In memoriam. Karl Günther Zimmer (1911-1988).
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Berlin↗
Biomedical subjects
P Herrlich
Publications and source records attributed to P Herrlich.
Phorbol ester-inducible genes contain a common cis element recognized by a TPA-modulated trans-acting factor.
The promoter regions of several phorbol diester-(TPA-) inducible genes (collagenase, stromelysin, hMT IIA, and SV40) share a conserved 9 bp motif. Synthetic copies of these closely related sequences conferred TPA inducibility upon heterologous promoters. Footprinting analysis indicated that these TPA-responsive elements (TREs) are recognized by a common cellular protein: the previously described transcription factor AP-1. A point mutation that eliminated the basal and induced activity of the TRE also interfered with its ability to bind AP-1. Treatment of cultured cells with TPA led to a rapid 3- to 4-fold increase in TRE binding activity, by a posttranslational mechanism. These results strongly suggest that AP-1 is at the receiving end of a complex pathway responsible for transmitting the effects of phorbol ester tumor promoters from the plasma membrane to the transcriptional machinery.
Animals↗
Posttranscriptional regulation of c-fos mRNA expression.
The transient induction of c-fos mRNA and protein suggests that regulation occurs not only by transcriptional activation but also at the level of turnover of the gene product. Here we present evidence for the rapid turnover of c-fos mRNA and some of the requirements for its specific degradation. The half life of induced mature cytoplasmic c-fos mRNA is 9 min in both serum-starved and growing primary human fibroblasts and in NIH 3T3 cells. A structure present at the 3' end of the c-fos mRNA molecule is involved in its low stability since the substitution or the removal of the untranslated 3' portion prolongues the RNA life time. The rapid turnover of fos mRNA requires, in addition, continued protein synthesis. Treatment of cells with cycloheximide stabilizes c-fos mRNA. Washing out cycloheximide reestablishes the rapid turnover. Both changes occur with lag periods of less than 17 minutes.
Adult↗
Alpha-radiation-induced amplification of integrated SV40 sequences is mediated by a trans-acting mechanism.
Treatment of Chinese hamster embryo cells with alpha irradiation (4 MeV, emitted by 241americium) induces a 15-fold amplification of integrated SV40 sequences. The extent of amplification depends on the dose of irradiation and on the presence of a functional T-antigen encoded by the SV40 A gene. The inducing signal can be transmitted to a non-irradiated nucleus following cell fusion. Amplification is therefore the result of some trans-acting process, and this could explain how SV40 amplification can occur after doses of alpha irradiation that are too low to cause direct DNA damage within the SV40 replicon.
Alpha Particles↗
12-O-tetradecanoyl-phorbol-13-acetate induction of the human collagenase gene is mediated by an inducible enhancer element located in the 5'-flanking region.
Genomic clones coding for human fibroblast collagenase were isolated. By constructing and transfecting mutants with 5' and 3' deletion mutations of the 5' control region of the gene into human or murine cells, we delimited a 32-base-pair sequence at positions -73 to -42 which is required for the induction of transcription by the tumor promoter 12-O-tetradecanoyl-phorbol-13-acetate. The DNA element behaves as a 12-O-tetradecanoyl-phorbol-13-acetate-inducible enhancer: it mediates the stimulation of transcription to the heterologous herpes simplex virus thymidine kinase promoter and acts in a position- and orientation-independent manner. Differences in enhancer efficiency in different cell lines are interpreted to indicate differences in the activity of a trans-acting factor.
Adult↗
Comparison of human stromelysin and collagenase by cloning and sequence analysis.
A comparison of the cDNA-derived amino acid sequences of human stromelysin and collagenase with the N-terminal sequences of purified enzymes reveals that these metalloproteinases are highly conserved and that they are secreted as proenzymes. A putative zinc-binding site was identified by its homology with the zinc-chelating sequence of thermolysin. These sequences permitted the identification of: transin, a protein induced in rat fibroblasts either exposed to growth factors or transformed by oncogenic viruses, as the rat homologue of stromelysin, and XHF1, a protein induced in human fibroblasts after treatment with tumourigenic agents, as collagenase.
Amino Acid Sequence↗
Interferon-gamma, mitomycin C, and cycloheximide as regulatory agents of MHC class II-associated invariant chain expression.
The murine and human major histocompatibility complex class II-associated invariant chain genes are expressed in mature B cells and in antigen-presenting cells. Several pre-B cell lines and fibroblasts do not naturally contain invariant chain mRNA. Expression is inducible, however, by interferons and other agents interfering with proliferation. Mitomycin C induces the transcription of the gene in pre-B cells, but not in fibroblasts. Interferon-gamma acts in both types of cells. Cycloheximide inhibits the induction of the invariant chain mRNA by interferon-gamma, suggesting that protein synthesis is required. In fact, cycloheximide itself increases the transcriptional rate at the invariant chain gene, suggesting the existence of a labile repressor or an indirect action through cycloheximide arrest of the cell cycle. Lipopolysaccharide (LPS) activation of B lymphocytes causes a rapid decrease of the invariant chain mRNA level and of the amount of invariant chain protein due to rapid turnover. Also class II alpha and beta mRNA expression decreases after LPS treatment. The decrease of invariant chain protein is accompanied by increased surface expression of alpha and beta. The murine invariant chain gene transfected into human fibroblasts is regulated by the same agents and the same dose of agents as is the endogenous gene. The differentiation marker invariant chain thus seems to be transcribed from a gene that is accessible to regulation even in nonlymphoid cells and the expression of which is linked to states of nonproliferation. The sequence responsible for these responses is contained within the cloned genomic fragment and is conserved between mouse and man.
Actins↗
The mammalian genetic stress response.
A number of carcinogenic and cocarcinogenic agents induce new gene products in mammalian cells including primary human skin fibroblasts. These have been defined by cDNA cloning techniques, by protein resolutions in 2D PAGE and by the detection of new enzymatic functions. The uniform and transient genetic reaction is tentatively called the genetic stress response.
Animals↗
Induction of metallothionein and other mRNA species by carcinogens and tumor promoters in primary human skin fibroblasts.
We used nucleic acid hybridization and cDNA cloning techniques to isolate human sequences that respond to the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA). These clones were used as probes to examine changes of gene expression that occurred after the proliferation of exponentially growing primary human fibroblasts was arrested. Transcript levels detected by these probes were increased coordinately by treatment of the cells with UV light, mitomycin C, TPA, or the UV light-induced extracellular protein synthesis-inducing factor EPIF (M. Schorpp, U. Mallick, H. J. Rahmsdorf, and P. Herrlich, Cell 37:861-868, 1984). Proteins coded for by these transcripts were characterized by hybrid-promoted translation and by cDNA sequencing. One of the cDNA clones was homologous to the metallothionein IIa gene, and one set of related clones selected RNA for the secreted TPA-inducible protein XHF1 (U. Mallick, H. J. Rahmsdorf, N. Yamamoto, H. Ponta, R.-D. Wegner, and P. Herrlich, Proc. Natl. Acad. Sci. USA 79:7886-7890, 1982).
Cell Transformation, Neoplastic↗
Selective gene amplification in mammalian cells after exposure to 60Co gamma rays, 241Am alpha particles, or uv light.
Simian Virus 40 wild type (SV40)-transformed Chinese hamster embryo cells (Co631) contain about five viral copies integrated per cell genome. These SV40 sequences were used as endogenous indicator genes to study the response of mammalian cells to radiation at the gene level. An increase in copy number was detected by dispersed cell blotting and Southern analysis in combination with specific DNA hybridization. All types of radiation tested induce a 15- to 25-fold amplification of SV40 sequences without producing intact virus. The amplification is dose dependent and increases with time after irradiation: a maximum effect is observed at Day 3 after alpha particle or uv exposure and at Day 6 after gamma-ray exposure. A RBE of 6 can be calculated for alpha particles if amplification rates at Day 3 are compared. However, when the maximum effect is considered independent of time, no difference between different types of radiation is observed. Southern blots of genomic DNA show that not all integrated SV40 sequences are amplified upon radiation. Amplified sequences are found either in restriction fragments of relatively high molecular weight or in unit size fragments. SV40 amplification is selective in that the amplification of other genes, e.g., of alpha-actin, dhfr (dihydrofolate reductase), and of two oncogenes of the ras family (Kirsten ras and Harvey ras), was below detection level.
Alpha Particles↗
12-O-Tetradecanoylphorbol-13-acetate (TPA)-induced gene sequences in human primary diploid fibroblasts and their expression in SV40-transformed fibroblasts.
We have isolated cDNA sequences from TPA-treated primary human fibroblasts, which indicate RNA species that are coordinately regulated after treatment of these cells with either ultraviolet light, mitomycin C, the UV-induced factor EPIF, or TPA. The levels of RNA are elevated in Bloom syndrome (cells of two out of three patients). After transformation with SV40 one of the sequences is overexpressed while another one is reduced. Both genes maintain their inducibility by the agents mentioned.
Bloom Syndrome↗
Double-strand break repair and G2 block in Chinese hamster ovary cells and their radiosensitive mutants.
Two X-ray-sensitive mutants of the CHO K1 cell line were examined for their cell-cycle progression after irradiation with gamma-rays, and for their ability to rejoin double-strand breaks (DSBs) as detected by neutral filter elution. Both mutants were impaired in DSB rejoining and both were irreversibly blocked in the G2 phase of the cell cycle as determined by cytofluorometry. From one mutant we have isolated several revertants. The revertants stem from genomic DNA transfection experiments and may have been caused by gene uptake. All revertants survived gamma-irradiation as did the wild-type CHO line. One of them has been examined for its ability to rejoin DSBs and was found to be similar to the wild type.
Animals↗
Mouse mammary tumour virus: a proviral gene contributes to the understanding of eukaryotic gene expression and mammary tumorigenesis.
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Animals↗
Genetic changes in mammalian cells reminiscent of an SOS response.
Prior to the isolation of mammalian DNA repair genes and identification of their gene products, the comparison between the bacterial SOS response and various similar reactions in mammalian cells remains rather speculative. The increasing number of observed phenomena including enhanced DNA repair, virus induction, induced cellular differentiation, and neoplastic transformation, all following DNA damage or arrest of replication, are, however, suggestive of an SOS-like system of growth control and may form an entry into this fascinating area.
Animals↗
In vitro template activity of 0.3 mRNA from wild type and initiation mutants of bacteriophage T7.
Bacteriophage T7 0.3 mRNA synthesised and processed in vitro has been purified starting from the DNA of T7+ as well as from that of two initiation mutants of T7 (CR17 with a U----C transition in the initiation codon and CR35b whose potential Shine and Dalgarno (S-D) interaction is interrupted by a G----A transition). These mRNAs were used as templates to direct the binding of fMet-tRNA and the synthesis of 0.3 protein in both E. coli and wheat germ cell-free systems. The initiation codon mutant displayed approximately 50% inhibition of fMet-tRNA binding and 0.3 protein synthesis in both systems. The S-D sequence mutant, on the other hand, was found to be less affected than the initiation triplet mutant (20%-40% inhibition) in both fMet-tRNA binding and template activity in the E. coli system. In the wheat germ system, which does not make use of the S-D interaction, however, this mutant displayed normal template activity suggesting that the inhibition obtained in the E. coli system, albeit slight, is due to the impairment of the S-D interaction and not to an alteration of the mRNA secondary or tertiary structure caused by the base substitution.
Cell-Free System↗
Glucocorticoid hormone interactions with cloned proviral DNA of mouse mammary tumor virus.
The molecular details of glucocorticoid hormone regulation of expression of the mouse mammary tumor virus (MMTV) proviral gene have been investigated. Cloned proviral DNA was introduced into cultured cells by a gene transfer procedure. DNA acquired by transfection was shown to be expressed in a hormone regulated fashion. The proviral DNA was fragmented and recombined in vitro with an indicator gene to delimit the hormone response sequence. Inducibility of the indicator gene (thymidine kinase gene from Herpes Simplex Virus, tk) was observed upon recombination with the long terminal repeat (LTR) sequence of MMTV. Further delimitation of the LTR DNA demonstrated that 202 nucleotides located 5' of the RNA initiation site are sufficient to confer glucocorticoid regulation. In vitro interaction of LTR DNA with glucocorticoid hormone receptor complex, showed a preferential affinity to the same sequence which mediated hormonal regulation in transfected cells. Evidence for a direct receptor gene interaction in the process of gene induction was gained by the measurement of the kinetics of induction and the use of a glucocorticoid antagonist (RU 486). The induction of the transfected gene is very rapid, independent of simultaneous protein synthesis and requires a functional glucocorticoid receptor hormone complex.
Animals↗
UV-induced extracellular factor from human fibroblasts communicates the UV response to nonirradiated cells.
Ultraviolet light enhances the synthesis of at least eight abundant proteins in human fibroblasts within 2 hr. These proteins are identical with those induced by the tumor promoter TPA. The inducing signal is generated by DNA damage, as these proteins are induced by lower doses of UV in fibroblasts from patients with Cockayne's syndrome or Xeroderma pigmentosum. In the supernatant of UV-treated cells, a heat-labile ammonium sulfate precipitable factor of more than 10 kd (EPIF) was detected which, upon transfer to nonirradiated cells, mimicked UV in the UV-induced synthesis of gene products. The response to UV, TPA, or EPIF was inhibited by fluocinolone acetonide, but not by retinoic acid, protease inhibitors, or superoxide dismutase.
Cell Communication↗
Regulation of MHC class II invariant chain expression: induction of synthesis in human and murine plasmocytoma cells by arresting replication.
The expression of the H2 Ia-associated invariant chain (Ii) has been determined by pulse labeling cells with [35S]methionine and resolving the proteins. Expression is maximal in noncycling peripheral B lymphocytes and is reduced upon maturation of B lymphocytes to plasma cells. B cell-derived cell lines behave correspondingly: IgM+ non-secretor cell lines synthesize Ii while plasmocytoma cells do not. Pre-B cell lines are also negative. In all Ii-negative cell lines, the synthesis of Ii is selectively induced by treating the cells with inhibitors of replication.
Animals↗