Identification of the type-B receptor for platelet-derived growth factor in human embryonal carcinoma cells

The human embryonal carcinoma (EC) cell line Tera 2 clone 13 (T2/13) can be induced to differentiate in vitro into neuroectodermal cell types with retinoic acid. Undifferentiated cells are characterized by rapid proliferation, whereas differentiated cells show a prolonged generation time, have a limited life span, and possess new cell-surface markers. In the present study we establish that both differentiated and undifferentiated T2/13 cells express the type-B platelet-derived growth factor (PDGF) receptor mRNA and bind PDGF-BB with high affinity. Differentiation causes a three-fold increase in receptor number per cell and leaves the affinity of the receptors unaffected. These data are the first to describe expression of this receptor in EC cells. The biosynthesis and degradation of this receptor were studied in undifferentiated as well as in differentiated T2/13 cells using an anti-type-B receptor antibody. These experiments revealed that high concentrations of recombinant PDGF-AA did not accelerate receptor metabolism in both cell types. In contrast, human PDGF or recombinant PDGF-BB added to the culture dishes readily increased receptor degradation. These results demonstrate that T2/13 cells express functional type-B PDGF receptors and suggest that cells responsive to PDGF might be present during mammalian development before the onset of mesoderm formation.     

Platelet-derived growth factor receptor beta regulates migration and DNA synthesis in metanephric mesenchymal cells

Platelet-derived growth factor (PDGF) B-chain and PDGF receptor beta (PDGFR beta) are essential for glomerulogenesis. Mice deficient in PDGF B-chain or PDGFR beta exhibit an abnormal glomerular phenotype characterized by total lack of mesangial cells. In this study, we localized PDGFR beta in the developing rat kidney and explored the biological effects of PDGF in metanephric mesenchymal cells in an attempt to determine the mechanism by which PDGF regulates mesangial cell development. Immunohistochemical and in situ hybridization studies of rat embryonic kidneys reveal that PDGFR beta localizes to undifferentiated metanephric mesenchyme and is later expressed in the cleft of the comma-shaped and S-shaped bodies and in more mature glomeruli in a mesangial distribution. We also isolated and characterized cells from rat metanephric mesenchyme. Metanephric mesenchymal cells express vimentin and alpha-smooth muscle actin but not cytokeratin. These cells also express functional PDGFR beta, as demonstrated by autophosphorylation of the receptor as well as activation of phosphatidylinositol 3 kinase in response to PDGF B-chain homodimer. PDGF B-chain also induces migration and proliferation of metanephric mesenchymal cells. Taken together with the fact that PDGF B-chain is expressed in the glomerular epithelium and mesangial area, as demonstrated in the human embryonic kidney, we suggest that PDGF B-chain acts in a paracrine fashion to stimulate the migration and proliferation of mesangial cell precursors from undifferentiated metanephric mesenchyme to the mesangial area. PDGF B-chain also likely stimulates proliferation of mesangial cell precursors in an autocrine fashion once these cells migrate to the glomerular tuft.     

Platelet-derived growth factor and its role in health and disease

Platelet-derived growth factor (PDGF) was first discovered in platelets because they are the principal source of mitogenic activity in whole blood serum for mesenchymal cells in culture. PDGF is ubiquitous in that it can be formed by a large number of normal cells as well as many varieties of transformed cells. However, its expression and biological activity appear to be controlled at a number of different levels. The molecule consists of two peptide chains (termed 'A' and 'B') and is found as one of at least three possible isoforms, (AB, AA or BB). Each of these isoforms binds to a high-affinity cell-surface receptor that is composed of two different subunits, each of which has specificity for one or the other of the peptide chains of PDGF. The two receptor subunits are present in differing amounts on different cell types, and therefore the capacity of the different isoforms of PDGF to induce mitogenesis depends on the specific PDGF isoform and the relative numbers of receptor subunits present on the responding cell. In addition to inducing cell replication, PDGF elicits a number of intracellular signals related to mitogenesis, is chemotactic, is a vasoconstrictor, activates leukocytes, and modulates extracellular matrix turnover. This growth factor is probably involved in a number of biologically important events including wound repair, embryogenesis and development, and inflammation, leading to fibrosis, atherosclerosis and neoplasia.     

PDGF induces c-myc mRNA expression in MG-63 human osteosarcoma cells but does not stimulate cell replication

The platelet-derived growth factor (PDGF) modulated growth response of the MG-63 human osteosarcoma cell line, which neither expresses c-sis mRNA nor secretes a PDGF analogue, was characterized. Scatchard analysis demonstrated that the MG-63 cells have 23,000 receptors per cell with a Kd of 5 X 10(-11) M. The receptor became phosphorylated, in a PDGF concentration-dependent manner, when 32P-orthophosphate-labeled cells were treated with PDGF for 3 h at 4 degrees C. The phosphorylated receptor was identified by autoradiography and gel electrophoresis after isolation of the 32P-labeled receptor using a solid-phase monoclonal antibody directed against phosphotyrosine. Binding of the receptor to the antibody was inhibited by 5 mM phenyl phosphate, further suggesting that PDGF stimulated tyrosine-specific receptor autophosphorylation. In addition, treatment of MG-63 cells with PDGF for 3 h at 37 degrees C induced a 7.5-fold increase in c-myc mRNA accumulation as analyzed on Northern gels. However, MG-63 cells grew equally well in either serum-(which contains PDGF) or plasma-(which does not) supplemented medium. Furthermore, PDGF did not stimulate DNA synthesis in growth arrested MG-63 cells, nor did it potentiate DNA synthesis modulated by somatomedin C. Thus MG-63 cells are a naturally occurring cell variant in which PDGF stimulates c-myc expression but does not modulate mitogenesis.     

PDGF-D is a specific, protease-activated ligand for the PDGF beta-receptor

The term 'platelet-derived growth factor' (PDGF) refers to a family of disulphide-bonded dimeric isoforms that are important for growth, survival and function in several types of connective tissue cell. So far, three different PDGF chains have been identified - the classical PDGF-A and PDGF-B and the recently identified PDGF-C. PDGF isoforms (PDGF-AA, AB, BB and CC) exert their cellular effects by differential binding to two receptor tyrosine kinases. The PDGF alpha-receptor (PDGFR-alpha) binds to all three PDGF chains, whereas the beta-receptor (PDGFR-beta) binds only to PDGF-B. Gene-targeting studies using mice have shown that the genes for PDGF-A and PDGF-B, as well as the two PDGFR genes, are essential for normal development. Furthermore, overexpression of PDGFs is linked to different pathological conditions, including malignancies, atherosclerosis and fibroproliferative diseases. Here we have identify and characterize a fourth member of the PDGF family, PDGF-D. PDGF-D has a two-domain structure similar to PDGF-C and is secreted as a disulphide-linked homodimer, PDGF-DD. Upon limited proteolysis, PDGF-DD is activated and becomes a specific agonistic ligand for PDGFR-beta. PDGF-DD is the first known PDGFR-beta-specific ligand, and its unique receptor specificity indicates that it may be important for development and pathophysiology in several organs.     

Targeting the PDGF signaling pathway in tumor treatment

Platelet-derived growth factor (PDGF) isoforms and PDGF receptors have important functions in the regulation of growth and survival of certain cell types during embryonal development and e.g. tissue repair in the adult. Overactivity of PDGF receptor signaling, by overexpression or mutational events, may drive tumor cell growth. In addition, pericytes of the vasculature and fibroblasts and myofibroblasts of the stroma of solid tumors express PDGF receptors, and PDGF stimulation of such cells promotes tumorigenesis. Inhibition of PDGF receptor signaling has proven to useful for the treatment of patients with certain rare tumors. Whether treatment with PDGF/PDGF receptor antagonists will be beneficial for more common malignancies is the subject for ongoing studies.     

Insulin inhibits platelet-derived growth factor-induced cell proliferation

Cellular behavior can be considered to be the result of a very complex spatial and temporal integration of intracellular and extracellular signals. These signals arise from serum-soluble factors as well as from cell-substrate or cell-cell interactions. The current approach in mitogenesis studies is generally to analyze the effect of a single growth factor on serum-starved cells. In this context, a metabolic hormone such as insulin is found to be a mitogenic agent in many cellular types. In the present study, we have considered the effect of insulin stimulation in platelet-derived growth factor (PDGF)-activated NIH-3T3 and C2C12 cells. Our results show that insulin is able to inhibit strongly both NIH-3T3 and C2C12 cell growth induced by PDGF, one of the most powerful mitotic agents for these cell types. This inhibitory effect of insulin is due primarily to a premature down-regulation of the PDGF receptor. Thus, when NIH-3T3 or C2C12 cells are stimulated with both PDGF and insulin, we observe a decrease in PDGF receptor phosphorylation with respect to cells treated with PDGF alone. In particular, we find that costimulation with insulin leads to a reduced production of H2O2 with respect to cell stimulation with PDGF alone. The relative low concentration of H2O2 in PDGF/insulin-costimulated cell leads to a limited down-regulation of protein tyrosine phosphatases, and, consequently, to a reduced PDGF receptor phosphorylation efficiency. The latter is very likely to be responsible for the insulin-dependent inhibition of PDGF-receptor mitogenic signaling.     

The bovine papillomavirus E5 protein requires a juxtamembrane negative charge for activation of the platelet-derived growth factor beta receptor and transformation of C127 cells

The bovine papillomavirus E5 gene encodes a 44-amino-acid, homodimeric transmembrane protein that is the smallest known transforming protein. The E5 protein transforms cultured fibroblasts by forming a stable complex with the endogenous platelet-derived growth factor (PDGF) beta receptor through transmembrane and juxtamembrane interactions, leading to sustained receptor activation. Aspartic acid 33 in the extracellular juxtamembrane region of the E5 protein is important for cell transformation and interaction with the PDGF beta receptor. A. N. Meyer et al. (Proc. Natl. Acad. Sci USA 91:4634-4638, 1994) speculated that this residue interacted with lysine 499 on the receptor. We constructed E5 mutants containing all possible substitutions at position 33, as well as several double mutants containing substitutions at aspartic acid 33 and at glutamic acid 36, and we examined the ability of these mutants to transform C127 mouse fibroblasts and to bind to and induce activation of the PDGF beta receptor. There was an excellent correlation between the transformation activities of the various mutants and their ability to bind to and activate the PDGF beta receptor. Analysis of the mutants demonstrated that a juxtamembrane negative charge on the E5 protein was required for cell transformation and for productive interaction with the PDGF beta receptor and indicated that aspartic acid 33 was more important for these activities than was glutamic acid 36. These results are consistent with the existence of an essential juxtamembrane salt bridge between lysine 499 on the PDGF beta receptor and an acidic residue in the C terminus of the E5 protein and lend support to our proposed model for the complex between the E5 dimer and the PDGF beta receptor.     

Characterization of two monoclonal antibodies reactive with the external domain of the platelet-derived growth factor receptor

Two monoclonal antibodies against the receptor for platelet-derived growth factor (PDGF) were obtained by immunizing mice with pure PDGF receptor preparations derived from porcine uterus. The antibodies, denoted PDGFR-B1 and PDGFR-B2, both bound to the external domain of the receptor, as demonstrated by indirect immunofluorescence and binding of 125I-labeled antibodies to intact human fibroblasts. Both antibodies precipitated pure 175-kDa 32P-labeled autophosphorylated porcine PDGF receptor as well as a Mr 175,000 glycoprotein from metabolically labeled cells. The monoclonal antibodies did not inhibit binding of 125I-PDGF to human fibroblasts and did not stimulate these cells to undergo mitosis. Both antibodies induced clustering and down-regulation of their antigen. However, this resulted in only a partial loss of cell surface binding sites for PDGF itself, consistent with the conclusion that the monoclonals recognized only one of two or several receptors for PDGF. Clustering and down-regulation were not seen when the cells were incubated with monovalent Fab' fragments of the PDGFR-B2 antibody. The antibodies also stimulated autophosphorylation of pure PDGF receptor, and PDGFR-B2 was shown to stimulate phosphorylation of phosphofructokinase, an exogenous substrate for the PDGF receptor kinase. High concentrations of PDGFR-B2 antibody, or Fab' fragments thereof, failed to enhance the PDGF receptor kinase activity, compatible with the possibility that dimerization was of importance in the antibody-stimulated kinase activity of purified PDGF receptors.     

Activation of the platelet-derived growth factor receptor by the bovine papillomavirus E5 transforming protein.

The bovine papillomavirus E5 gene encodes a 44 amino acid membrane-associated protein that can induce tumorigenic transformation of rodent fibroblast cell lines. Genetic studies suggest that the E5 protein may transform cells by influencing the activity of cellular proteins involved in growth regulation. We report here that the endogenous cellular beta type receptor for the platelet-derived growth factor (PDGF) is constitutively activated in C127 and FR3T3 cells stably transformed by the E5 protein, but not in these cell types transformed by a variety of other oncogenes. In C127 cells, a metabolic precursor as well as the mature form of the receptor is activated by E5 transformation. Activation of the receptor also occurs upon acute E5-mediated transformation of these cells and precedes mitogenic stimulation in this system. Moreover, activation of the receptor by addition of PDGF or the v-sis gene to untransformed cells is sufficient to induce DNA synthesis and stable growth transformation. We propose that the PDGF receptor is an important cellular intermediate in the transforming activity of the bovine papillomavirus E5 protein. There is a short region of sequence similarity between the fibropapillomavirus E5 proteins and PDGF, suggesting that the E5 proteins may activate the PDGF receptor by binding directly to it.     

Ligand-induced interaction between alpha- and beta-type platelet-derived growth factor (PDGF) receptors: role of receptor heterodimers in kinase activation.

Two types of PDGF receptors have been cloned and sequenced. Both receptors are transmembrane glycoproteins with a ligand-stimulatable tyrosine kinase site. We have shown earlier that ligand-induced activation of the beta-type PDGF receptor is due to the conversion of the monomeric form of the receptor to the dimeric form [Bishayee et al. (1989) J. Biol. Chem. 264, 11699-11705]. In the present studies, we have established the ligand-binding specificity of two receptor types and extended it further to investigate the ligand-induced association state of the alpha-receptor and the role of alpha-receptor in the activation of beta-receptor. These studies were conducted with cells that express one or the other type of PDGF receptor as well as with cells that express both types of receptors. Moreover, ligand-binding characteristics of the receptor were confirmed by immunoprecipitation of the receptor-125I-PDGF covalent complex with type-specific anti-PDGF receptor antibodies. These studies revealed that all three isoforms of PDGF bind to alpha-receptor, and such binding leads to dimerization as well as activation of the receptor. In contrast, beta-receptor can be activated only by PDGF BB and not by PDGF AB or PDGF AA. However, by using antipeptide antibodies that are specific for alpha- or beta-type PDGF receptor, we demonstrated that in the presence of alpha-receptor, beta-receptor kinase can be activated by PDGF AB. We present here direct evidence that strongly suggests that such PDGF AB induced activation of beta-receptor is due to the formation of a noncovalently linked alpha-beta receptor heterodimer.     

Ligand stimulation reduces platelet-derived growth factor beta-receptor susceptibility to tyrosine dephosphorylation

Ligand binding to the platelet-derived growth factor (PDGF) beta-receptor leads to increased receptor tyrosine phosphorylation as a consequence of dimerization-induced activation of the intrinsic receptor tyrosine kinase activity. In this study we asked whether ligand-stimulated PDGF beta-receptor tyrosine phosphorylation, to some extent, also involved reduced susceptibility to tyrosine dephosphorylation. To investigate this possibility we compared the sensitivity of ligand-stimulated and non-stimulated forms of tyrosine-phosphorylated PDGF beta-receptors to dephosphorylation using various preparations containing protein-tyrosine phosphatase activity. Ligand-stimulated or unstimulated tyrosine-phosphorylated receptors were obtained after incubation of cells with pervanadate only or pervanadate, together with PDGF-BB, respectively. Dephosphorylation of receptors immobilized on wheat germ agglutinin-Sepharose, as well as of receptors in intact cell membranes, was investigated under conditions when rephosphorylation did not occur. As compared with unstimulated receptors the ligand-stimulated PDGF beta-receptors showed about 10-fold reduced sensitivity to dephosphorylation by cell membranes, a recombinant form of the catalytic domain of density-enhanced phosphatase-1, or recombinant protein-tyrosine phosphatase 1B. We conclude that ligand-stimulated forms of the PDGF beta-receptor display a reduced susceptibility to dephosphorylation. Our findings suggest a novel mechanism whereby ligand stimulation of PDGF beta-receptor, and possibly other tyrosine kinase receptors, leads to a net increase in receptor tyrosine phosphorylation.     

Neurofibrosarcoma-derived Schwann cells overexpress platelet-derived growth factor (PDGF) receptors and are induced to proliferate by PDGF BB

Neurofibromatosis type 1 (NF1) is characterized by the formation of neurofibromas, benign tumors of the peripheral nerve consisting essentially of Schwann cells, which can sometimes turn malignant to form neurofibrosarcomas. The mechanism of progression toward a malignant phenotype remains largely unknown. In this report, we show that platelet-derived growth factor (PDGF) BB, and to a lesser extent fibroblast growth factor 2, are mitogenic for two neurofibrosarcoma-derived Schwann cell lines, but not for a Schwann cell line derived from a schwannoma (from a non-NF1 patient) or for transformed rat Schwann cells. Levels of expression of both PDGF receptor α and β are significantly increased in the two neurofibrosarcoma-derived cell lines compared to the non-NF1 Schwann cell lines. The level of tyrosyl-phosphorylated PDGF receptor β is strongly increased upon stimulation by PDGF BB. In comparison, only modest levels of tyrosyl-phosphorylated PDGF receptor α are observed, upon stimulation by PDGF AA or PDGF BB. Accordingly, PDGF AA is only a weak mitogen for the neurofibrosarcoma-derived cells by comparison to PDGF BB. These results indicate that the mitogenic effect of PDGF BB for the neurofibrosarcoma-derived Schwann cell lines is primarily transduced by PDGF receptor β. Neu differentiation factor β, a potent mitogen for normal Schwann cells, was unable to stimulate proliferation of the transformed Schwann cell lines, due to a dramatic down-regulation of the erbB3 receptor. Therefore, aberrant expression of growth factor receptors by Schwann cells, such as the PDGF receptors, could represent an important step in the process leading to Schwann cell hyperplasia in NF1. J. Cell. Physiol. 177:334–342, 1998. © 1998 Wiley-Liss, Inc. The information in the article does not reflect government policy and no official endorsement should be inferred.     

Dissociation of PDGF receptor tyrosine kinase activity from PDGF-mediated inhibition of gap junctional communication

Gap junction-mediated intercellular communication (GJC) may play an important role in cell proliferation and transformation since GJC is inhibited by growth factors, oncogenes, tumor promoters, and carcinogens. We have studied inhibition of GJC by platelet-derived growth factor-BB (PDGF) in the mouse fibroblast cell line C3H/10T1/2 and have sought to determine whether PDGF-induced inhibition of GJC is mediated by the PDGF receptor tyrosine kinase (RTK). PDGF-mediated inhibition of GJC was rapid and transient, with maximal inhibition occurring 40 min after PDGF addition and GJC returning to control levels after 70 min. The effect of PDGF on GJC was concentration-dependent, with maximal inhibition of 90% or greater occurring at 10 ng/ml PDGF. Stimulation of RTK activity, as determined by antiphosphotyrosine immunoblot analysis of PDGF receptor and the receptor substrates phospholipase C-γl (PLC-γl) and guanosine triphosphatase activating protein (GAP), was also concentration-dependent. Inhibition of GJC required a greater concentration of PDGF than did stimulation of RTK activity. The tyrosine kinase inhibitor genistein blocked PDGF-induced RTK activity, as measured by PDGF receptor, PLC-γl, and GAP tyrosine phosphorylation, in a concentration-dependent manner but did not affect PDGF-mediated inhibition of GJC. Genistein alone had no effect on GJC or PDGF receptor expression. PDGF treatment in the presence or absence of genistein resulted in phosphorylation of the connexin 43 protein on nontyrosine residues. These results suggest that inhibition of GJC by ligand-activated PDGF receptor is dissociable from the RTK activity responsible for PDGF, PLC-γl, and GAP phosphorylation. © 1994 Wiley-Liss, Inc.     

Activated platelet-derived growth factor autocrine pathway drives the transformed phenotype of a human glioblastoma cell line

Human glioblastoma cells (A172) were found to concomitantly express PDGF-BB and PDGF β-receptors. The receptors were constitutively autophosphorylated in the absence of exogenous ligand, suggesting the presence of an autocrine PDGF pathway. Neutralizing PDGF antibodies as well as suramin inhibited the autonomous PDGF receptor tyrosine kinase activity and resulted in up-regulation of receptor protein. The interruption of the autocrine loop by the PDGF antibodies reversed the transformed phenotype of the glioblastoma cell, as determined by (1) diminished DNA synthesis, (2) inhibition of tumor colony growth, and (3) reversion of the transformed morphology of the tumor cells. The PDGF antibodies showed no effect on the DNA synthesis of another glioblastoma cells line (U343MGa 31L) or on Ki-ras-transformed fibroblasts. The present study demonstrates an endogenously activated PDGF pathway in a spontaneous human glioblastoma cell line. Furthermore, we provide evidence that the autocrine PDGF pathway drives the transtormed phenotype of the tumor cells, a process that can be blocked by extracellular antagonists. © 1994 Wiley-Liss, Inc.     

Dissecting tumor maintenance requirements using bioluminescence imaging of cell proliferation in a mouse glioma model

Bioluminescence imaging has previously been used to monitor the formation of grafted tumors in vivo and measure cell number during tumor progression and response to therapy. The development and optimization of successful cancer therapy strategies may well require detailed and specific assessment of biological processes in response to mechanistic intervention. Here, we use bioluminescence imaging to monitor the cell cycle in a genetically engineered, histologically accurate model of glioma in vivo. In these platelet-derived growth factor (PDGF)-driven oligodendrogliomas, G1 cell-cycle arrest is generated by blockade of either the PDGF receptor or mTOR using small-molecule inhibitors.     

A phosphatidylinositol-3 kinase binds to platelet-derived growth factor receptors through a specific receptor sequence containing phosphotyrosine.

Platelet-derived growth factor (PDGF) stimulates autophosphorylation of the PDGF receptor and association of the receptor with several cytoplasmic molecules, including phosphatidylinositol-3 kinase (PI3 kinase). In this study we examined the association of PI3 kinase with immunoprecipitated autophosphorylated PDGF receptor in vitro. The PI3 kinase from cell lysates bound to the wild-type receptor but not to a mutant receptor that had a deletion of the kinase insert region. A protein of an apparent size of 85 kDa bound to the receptor, consistent with previous observations that a protein of this size is associated with PI3 kinase activity. In addition, 110- and 74-kDa proteins bound to the phosphorylated receptor. Dephosphorylated receptors lost the ability to bind PI3 kinase activity as well as the 85-kDa protein. A 20-amino-acid peptide composed of a sequence in the kinase insert region that included one of the autophosphorylation sites of the receptor (tyrosine 719) as well as a nearby tyrosine (Y708) blocked the binding of PI3 kinase to the receptor, but only when the peptide was phosphorylated on tyrosine residues. A scrambled version of the peptide did not block PI3 kinase binding to the receptor even when it was phosphorylated on tyrosine. These tyrosine-phosphorylated peptides did not block binding of phospholipase C-gamma or GTPase-activating protein to the receptor. In separate experiments (receptor blots), soluble radiolabeled receptor bound specifically to an 85-kDa protein present in sodium dodecyl sulfate-polyacrylamide gel electrophoresis-fractionated 3T3 cell lysates that were transferred to nitrocellulose paper. The binding was blocked by the same tyrosine-phosphorylated peptides that prevented binding of PI3 kinase activity to immobilized receptors. These findings show that the PDGF receptor binds directly to an 85-kDa protein and to a PI3 kinase activity through specific sequences in the kinase insert region. The association of a 110-kDa protein with the receptor also involve these sequences, suggesting that this protein may be a subunit of the PI3 kinase. Phosphotyrosine is an essential structure required for the interactions of these proteins with the PDGF receptor.     

Actions of platelet-derived growth factor isoforms in mesangial cells

Platelet-derived growth factor (PDGF) occurs as homodimers or heterodimers of related polypeptide chains PDGF-BB, -AA, and -AB. There are two receptors that bind PDGF, termed alpha and beta. The beta receptor recognizes PDGF B chain and is dimerized in response to PDGF BB. The alpha receptor recognizes PDGF B as well as A chains and can be dimerized by the three dimeric forms of PDGF AA, AB, and BB. To characterize PDGF receptor signaling mechanisms and biologic activities in human mesangial cells (MC), we explored the effects of the three PDGF isoforms on DNA synthesis, phospholipase C activation, and PDGF protooncogene induction. PDGF-BB homodimer and AB heterodimer induced a marked increase in DNA synthesis, activation of phsopholipase C, and autoinduction of PDGF A and B chain mRNAs, whereas PDGF-AA homodimer was without effect. The lack of response to PDGF AA could be accounted for by down regulation of the PDGF-alpha receptor since preincubation of MC with suramin restored PDGF AA-induced DNA synthesis. Ligand binding studies demonstrate specific binding of labeled PDGF BB and AB and to a lower extent PDGF AA isoforms to mesangial cells. These results are consistent with predominant expression of PDGF beta receptor in MC, which is linked to phospholipase-C activation. The potent biologic effects of PDGF-AB heterodimer in cells that express very few alpha receptors and do not respond to PDGF AA are somewhat inconsistent with the currently accepted model of PDGF receptor interaction and suggest the presence of additional mechanisms for PDGF isoform binding and activation. © 1994 Wiley-Liss, Inc.     

Specific interaction between the bovine papillomavirus E5 transforming protein and the beta receptor for platelet-derived growth factor in stably transformed and acutely transfected cells.

The E5 protein of bovine papillomavirus is a 44-amino-acid membrane protein which induces morphologic and tumorigenic transformation of fibroblasts. We previously showed that the E5 protein activates and forms a complex with the endogenous beta receptor for platelet-derived growth factor (PDGF) in transformed rodent fibroblasts and that the PDGF beta receptor can mediate tumorigenic transformation by the E5 protein in a heterologous cell system. Other workers have identified the receptor for epidermal growth factor (EGF) as a potential target of the E5 protein in NIH 3T3 cells. Here, we investigate the specificity of the interaction of the E5 protein with various growth factor receptors, with particular emphasis on the PDGF beta receptor and the EGF receptor. Under conditions where both the PDGF beta receptor and the EGF receptor are stably expressed in E5-transformed mouse and bovine fibroblasts and in E5-transformed epithelial cells, the E5 protein specifically forms a complex with and activates the PDGF receptor and not the EGF receptor. Under conditions of transient overexpression in COS cells, the E5 protein has the potential to associate with several growth factor receptors, including the EGF receptor. However, upon coexpression of PDGF beta receptors and EGF receptors in COS cells, the E5 protein preferentially forms a complex with the PDGF receptor. Therefore, we conclude that the PDGF beta receptor is the primary target for the E5 protein in a variety of cell types, including bovine fibroblasts.