Biomedical subjects

K Miyazono

Publications and source records attributed to K Miyazono.

At least 145 records · Page 8Linked to original sources

PubMed1995

Enhanced expression of transforming growth factor-beta and its type-I and type-II receptors in human glioblastoma.

Immunohistochemical studies of transforming growth factor-beta (TGF-beta) and its receptors have been carried out on 16 glioma tissues and compared with 5 cases of gliosis. Significantly higher expressions of TGF-beta I, as well as type-I and type-II TGF-beta receptors (T beta R-I and T beta R-II, respectively), were observed in advanced-malignant-glioma tissues when compared with non-tumorous gliosis. Immunoreactivities of TGF-beta and T beta R-I were localized in the cytoplasm of spindle-shaped tumor cells surrounding proliferating vessels or around areas of necrosis. The advancing edge of the tumor clusters frequently stained positive. Similar expression patterns were found for TGF-beta 2 and TGF-beta 3, whereas only weak or no expression was found for endoglin. In low-grade astrocytomas and in gliosis cases, the expression was moderate for T beta R-I and weak for TGF-beta and T beta R-II. In 3 examined human malignant glioma cell lines, clear immunostainings were detected for TGF-beta and its receptors. Ligand-induced heteromeric complexes of the receptors were formed in these cell lines, but the amount of the receptors was less than that of mink lung epithelial cells, which are sensitive target cells for TGF-beta. TGF-beta I showed no growth-inhibitory activity on any of these glioma cell lines. These results suggest that malignant gliomas produce TGF-beta and receptors, but are refractory to TGF-beta, implying dysregulation in the signalling pathway in the tumor cells. It is possible that the released TGF-beta acts on neighboring cells and affects stromal growth, angiogenesis, metastasis or immune surveillance in human glioma.

Brain Neoplasms↗

PubMed1995

A human keratinocyte cell line produces two autocrine growth inhibitors, transforming growth factor-beta and insulin-like growth factor binding protein-6, in a calcium- and cell density-dependent manner.

Two growth inhibitors were identified in culture medium conditioned by a human keratinocyte cell line, HaCat. TGF-beta was detected in media conditioned by growing or confluent HaCat cells, as well as in media conditioned at physiological (1 mM) or low (0.03 mM) Ca2+ concentrations. However, a considerable part of transforming growth factor beta (TGF-beta) in media conditioned at a physiological Ca2+ concentration was in active form, whereas most TGF-beta in media conditioned at a low Ca2+ concentration was latent. The other growth-inhibitory activity, which was detected only in media conditioned by confluent cells at a physiological Ca2+ concentration, was purified to homogeneity by a four-step procedure. The N-terminal amino acid sequence of the 33-kDa protein was identical with that of insulin-like growth factor binding protein-6 (IGFBP-6). Purified IGFBP-6 inhibited the growth of HaCat and Balb/MK keratinocyte cell lines, as well as Mv1Lu cells. The growth activity was also demonstrated by human recombinant IGFBP-6. In summary, HaCat cells secrete at least two possible autocrine growth inhibitors: TGF-beta which is secreted constitutively, but activated in a Ca(2+)-dependent manner, and IGFBP-6 which is secreted in a cell density- and Ca(2+)-dependent manner.

Amino Acid Sequence↗

PubMed1995

[Transforming growth factor-beta. An interesting candidate for clinical use].

Transforming growth factor-beta (TGF-beta) is the prototype of a large family of growth regulatory factors affecting the growth and differentiation of many cell types. Their action is mediated by complex formation of type I and type II receptors, both of which are serine/threonine kinases. TGF-beta inhibitors inhibit the growth of most cell types are potent immunosuppressive agents. They also stimulate the formation of connective tissue, thus promoting wound healing. TGF-beta and TGF-beta antagonists may have potential clinical uses in the treatment of various disorders.

Fibrosis↗

PubMed1995

The GS domain of the transforming growth factor-beta type I receptor is important in signal transduction.

Signal transduction by transforming growth factor-beta (TGF-beta) involves the formation of a heteromeric complex of two transmembrane serine/threonine kinase receptors, type I (T beta R-I) and type II (T beta R-II). In the region preceeding the kinase domain of T beta R-I there is a glycine- and serine-rich sequence, termed the GS domain, which has been shown to be phosphorylated by T beta R-II. In order to determine the importance of the serine residues in this domain, receptor mutants with one or more serine residues mutated were analyzed. All the mutants of T beta R-I were able to bind ligand and their kinase activity was not abolished by the mutations. The receptor mutants with single mutated serine residues mediated transcriptional responses to TGF-beta with similar efficiency as the wild type receptor, whereas those with two or three of the serine residues mutated showed only weak transcriptional responses. These results suggest that serine residues in the GS domain are important for signal transduction by T beta R-I; however, the signaling activity of T beta R-I does not depend on any particular serine residue in the GS domain, but rather on how many of the serine residues in the region are intact.

Activin Receptors, Type I↗

PubMed1995

A rat pituitary tumor cell line (GH3) expresses type I and type II receptors and other cell surface binding protein(s) for transforming growth factor-beta.

A rat pituitary tumor cell line (GH3) has been reported to express transforming growth factor-beta (TGF-beta) binding components of 70-74 kDa (ligand included), denoted TGF-beta type IV receptor. We investigated whether the type IV receptor corresponds to any of the recently cloned type I receptors for proteins in the TGF-beta super-family. TGF-beta type I receptor (T beta R-I) complexes of 69-72 kDa formed a heteromeric complex with T beta R-II in GH3 cells, as detected by immunoprecipitation. In addition, TGF-beta formed complexes of 72-74 kDa, which were different from T beta R-I and the other known type I receptors, and were not dependent on T beta R-II for binding. The GH3 cells were resistant to the growth inhibitory activity of TGF-beta, but a transcriptional response was activated by TGF-beta in this cell line, presumably through the T beta R-II and T beta R-I complex. These results indicate that GH3 cells have T beta R-I and T beta R-II and, in addition, other binding protein(s) which form 72-74-kDa complexes with TGF-beta; the function of the latter component(s) remains to be elucidated.

Activins↗

PubMed1995

Enhanced expression of type I receptors for bone morphogenetic proteins during bone formation.

Type I receptors for bone morphogenetic proteins (BMPs), i.e., BMPR-IA and BMPR-IB, are transmembrane serine/threonine kinases, that bind osteogenic protein-1 (OP-1, also termed BMP-7) and BMP-4. Using antibodies specific to BMPR-IA and -IB, we have studied the expression of BMP type I receptors in the bone formation process during embryonic development and fracture healing. In the mouse embryo, both BMPR-IA and -IB were expressed in condensing mesenchymal cells at 13.5 days post coitum (p.c.). At 15.5 days p.c., expression of BMPR-IB, but not of BMPR-IA, was observed in the cells in perichondrium of developing cartilage. At 17.5 and 19.5 days p.c., expression of both receptors was observed in chondrocytes and in osteoblasts. In normal rat adult bone, expression of BMPR-IA, but not of BMPR-IB, was observed in osteoblasts in the periosteum. Three days after the femoral fracture, expression of BMPR-IA and -IB was up-regulated in cells at the proliferating osteogenic layer of the periosteum. On day 7, both receptors were found in fibroblast-like spindle cells and chondrocytes in the endochondral ossification sites, and osteoblasts in the newly formed trabecular bone. Expression of BMPR-IA was higher than that BMPR-IB in osteogenic layer on day 3 and in osteoblasts in the trabecular bone on day 7. On day 14, expression of BMP type I receptors was observed at similar sites, albeit with lower expression levels than were observed on day 7. The present data suggest that expression of BMP type I receptors is up-regulated during bone formation, and that they may play important roles in bone morphogenesis.

Animals↗

PubMed1995

Localization of transforming growth factor-beta type I and type II receptors in mouse development.

We have investigated the localization pattern of the transforming growth factor-beta (TGF-beta) receptors type I (T beta R-I) and type II (T beta R-II) during mouse organogenesis by immunohistochemical analysis. Staining of both receptors was found in many developing organs, e.g., bone, teeth, Meckel's cartilage, and neural tissues, where the expression of their ligands has been previously reported. During the investigated stages, expression of T beta R-I was more ubiquitous than that of T beta R-II. T beta R-II preferentially localized in the undifferentiated mesenchymal cells which subsequently differentiated into bone. There was no staining of T beta R-II in the central nervous system, while intense T beta R-I staining was found specifically in nervous tissues. Expression of T beta R-I and T beta R-II was mostly coincident with that of their ligands, suggesting that TGF-beta s act as multiple mediators during organogenesis. In addition, colocalization of both receptors in the epithelia of the tooth bud and submandibular gland, which were actively invaginating into the mesenchyme, leads us to speculate that both receptors may be necessary for dynamic epithelial morphogenesis.

Activin Receptors, Type I↗

PubMed1995

Localization of transforming growth factor beta 1 and its latent binding protein in human chronic pancreatitis.

BACKGROUND/AIMS: Transforming growth factor beta 1 (TGF-beta 1) is thought to be the mediator of fibrosis in liver, glomerular, and pulmonary fibrosis. This study investigated the expression of TGF-beta 1 precursor (beta 1 latency-associated peptide), latent TGF-beta 1-binding protein (LTBP), and TGF-beta 1 messenger RNA (mRNA) in chronic pancreatitis. METHODS: Beta 1 latency-associated peptide and LTBP expression were studied by immunohistochemistry, and TGF-beta 1 mRNA expression was studied by reverse-transcription polymerase chain reaction analysis in normal pancreatic parenchyma and in tissues from patients with chronic pancreatitis of different etiologies. RESULTS: In normal specimens, TGF-beta 1 precursor was present in islet cells and in a few ductal and acinar cells but not in periductal connective tissue. No immunoreactivity for LTBP was detected. In chronic pancreatitis, TGF-beta 1 precursor was detected mainly in mononuclear cells located in the fibrotic areas and also in ducts damaged by fibrosis, more frequently in calcifying chronic pancreatitis. LTBP was present predominantly in mononuclear cells and in the extracellular matrix around them. TGF-beta 1 mRNA was either not expressed or was faintly expressed in normal tissue, whereas intense signals were detected in chronic pancreatitis. CONCLUSIONS: The findings suggest the involvement of TGF-beta 1 in the development of fibrosis in chronic pancreatitis and the important role of inflammatory cells.

Adult↗

PubMed1995

Axotomy of rat facial nerve induces TGF-beta and latent TGF-beta binding protein.

Transforming growth factor-beta (TGF-beta) has been found to be abundantly and specifically expressed in the nervous system. However, the function of TGF-beta during nerve regeneration is still unknown. We have examined the expression of TGF-beta isoforms and the latent TGF-beta binding protein (LTBP) by immunohistochemistry in the rat facial nuclei after unilateral axotomy. An increased immunoreactivity for all the TGF-beta isoforms and the LTBP was observed in the facial nuclei of the injured side during the regeneration period examined until Day 24. These differences were tested statistically by nominal logistic regression analysis. When the intensity of the immunoreactivity in the injured side was compared to that of the contralateral side, significantly increasing differences were found for TGF-beta 2 (p < 0.003) and LTBP (p < 0.002). Strong immunostaining was detected in the neuronal perikarya and their axons. No clear immunoreactivity was seen in either microglia or astrocytes. The enhanced immunoreactivity was seen in the operated side already at Day 3, remaining at high level with some fluctuations until Day 12 or 24 after axotomy. These findings suggest that TGF-beta might play a functional role in the regeneration of motor neurons.

Amino Acid Sequence↗

PubMed1995

Expression of type I and type IB receptors for activin in midgestation mouse embryos suggests distinct functions in organogenesis.

Activins exert their effects by inducing heteromeric complexes of either of two different type I receptors, ActR-I or ActR-IB, and either of two type II receptors, ActR-II or ActR-IIB. We describe the cDNA cloning of the mouse homologue of human ActR-IB and analyze binding of radio-iodinated activin on type I/type II combinations of mouse receptors expressed from cDNA. We studied the distribution of ActR-I and ActR-IB mRNAs in postimplantation mouse embryos by in situ hybridization. In the 12.5-day postcoitum embryo, both mRNAs are found in the brain, spinal cord, some ganglia, vibrissae, lungs, body wall, stomach, gonads, ribs, limbs and shoulders. ActR-I mRNA, but not ActR-IB, is expressed in blood vessels, the heart, tongue, intervertebral discs and diaphragm. Conversely, only ActR-IB mRNA is detected in the olfactory region, eye, tooth primordium, esophagus, mesonephros, dorsal root ganglia and is strongly expressed in the spinal cord. Our results demonstrate similarities, but also differences and complementarities (mesenchymal versus epithelial expression) between the expression patterns of these type I receptors. Moreover, their expression patterns overlap with at least one of the type II activin receptors and/or one of activin subunits in some regions of the embryo, such as the brain, spinal cord, pituitary, whisker follicles, and the inner nuclear neuroblastic layer of the eye.

Activin Receptors↗

PubMed1995

Osteogenic protein-1 binds to activin type II receptors and induces certain activin-like effects.

Proteins in the TGF-beta superfamily transduce their effects through binding to type I and type II serine/threonine kinase receptors. Osteogenic protein-1 (OP-1, also known as bone morphogenetic protein-7 or BMP-7), a member of the TGF-beta superfamily which belongs to the BMP subfamily, was found to bind activin receptor type I (ActR-I), and BMP receptors type IA (BMPR-IA) and type IB (BMPR-IB) in the presence of activin receptors type II (ActR-II) and type IIB (ActR-IIB). The binding affinity of OP-1 to ActR-II was two- to threefold lower than that of activin A. A transcriptional activation signal was transduced after binding of OP-1 to the complex of ActR-I and ActR-II, or that of BMPR-IB and ActR-II. These results indicate that ActR-II can act as a functional type II receptor for OP-1, as well as for activins. Some of the known biological effects of activin were observed for OP-1, including growth inhibition and erythroid differentiation induction. Compared to activin, OP-1 was shown to be a poor inducer of mesoderm in Xenopus embryos. Moreover, follistatin, an inhibitor of activins, was found to inhibit the effects of OP-1, if added at a 10-fold excess. However, certain effects of activin, like induction of follicle stimulating hormone secretion in rat pituitary cells were not observed for OP-1. OP-1 has overlapping binding specificities with activins, and shares certain but not all of the functional effects of activins. Thus, OP-1 may have broader effects in vivo than hitherto recognized.

Activin Receptors↗

PubMed1995

Dual role for the latent transforming growth factor-beta binding protein in storage of latent TGF-beta in the extracellular matrix and as a structural matrix protein.

The role of the latent TGF-beta binding protein (LTBP) is unclear. In cultures of fetal rat calvarial cells, which form mineralized bonelike nodules, both LTBP and the TGF-beta 1 precursor localized to large fibrillar structures in the extracellular matrix. The appearance of these fibrillar structures preceded the appearance of type I collagen fibers. Plasmin treatment abolished the fibrillar staining pattern for LTBP and released a complex containing both LTBP and TGF-beta. Antibodies and antisense oligonucleotides against LTBP inhibited the formation of mineralized bonelike nodules in long-term fetal rat calvarial cultures. Immunohistochemistry of fetal and adult rat bone confirmed a fibrillar staining pattern for LTBP in vivo. These findings, together with the known homology of LTBP to the fibrillin family of proteins, suggest a novel function for LTBP, in addition to its role in matrix storage of latent TGF-beta, as a structural matrix protein that may play a role in bone formation.

Animals↗

PubMed1995

Distinct spatial and temporal expression patterns of two type I receptors for bone morphogenetic proteins during mouse embryogenesis.

Bone morphogenetic proteins (BMPs) are multifunctional proteins structurally related to transforming growth factor-beta (TGF beta) and activin that can induce cartilage and bone growth in vivo. Members of the TGF beta superfamily exert their biological effects via heteromeric serine/threonine kinase complexes of type I and type II receptors. We previously obtained six different type I receptors, termed activin receptor-like kinase-1 (ALK-1) to -6. ALK-5 is a TGF beta type I receptor, ALK-2 and ALK-4 are activin type I receptors, and ALK-3 and ALK-6 are type I receptors for osteogenic protein-1 (OP-1)/bone morphogenetic protein-7 (BMP-7) and BMP-4. Here we report the complementary DNA cloning of the mouse homolog of ALK-3, which is highly conserved between mouse and man. ALK-3 messenger RNA (mRNA) is ubiquitously expressed in various adult mouse tissues, whereas ALK-6 mRNA is only found in brain and lung. The distribution of ALK-3 and ALK-6 mRNA in the postimplantation mouse embryo [6.5-15.5 days postcoitum (pc)] was studied by in situ hybridization. ALK-3 was nearly ubiquitously expressed throughout these stages of development, but was notably absent in the liver. In contrast, ALK-6 showed a more restricted expression pattern. ALK-6 mRNA was absent in early postimplantation embryos, was detected first in 9.5 days pc embryos, and persisted until 15.5 days pc. In midgestation embryos, ALK-6 transcripts were detected in mesenchymal precartilage condensations, premuscle masses, blood vessels, central nervous system, parts of the developing ear and eye, and epithelium. The expression in sites of developing cartilage and bone supports the idea that ALK-3 and -6 are receptors for BMPs in vivo. In addition, the expression of these genes in many soft tissues suggests broader functions for BMPs in embryogenesis.

Activin Receptors↗

PubMed1995

Matrix-associated latent TGF-beta with latent TGF-beta binding protein in the progressive process in adriamycin-induced nephropathy.

BACKGROUND: A progressive increase in latent transforming growth factor-beta (TGF-beta) secretion from diseased tissue was revealed in our previous work using adriamycin (ADR)-nephropathy (Kidney Int 45:525-36, 1994). Latent TGF-beta is composed of mature TGF-beta and latency-associated peptide (LAP) with or without latent TGF-beta-binding protein (LTBP). LTBP has been reported to contribute to either matrix-association or activation of latent TGF-beta. LTBP also seems to play a key role in the renal lesions of this model. The present study was designed to show the secretion of latent TGF-beta with LTBP and the location of LTBP in renal tissue in ADR-nephropathy. EXPERIMENTAL DESIGN: The renal cortical tissue specimens were sampled at Weeks 4, 8, and 16 after the injection of ADR or saline (control) for cortical tissue culture and immunohistology. TGF-beta in the conditioned medium was assayed by immunoprecipitation and bioassay using mink lung epithelial cells. An immunohistochemical study was performed to examine the localization of LTBP, ED-1-positive macrophages, and extracellular matrix proteins including laminin, fibronectin, and collagen type I and type III. RESULTS: A TGF-beta bioassay revealed a progressive increase in latent TGF-beta secretion from the cortex of diseased kidney. Free LTBP and LTBP-LAP complex with mature TGF-beta were immunoprecipitated by anti-LTBP Ab from the cortical culture medium. An immunohistochemical study using anti-LTBP Ab demonstrated that LTBP localization was restricted to the glomeruli and the arterioles in the control cortex. In the ADR rats at Week 4, a faint deposition of LTBP was observed in the interstitium around the glomeruli. At Week 8 or 16, LTBP was accumulated in the sclerosing glomeruli or fibrous interstitium, where ECM proteins and infiltrating ED-1-positive macrophages were intensely located. CONCLUSIONS: Our results indicated that latent TGF-beta with LTBP was localized in association with the extracellular matrix in the sclerotic and fibrotic tissue in this model. Matrix-associated latent TGF-beta with LTBP may thus play an important role in the progressive process of glomerulosclerosis and interstitial fibrosis in ADR-nephropathy.

Animals↗

PubMed1995

Expression and prognostic significance of TGF-beta isotypes, latent TGF-beta 1 binding protein, TGF-beta type I and type II receptors, and endoglin in normal ovary and ovarian neoplasms.

BACKGROUND: The etiology and biology of ovarian carcinogenesis is largely unknown. Recent results have indicated prognostic significance of growth factors in this malignancy. TGF-beta is a widely distributed growth factor with multifactorial effects in in vitro systems. Studies on the in vivo expression pattern of TGF-beta and its receptors might help us to understand its biologic significance in this malignancy. EXPERIMENTAL DESIGN: Tissue samples of normal ovary and benign as well as malignant ovarian neoplasms were examined for expression of transforming growth factor (TGF)-beta 1, -beta 2, and -beta 3, the latent TGF-beta-binding protein (LTBP), TGF-beta type I (T beta R-II) receptors and endoglin by immunohistochemistry and in situ hybridization. Furthermore, the results of the immunohistochemical analysis were compared with patient survival. RESULTS: Expression of all ligands was significantly increased in tumor cells compared with the normal epithelial cells. In contrast, LTBP immunoreactivity was detected significantly more often in normal epithelium than in tumor cells. T beta R-I and T beta R-II as well as endoglin were found in tumor tissues and normal ovary without any difference among the groups. In the blood vessels of malignant tumors, significantly increased TGF-beta 1 reactivity and decreased TGF-beta 2 reactivity were found when they were compared with those of normal ovaries and benign tumors. Patients with malignant tumors expressing TGF-beta 1, T beta R-I, or endoglin in blood vessels demonstrated longer survival than those having negatively stained tumors. In contrast, positive endoglin staining in tumor cells correlated with decreased survival even in advanced disease or in patients having residual tumor bulk after surgery. CONCLUSIONS: The differential expression of TGF-beta ligand and the significant correlations between expression of ligands or receptors and patient survival indicate involvement of the TGF-beta system in ovarian tumor development.

Antigens, CD↗

PubMed1994

Identification and characterization of LTBP-2, a novel latent transforming growth factor-beta-binding protein.

Latent transforming growth factor-beta (TGF-beta)-binding protein (LTBP) is a component of the latent TGF-beta complex in human platelets. LTBP is composed of two different cysteine-rich repeat sequences, i.e. epidermal growth factor (EGF)-like repeats and a repeat containing 8 cysteine residues. The overall structure of LTBP is similar to those of the microfibrillar proteins fibrillin-1 and fibrillin-2. Here we report the identification of a novel protein termed LTBP-2, which is structurally related to LTBP. cDNA for LTBP-2 was obtained from human foreskin fibroblast cDNA libraries using a fragment of the LTBP cDNA as a probe. LTBP-2 is composed of 20 EGF-like repeats and four copies of the 8-cysteine repeat. The amino acid sequence of LTBP-2 is 41% identical to that of LTBP and 25% identical to that of fibrillin-1. LTBP-2 is synthesized as a 240-kDa protein by human foreskin fibroblasts and also by COS cells transfected with the isolated LTBP-2 cDNA. Similar to LTBP, a considerable part of LTBP-2 was found to be associated with extracellular matrix. Co-transfection of cDNAs for LTBP-2 and TGF-beta 1 revealed that LTBP-2 forms a high molecular weight complex with the TGF-beta 1 precursor. The LTBP-2 gene was assigned to chromosome 14q24. These results indicate that different forms of latent TGF-beta complexes occur and suggest that the different associated proteins may function to target the complexes to specific sites.

Adaptor Proteins, Signal Transducing↗

PubMed1994

Distinct roles of the intracellular domains of transforming growth factor-beta type I and type II receptors in signal transduction.

Transforming growth factor-beta (TGF-beta) transduces signals through binding to type I (T beta R-I) and type II (T beta R-II) serine/threonine kinase receptors. T beta R-I requires T beta R-II for ligand binding, whereas T beta R-II requires T beta R-I for signaling. We generated two different chimeric TGF-beta receptors, i.e. T beta R-1.2 containing the extracellular domain of T beta R-I and the intracellular domain of T beta R-II, and T beta R-2.1 containing the extracellular domain of T beta R-II and the intracellular domain of T beta R-I. T beta R-2.1 bound 125I-TGF-beta 1 alone, whereas T beta R-1.2 bound the ligand only in the presence of T beta R-II or T beta R-2.1. When transfected into a mutant mink lung epithelial cell line that lacks functional T beta R-II, T beta R-II cDNA, but not T beta R-2.1 cDNA, restored the responsiveness to TGF-beta 1 with regard to transcriptional activation of plasminogen activator inhibitor-1 gene promoter and 12-O-tetradecanoylphorbol-13-acetate-responsive elements. In a mutant mink lung epithelial cell line lacking T beta R-I, T beta R-I cDNA stimulated promoter activity, but the T beta R-1.2 cDNA did not. T beta R-2.1 formed an oligomer with T beta R-II when transfected into COS cells, but the complex did not transduce the signal after ligand stimulation. On the other hand, co-transfection of T beta R-1.2 and T beta R-2.1 cDNAs restored the responsiveness to TGF-beta 1. These results indicate that an interaction between the intracellular regions of T beta R-I and T beta R-II, triggered by ligand binding to the extracellular domains of these receptors, leads to efficient signal transduction by TGF-beta.

Animals↗

PubMed1994

Formation of hetero-oligomeric complexes of type I and type II receptors for transforming growth factor-beta.

Transforming growth factor-beta (TGF-beta) transduces signals through a heteromeric complex of type I (T beta R-I) and type II (T beta R-II) TGF-beta receptors. To determine the stoichiometry of this complex we used analysis by affinity labeling with 125I-TGF-beta 1 and covalent cross-linking with disuccinimidyl suberate and immunoprecipitation using T beta R-I- or T beta R-II-specific antisera, followed by two-dimensional gel electrophoresis under nonreducing (first dimension) and reducing (second dimension) conditions. Dimers composed of T beta R-I and/or T beta R-II were observed on mink lung epithelial cells as well as COS-1 cells transfected with T beta R-I and T beta R-II cDNAs. Homodimers of T beta R-I could be demonstrated on these cells after dissociation of T beta R-II by sodium dodecyl sulfate treatment. On the cell surface of mink lung epithelial cell mutants that express only T beta R-II and do not respond to TGF-beta, and on COS-1 cells transfected with only T beta R-II, only homodimers of T beta R-II were seen. The facts that T beta R-I does not bind TGF-beta in the absence of T beta R-II and that T beta R-II forms a signaling complex with T beta R-I, together with our observations that homodimers of T beta R-I and T beta R-II are seen on responsive cells, support the notion that TGF-beta induces the formation of hetero-oligomeric receptor complexes, most likely a heterotetramer containing two molecules each of T beta R-I and T beta R-II.

Affinity Labels↗