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P Lengyel

Publications and source records attributed to P Lengyel.

At least 19 recordsLinked to original sources
PubMed2002

The increase in levels of interferon-inducible proteins p202a and p202b and RNA-dependent protein kinase (PKR) during myoblast differentiation is due to transactivation by MyoD: their tissue distribution in uninfected mice does not depend on interferons.

The murine 200 family proteins p202a, p202b, and p204, and also RNA-dependent protein kinase (PKR) are inducible by interferons (IFNs). p202a, p202b, and p204 modulate the activity of a large variety of transcription factors and also are involved in muscle differentiation. PKR is a multifunctional serine/threonine kinase, which is involved in antiviral defense and cell growth control and in the response to various stress signals. We reported earlier that the level of p204 increases during cultured C2C12 myoblast differentiation to myotubes in consequence of transactivation by the skeletal muscle-specific MyoD protein. The levels of p202a, p202b, and PKR also increase during the differentiation. We report here that these increased protein levels also are due to the transactivation of their genes by MyoD. This is made possible by the occurrence in each of these genes of at least six E boxes, which are recognition sites for MyoD. We also show that the distribution of the p204, p202a, p202b, and PKR proteins in five tissues of adult C129 mice is the same in wild-type mice and mice lacking the IFN-alpha, IFN-beta, and IFN-gamma receptors. This indicates that the synthesis and distribution of these proteins in uninfected adult mice are not affected by endogenous IFNs.

Animals↗
PubMed2001

p202, an interferon-inducible negative regulator of cell growth, is a target of the adenovirus E1A protein.

Studies have revealed that human adenovirus-encoded E1A protein promotes cell proliferation through the targeted interaction with cellular proteins that act as key negative regulators of cell growth. The targets of E1A protein include the retinoblastoma tumor suppressor protein (pRb). Because p202, an interferon (IFN)-inducible murine protein (52-kDa), negatively regulates cell growth in part through the pRb/E2F pathway, we tested whether the p202 is a target of the adenovirus-encoded E1A protein for functional inactivation. Here we report that the expression of E1A protein overcame p202-mediated inhibition of cell growth and this correlated with an alleviation of p202-mediated inhibition of the transcriptional activity of E2F. Furthermore, E1A protein relieved p202-mediated inhibition of the specific DNA-binding activity of E2F complexes, including those containing the pocket proteins. Additionally, the E1A protein bound to p202 both in vitro and in vivo and a deletion of four amino acids in the conserved region 2 (CR2) of E1A protein significantly reduced the binding of E1A to p202. Interestingly, ectopic expression of p202 under reduced serum conditions significantly reduced E1A-mediated apoptosis. Taken together, our observations provide support to the idea that the p202 and adenovirus E1A protein functionally counteract each other and E1A protein targets p202 to promote cell proliferation.

Adenovirus E1A Proteins↗
PubMed2000

The interferon- and differentiation-inducible p202a protein inhibits the transcriptional activity of c-Myc by blocking its association with Max.

p202a is a murine protein that is induced during the fusion of myoblasts to myotubes and can also be induced by interferon. Even 2-3-fold overexpression of p202a in cells retards proliferation. p202a was shown to modulate transcription by binding, and inhibiting the activity of several transcription factors including c-Fos, c-Jun, AP-2, E2F1, E2F4, NF-kappaB, MyoD, and myogenin. Here we report that p202a also bound the c-Myc protein in vitro and in vivo; the C-terminal p202a b segment bound the C-terminal basic region helix-loop-helix-leucine zipper (bHLHLZ) region of c-Myc. The transfection of a p202a expression plasmid inhibited the c-Myc-dependent expression of reporter plasmids in transient assays; moreover, overexpression of p202a in stable cell lines decreased the endogenous levels of mRNAs whose expression is driven by c-Myc. These effects of p202a are consistent with our finding that the binding of p202a to c-Myc inhibited the binding of c-Myc to Max in vitro and in vivo. p202a also inhibited the c-Myc-induced anchorage-independent growth and apoptosis of Rat-1 cells. The inhibition of c-Myc-dependent transcription, proliferation, and apoptosis by p202a is in line with the involvement of p202a in differentiation.

Animals↗
PubMed2000

MyoD-dependent induction during myoblast differentiation of p204, a protein also inducible by interferon.

p204, an interferon-inducible p200 family protein, inhibits rRNA synthesis in fibroblasts by blocking the binding of the upstream binding factor transcription factor to DNA. Here we report that among 10 adult mouse tissues tested, the level of p204 was highest in heart and skeletal muscles. In cultured C2C12 skeletal muscle myoblasts, p204 was nucleoplasmic and its level was low. During myoblast fusion this level strongly increased, p204 became phosphorylated, and the bulk of p204 appeared in the cytoplasm of the myotubes. Leptomycin B, an inhibitor of nuclear export that blocked myoblast fusion, inhibited the nuclear export signal-dependent translocation of p204 to the cytoplasm. The increase in the p204 level during myoblast fusion was a consequence of MyoD transcription factor binding to several MyoD-specific sequences in the gene encoding p204, followed by transcription. Overexpression of p204 (in C2C12 myoblasts carrying an inducible p204 expression plasmid) accelerated the fusion of myoblasts to myotubes in differentiation medium and induced the fusion even in growth medium. The level of p204 in mouse heart muscle strongly increased during differentiation; it was barely detectable in 10. 5-day-old embryos, reached the peak level in 16.5-day-old embryos, and remained high thereafter. p204 is the second p200 family protein (after p202a) found to be involved in muscle differentiation. (p202a was formerly designated p202. The new designation is due to the identification of a highly similar protein-p202b [H. Wang, G. Chatterjee, J. J. Meyer, C. J. Liu, N. A. Manjunath, P. Bray-Ward, and P. Lengyel, Genomics 60:281-294, 1999].) These results reveal that p204 and p202a function in both muscle differentiation and interferon action.

Actinin↗
PubMed1999

Characteristics of three homologous 202 genes (Ifi202a, Ifi202b, and Ifi202c) from the murine interferon-activatable gene 200 cluster.

The Ifi202 gene is part of the interferon-activatable murine gene 200 cluster on chromosome 1. Ifi202 encodes the p202 protein whose overexpression is growth inhibitory and which can bind and inhibit the activity of numerous transcription factors including c-Jun, c-Fos, NF-kappaB, E2F-1, E2F-4, MyoD, and myogenin. We report here the exon-intron structure of Ifi202 and the discovery of Ifi202b and Ifi202c, close homologs of Ifi202 (whose designation we now change to Ifi202a). Ifi202a, b, and c were colocalized to chromosome 1 bands H4-H5 by fluorescence in situ hybridization. Ifi202b encodes p202b, which is interferon-inducible and differs from p202a in only 7 of 445 amino acids. 202b mRNA is constitutively expressed in tissues in which 202a mRNA is expressed. Ifi202c is apparently an unexpressed pseudogene. In murine embryonic fibroblasts (MEFs) from 129 mice, the level of 202b mRNA is approximately half that of 202a mRNA. We knocked out the Ifi202a gene from 129 mice. The expression of 202b mRNA, but not 202a mRNA, persisted in the knockout mice and their MEFs at the same level as in wildtype mice. However, in MEFs from the knockout mice, the constitutive and interferon-induced levels of p202b were approximately as high as the constitutive and the interferon-induced levels of p202a plus p202b, respectively, in MEFs from wildtype mice. These findings suggest dosage compensation at the posttranscriptional level. This might account for the apparent lack of phenotype of the knockout mice.

Animals↗
PubMed1999

The interferon-inducible nucleolar p204 protein binds the ribosomal RNA-specific UBF1 transcription factor and inhibits ribosomal RNA transcription.

p204, a member of the interferon-inducible p200 family of murine proteins, is primarily nucleolar. We generated cell lines in which p204 was inducible by muristerone. This induction resulted in retardation of cell proliferation and inhibition of rRNA transcription in vivo. Interferon treatment, resulting in p204 induction and retardation of proliferation, also caused inhibition of rRNA transcription in vivo. p204 also inhibited rRNA transcription in vitro. This inhibition was overcome by addition of UBF1, the rRNA-specific transcription factor. A direct interaction between p204 and UBF1 was revealed in vitro in pull-down assays, and in vivo by co-immunoprecipitation from cell extracts. UBF1 bound strongly to at least two regions of p204: the N-terminal segment linked to the conserved 200 amino acid a segment, and the conserved 200 amino acid b segment. Cleavage of the a or b segments into two segments (encoded by single exons) resulted in a strong decrease or loss of binding. The inhibition of rRNA transcription by p204 may be due to the inhibition by p204 of the specific DNA binding of UBF1. This was revealed in electrophoretic mobility shift, magnetic bead and footprinting assays. Thus, p204 serves as a mediator of the inhibition of rRNA transcription by interferon.

Animals↗
PubMed1998

Increase in p202 expression during skeletal muscle differentiation: inhibition of MyoD protein expression and activity by p202.

p202 is a primarily nuclear, interferon-inducible murine protein that is encoded by the Ifi 202 gene. Overexpression of p202 in transfected cells retards cell proliferation. p202 modulates the pattern of gene expression by inhibiting the activity of various transcription factors including NF-kappaB, c-Fos, c-Jun, E2F-1, and p53. Here we report that p202 was constitutively expressed in mouse skeletal muscle and that the levels of 202 RNA and p202 greatly increased during the differentiation of cultured C2C12 myoblasts to myotubes. When overexpressed in transfected myoblasts, p202 inhibited the expression of one muscle protein (MyoD) without affecting the expression of a second one (myogenin). Thus, the decrease in the level of MyoD (but not of myogenin) during muscle differentiation may be the consequence of the increase in p202 level. Overexpressed p202 also inhibited the transcriptional activity of both MyoD and myogenin. This inhibition was correlated with an interaction of p202 with both proteins, as well as the inhibition by p202 of the sequence-specific binding of both proteins to DNA. This inhibition of the expression of MyoD and of the transcriptional activity of MyoD and myogenin may account for the inhibition of the induction of myoblast differentiation by premature overexpression of p202.

Animals↗
PubMed1996

p202, an interferon-inducible modulator of transcription, inhibits transcriptional activation by the p53 tumor suppressor protein, and a segment from the p53-binding protein 1 that binds to p202 overcomes this inhibition.

p202, an interferon-inducible murine protein, is a member of the "200 family" of proteins and is primarily nuclear. p202 is a modulator of transcription; it binds several transcription factors, including NF-kappaB p50 and p65, AP-1 c-Fos and c-Jun, and E2F1, and inhibits their transcriptional activity. p202 also binds pRb, the retinoblastoma protein, and if overexpressed it retards cell proliferation. Here we report that using the yeast two-hybrid assay we found that p202 bound the murine homolog of the human p53-binding protein 1 (53BP1), a protein shown to interact with the DNA binding domain of the p53 tumor suppressor protein. p202 bound a 98amino acid segment from 53BP1. This binding was inhibited by the replacement in p202 of a histidine (from the M(F/L)HATVA(T/S) sequence that is conserved among all of the 200 family proteins) by phenylalanine. We also report that overexpression of p202 inhibited the p53-dependent expression of reporter genes containing p53-activable segments from the mdm2 and p21 genes, whereas a decrease in the p202 level (in consequence of the expression of 202 antisense RNA) resulted in an increase in the p53-dependent expression of these reporters. Expression of the 53BP1 segment binding to p202 overcame the inhibition by overexpressed p202 of the transcription of reporters mediated by the p53 or the AP-1 transcription factors and of the proliferation of yeast.

Amino Acid Sequence↗
PubMed1996

Inhibition of E2F-mediated transcription by p202.

Many of the antimicrobial, immunomodulatory and cell growth inhibitory activities of the interferons are mediated by interferon-inducible proteins. Earlier we characterized an interferon-inducible murine protein, p202, whose expression in transfected cells inhibits cell proliferation and which can form a complex with retinoblastoma protein (pRb). Here we report that in transfected cells expression of p202 inhibits E2F-stimulated transcription of a reporter gene and of endogenous genes. Inhibition of the transcriptional activity of E2F by p202 does not depend on fully functional pRb and is correlated with inhibition of the sequence-specific DNA binding of E2F. p202 interacts with the transcription factor E2F (E2F-1/DP-1) in vitro and in vivo. Inhibition of E2F activity by p202 may contribute to growth inhibition by the interferons.

Animals↗
PubMed1996

The interferon-inducible p202 protein as a modulator of transcription: inhibition of NF-kappa B, c-Fos, and c-Jun activities.

The antimicrobial, immunomodulatory, and cell growth-regulatory activities of the interferons are mediated by interferon-inducible proteins. One of these is p202, a nuclear protein that is encoded by the Ifi 202 gene from the interferon-activatable gene 200 cluster. Overexpression of p202 in transfected cells slows down cell proliferation. As shown earlier, p202 binds to the hypophosphorylated form of the retinoblastoma susceptibility protein. Here we report that p202 inhibits the activities of the NF-kappa B and the AP-1 enhancers both in transiently transfected cells and in transfected stable cell lines overexpressing p202. Furthermore, p202 binds the NF-kappa B p50 and p65 and the AP-1 c-Fos and c-Jun transcription factors in vitro and in vivo. NF-kappa B, c-Fos, and c-Jun participate in the transcription of various cellular and viral genes, and thus p202 can modulate the expression of these genes in response to interferons.

Animals↗
PubMed1995

Binding of an interferon-inducible protein (p202) to the retinoblastoma protein.

Many of the antimicrobial, immunomodulatory, and cell growth regulatory activities of the interferons are mediated by interferon-inducible proteins. One family of such murine proteins is encoded by six or more adjacent and structurally related genes (gene 200 cluster). Two homologous human genes have also been reported. p202, encoded by the Ifi202 gene in the gene 200 cluster, is a 52-kDa nuclear phosphoprotein. Constitutive overexpression of p202 in transfected cells is growth-inhibitory. We report here that p202 binds the cell growth regulatory retinoblastoma protein (pRb) in vitro and in vivo. The binding is due to direct interaction between the two proteins. p202 has two nonoverlapping segments for binding pRb, and pRb has two nonoverlapping segments (one of them including the pocket region) for binding p202. The hypophosphorylated form of pRb binds to p202, p202 is the first interferon-inducible protein found to bind pRb.

Amino Acid Sequence↗
PubMed1993

Tumor-suppressor genes: news about the interferon connection.

The interferons are a family of secreted, multifunctional proteins which are components of the defenses of vertebrates against viral, bacterial, and parasitic infections and certain tumors. They exert their various activities by inducing the synthesis of a large variety of proteins. There are direct and indirect indications that several of these proteins may have tumor-suppressor activities. The interferon-inducible proteins implicated include: (i) a double-stranded RNA-activatable protein kinase that can phosphorylate and thereby inactivate the eukaryotic peptide chain initiation factor eIF-2; (ii) the interferon regulatory factors IRF-1 and IRF-2, which can modulate the expression of the interferons and of some interferon-inducible proteins; and (iii) RNase L, a latent endoribonuclease which can be activated by (2'-5')oligoadenylates, the products of a family of enzymes which are also interferon-inducible. It is note-worthy that some of the proteins encoded by tumor virus oncogenes (e.g., E1A from adenovirus, EBNA-2 from Epstein-Barr virus, and terminal protein from hepatitis B virus) impair the induction of at least some proteins by interferons.

Animals↗
PubMed1993

Interferon action: cytoplasmic and nuclear localization of the interferon-inducible 52-kD protein that is encoded by the Ifi 200 gene from the gene 200 cluster.

Recently, we reported that an interferon (IFN)-inducible, murine 72-kD phosphoprotein (the 204 protein) that is encoded by the Ifi 204 gene from the gene 200 cluster is localized in the nucleolus and the nucleoplasm. We have now raised a polyclonal antiserum against the 202 protein that is encoded by the Ifi 202 gene from the same gene cluster and regions of which are homologous to those from the 204 protein. Using the antiserum, we established that the 202 protein is a 52-kD phosphoprotein whose level in cells from various murine lines can be increased up to 16-fold upon treatment with IFN-alpha. Experiments involving fractionation of cell lysates and indirect immunofluorescence microscopy of cultured cells revealed that the 202 protein was localized in the cytoplasm and the nucleus. Upon treatment of cells with IFN, the 202 protein first accumulated on the surface of a cytoplasmic, membranous fraction and after prolonged treatment with IFN it was localized mainly in the nucleus. In IFN-treated mitotic AKR cells, the 202 protein was colocalized with chromosomes. 202 protein extracted from IFN-treated AKR cells bound double-stranded DNA in vitro. Studies on 202 protein function should be facilitated by the availability of complete cDNA clones and the finding of cell lines and an inbred strain of mice in which the expression of this protein was impaired.

Animals↗
PubMed1992

Impaired transcription of the poly rI:rC- and interferon-activatable 202 gene in mice and cell lines from the C57BL/6 strain.

Activation of 202 and (2'-5')(A)n synthetase genes after injection of interferon (IFN)-inducing, double-stranded, poly rI:rC was compared in various mouse strains. The 202 mRNA level increased 4.5- to 10-fold in DBA/2, BALB/c, and C3H/HeJ mice, whereas in C57BL/6 mice it rose only to about that in untreated DBA/2, BALB/c, and C3H/HeJ mice. To determine whether this low level was due to a reduced transcription rate, a nuclear "run-on" assay was performed with NIH 3T3 cells or BLK cells derived from C57BL/6 mice. IFN-alpha increased the 202 mRNA transcription severalfold in NIH 3T3 cells only, and that of a (2'-5')(A)n synthetase gene in both cell lines. The possibility that an alteration in transacting factors could be responsible for this difference was examined. For this purpose the 5' terminal flanking region (called the b segment, about 0.8 kb) of the 202 gene was linked to a heterologous reporter gene--chloramphenicolacetyl-transferase (CAT) and transfected into normal or transformed NIH 3T3 cells and into various C57BL/6-derived cell lines. IFN-alpha induced strong CAT activity in transfected normal or transformed NIH 3T3 cells, but a much lower activity in those from C57BL/6 mice. The b segment contains an IFN-responsive element (ISRE) (35 bp) homologous to that present in several other IFN-inducible genes. Three tandem copies of the 202 ISRE were linked to an enhancerless SV40 early promoter driving an influenza virus hemagglutinin (HA) cDNA segment. No increase in HA mRNA expression was detected in the transfected BLK cell line derived from C57BL/6 mice following IFN treatment, whereas in the NIH 3T3 cell line, the IFN treatment resulted in a 2.5-fold increase. These and other results suggest that C57BL/6 mice and cell lines derived from them might carry defective transacting factors impairing the ability of IFN-alpha to activate the 202 gene without impairing its ability to activate a (2'-5')(A)n synthetase gene.

2',5'-Oligoadenylate Synthetase↗
PubMed1992

Interferon action: nucleolar and nucleoplasmic localization of the interferon-inducible 72-kD protein that is encoded by the Ifi 204 gene from the gene 200 cluster.

The interferons are cytokines with antiviral, cell growth regulatory, and immunomodulatory activities. These activities are mediated by the proteins induced by the interferons. Earlier we described a gene cluster (the 200 cluster) consisting of at least six adjacent, interferon-activatable genes located next to the erythroid alpha-spectrin locus on murine chromosome 1. The genes of the cluster arose by repeated gene duplication and they specify proteins with pronounced sequence similarity. We have now raised polyclonal antibodies against a segment from one of these proteins (the 204 protein of 72 kD). Using these, we established that the 204 protein is a phosphoprotein whose level in cells from various murine lines can be increased up to 75-fold upon treatment with alpha interferon. Experiments involving fractionation of cell lysates and indirect immunofluorescence microscopy of control and interferon-treated cells revealed that the 204 protein is nucleolar and nucleoplasmic. This conclusion was confirmed by co-localization with B23, a known nucleolar protein. The 204 protein is the first interferon-induced protein found to be located in the nucleoli, the subcellular organelles of ribosomal RNA production and ribosome assembly. It remains to be seen whether the 204 protein affects any of these processes. Studies on 204 protein function should be facilitated by the availability of complete cDNA clones and the finding of cell lines in which the expression of this protein is impaired.

Amino Acid Sequence↗
PubMed1991

Characterization of recessive (mediator-) revertants from NIH 3T3 cells transformed with a c-H-ras oncogene.

We have reported earlier the isolation of two recessive, serum- and anchorage-dependent revertants from an NIH 3T3 line which had been transformed with multiple copies of a c-H-ras oncogene. In both revertants the oncogene was fully expressed and fusion of either revertant with normal (untransformed) cells or of the two revertants with one another resulted in transformed progeny. These, and other data indicate that the transforming activity of the c-H-ras oncogene is impaired in the two revertants, in consequence of defects in distinct genes needed to mediate this activity. Here, we describe some of the biochemical features of the revertants. In both of these (as in the transformed line) the bulk of the ras-p21 protein was found in the membrane fraction. This suggests proper posttranslational processing. Furthermore, no difference was detected either in the ras-p21 protein GTPase stimulating activity of GAP or in the extent of GAP-tyrosine phosphorylation among growing cultures of the two revertants, the transformed line and the parental NIH 3T3 line. The level of glucose transporter mRNA was severalfold higher in the transformed line than in the NIH 3T3 line. In the two revertants, however, the level was as low as that in the NIH 3T3 line. This indicates that the reversion impaired the effect of the c-H-ras oncogene on transcription. The raf oncogene (proposed to increase transcription factor activity) could retransform both revertants. Moreover, as revealed in experiments with growing cultures, neither transformation by the c-H-ras oncogene nor reversion from the transformed state altered the electrophoretic mobility of the raf protein or the level of its actin kinase activity. These results suggest that transformation by the c-H-ras oncogene is not mediated by the activation of raf protein kinase. The tyrosine phosphorylation of the p34cdc2 protein kinase (a cell cycle regulatory enzyme) was severalfold higher in the transformed line than in the NIH 3T3 line. The level of p34cdc2 protein kinase phosphorylation was as high in the R260 revertant as in the transformed line and as low in the R116 revertant as in the NIH 3T3 line. We are attempting to identify the defective mediator genes impairing the transforming activity of the c-H-ras oncogene in the two revertants.

Cell Line, Transformed↗