular response to the TGF-beta already present may play a role in the development of breast disease.

Breast↗