Showing papers on "Gene expression published in 1985"

Identification of DNA sequences required for activity of the cauliflower mosaic virus 35S promoter

Abstract: Although promoter regions for many plant nuclear genes have been sequenced, identification of the active promoter sequence has been carried out only for the octopine synthase promoter. That analysis was of callus tissue and made use of an enzyme assay. We have analysed the effects of 5' deletions in a plant viral promoter in tobacco callus as well as in regenerated plants, including different plant tissues. We assayed the RNA transcription product which allows a more direct assessment of deletion effects. The cauliflower mosaic virus (CaMV) 35S promoter provides a model plant nuclear promoter system, as its double-strand DNA genome is transcribed by host nuclear RNA polymerase II from a CaMV minichromosome. Sequences extending to -46 were sufficient for accurate transcription initiation whereas the region between -46 and -105 increased greatly the level of transcription. The 35S promoter showed no tissue-specificity of expression.

Cloning and expression of the human erythropoietin gene.

Abstract: The human erythropoietin gene has been isolated from a genomic phage library by using mixed 20-mer and 17-mer oligonucleotide probes. The entire coding region of the gene is contained in a 5.4-kilobase HindIII-BamHI fragment. The gene contains four intervening sequences (1562 base pairs) and five exons (582 base pairs). It encodes a 27-amino acid signal peptide and a 166-amino acid mature protein with a calculated Mr of 18,399. The erythropoietin gene, when introduced into Chinese hamster ovary cells, produces erythropoietin that is biologically active in vitro and in vivo.

Trans-activator gene of human T-lymphotropic virus type III (HTLV-III).

Abstract: Human T-lymphotropic virus type III (HTLV-III) encodes a trans-acting factor that activates the expression of genes linked to the HTLV-III long terminal repeat. By functional mapping of complementary DNA transcripts of viral messenger RNA's the major functional domain of the gene encoding this factor was localized to a region immediately before the env gene of the virus, a region previously thought to be noncoding. This newly identified gene consists of three exons, and its transcription into messenger RNA involves two splicing events bringing together sequences from the 5' part (287 base pairs), middle (268 base pairs), and 3'part (1258 base pairs) of the HTLV-III genome. A similar messenger RNA with a truncated second exon (70 base pairs) does not encode a trans-acting function. It is proposed that this second messenger RNA is the transcript of a gene (3'-orf) located after the env gene. Messenger RNA's were also identified for the env and gag-pol genes of HTLV-III.

Plasticity of the differentiated state.

Abstract: Heterokaryons provide a model system in which to examine how tissue-specific phenotypes arise and are maintained. When muscle cells are fused with nonmuscle cells, muscle gene expression is activated in the nonmuscle cell type. Gene expression was studied either at a single cell level with monoclonal antibodies or in mass cultures at a biochemical and molecular level. In all of the nonmuscle cell types tested, including representatives of different embryonic lineages, phenotypes, and developmental stages, muscle gene expression was induced. Differences among cell types in the kinetics, frequency, and gene dosage requirements for gene expression provide clues to the underlying regulatory mechanisms. These results show that the expression of genes in the nuclei of differentiated cells is remarkably plastic and susceptible to modulation by the cytoplasm. The isolation of the genes encoding the tissue-specific trans-acting regulators responsible for muscle gene activation should now be possible.

Cell-specific expression of the rat insulin gene: evidence for role of two distinct 5' flanking elements

Abstract: The 59 flanking DNA of the rat insulin I gene contains sequences controlling cell-specific expression. Analysis of this region by replacement of specific portions with nondiscriminatory control elements from viral systems shows that a transcriptional enhancer is located in the distal portion of the 59 flanking DNA; its position has been mapped by deletion analysis. Additional experiments suggest that another distinct regulatory element is located more proximal to the transcription start site. The activity of both elements is restricted to pancreatic B cells. The combinatorial effect of multiple control elements could explain the cell-specific expression of insulin genes.

Molecular cloning of the complementary dna for human tumor necrosis factor

Abstract: Tumor necrosis factor (TNF) is a soluble protein that causes damage to tumor cells but has no effect on normal cells. Human TNF was purified to apparent homogeneity as a 17.3-kilodalton protein from HL-60 leukemia cells and showed cytotoxic and cytostatic activities against various human tumor cell lines. The amino acid sequence was determined for the amino terminal end of the purified protein, and oligodeoxyribonucleotide probes were synthesized on the basis of this sequence. Complementary DNA (cDNA) encoding human TNF was cloned from induced HL-60 messenger RNA and was confirmed by hybrid-selection assay, direct expression in COS-7 cells, and nucleotide sequence analysis. The human TNF cDNA is 1585 base pairs in length and encodes a protein of 233 amino acids. The mature protein begins at residue 77, leaving a long leader sequence of 76 amino acids. Expression of high levels of human TNF in Escherichia coli was accomplished under control of the bacteriophage lambda PL promoter and gene N ribosome binding site.

Expression in brain of a messenger RNA encoding a novel neuropeptide homologous to calcitonin gene-related peptide.

Abstract: As a consequence of alternative RNA processing events, a single rat gene can generate messenger RNA's (mRNA's) encoding either calcitonin or a neuropeptide referred to as alpha-type calcitonin gene-related peptide (alpha-CGRP). An mRNA product of a related gene has been identified in rat brain and thyroid encoding the protein precursor of a peptide differing from alpha-CGRP by only a single amino acid. The RNA encoding this peptide, which is referred to as beta-CGRP, appears to be the only mature transcript of the beta-CGRP gene. Hybridization histochemistry reveals a similar distribution of alpha- and beta-CGRP mRNA's, but their relative levels of expression vary in different cranial nerve nuclei. Thus beta-CGRP is a new member of a family of related genes with potential functions in regulating the transduction of sensory and motor information.

Molecular Cloning of a Complementary DNA Encoding Human Macrophage-Specific Colony-Stimulating Factor (CSF-1)

Abstract: Complementary DNA (cDNA) clones encoding human macrophage-specific specific colony-stimulating factor (CSF-1) were isolated. One cDNA clone codes for a mature polypeptide of 224 amino acids and a putative leader of 32 amino acids. This cDNA, which was cloned in the Okayama-Berg expression vector, specifies the synthesis of biologically active CSF-1 in COS cells, as determined by a specific radioreceptor assay, macrophage bone marrow colony formation, and antibody neutralization. Most of the cDNA isolates contain part of an intron sequence that changes the reading frame, resulting in an abrupt termination of translation; these cDNA's were inactive in COS cells. The CSF-1 appears to be encoded by a single-copy gene, but its expression results in the synthesis of several messenger RNA species, ranging in size from about 1.5 to 4.5 kilobases.

A Macrophage Factor Inhibits Adipocyte Gene Expression: An in Vitro Model of Cachexia

Abstract: Certain infections and malignancies in mammals cause the development of a condition known as cachexia in which the animal continues to lose weight, often while consuming an adequate diet. When macrophages are stimulated with an endotoxin, they produce a factor or factors, termed cachectin, that inhibits the activity of fat-producing (lipogenic) enzymes in cultured adipocytes. This effect may reflect one of the physiological bases for cachexia. In the present study, clones of complementary DNA from genes whose expression is increased during the differentiation of adipocytes were used to study the molecular basis of cachectin's actions. In the presence of cachectin, the expression of the corresponding genes was reversibly and specifically inhibited. Furthermore, when mature adipocytes were exposed to cachectin, the messenger RNA's of those genes diminished and rapidly approached the levels present before differentiation.

Cloning of firefly luciferase cDNA and the expression of active luciferase in Escherichia coli.

Abstract: A cDNA library was constructed from firefly (Photinus pyralis) lantern poly(A)+ RNA, using the Escherichia coli expression vector lambda gt11. The library was screened with anti-P. pyralis luciferase (Photinus luciferin:oxygen 4-oxidoreductase, EC 1.13.12.7) antibody, and several cDNA clones expressing luciferase antigens were isolated. One clone, lambda Luc1, contained 1.5 kilobase pairs of cDNA that hybridized to a 1.9- to 2.0-kilobase band on a nitrocellulose blot of electrophoretically fractionated lantern RNA. Hybridization of the cloned cDNA to lantern poly(A)+ RNA selected an RNA that directed the in vitro synthesis of a single polypeptide. This polypeptide comigrated with luciferase on NaDodSO4/PAGE and produced bioluminescence upon the addition of luciferin and ATP. A 1.8-kilobase-pair cDNA was isolated by probing the firefly cDNA library with the cDNA from lambda Luc1. This cDNA contained sufficient coding information to direct the synthesis of active firefly luciferase in E. coli.

Light Regulation of Gene Expression in Higher Plants

Abstract: In this review areas of currently active research are considered which have demonstrated that a plant's response to light involves changes in the expression of specific genes at the level of RNA. The regulation of gene expression by phytochrome and the UV-sensitive photoreceptor have been studied most extensively at the molecular level, and this review particularly focuses on such studies in higher plants. Some of the observations made on the differences in gene expression between light-grown and dark-grown plants are also included, although the photoreceptor(s) responsible for the differences may not have been ascertained. In some of these cases, phytochrome involvement has been or may be demonstrated in later studies, while in others the observed differences may be a result of the action of other photoreceptors or of multiple light-affected processes. One such process is the development of chloroplasts, a major developmental step triggered by light in angiosperms. In addition, many of the genes whose expression is changed by light and which have been studied at a molecular level encode chloroplast proteins. 156 references.

Identification of a set of genes expressed during the G0/G1 transition of cultured mouse cells.

Abstract: To identify previously undetected genes that may be involved in the transition from a resting state (G0) to a proliferative state (G1) of mammalian cells, we set out to isolate cDNA clones derived from mRNAs that appear in serum-stimulated cells in the absence of protein synthesis A lambda cDNA library was prepared using poly(A)+ RNA from BALB/c 3T3 cells that had been brought to quiescence and subsequently stimulated with serum in the presence of cycloheximide Approximately 50 000 recombinant phage plaques were screened, and 357 clones were isolated that hybridized to probes derived from stimulated-cell RNA but not to probes from resting-cell RNA Cross hybridization analysis showed that four RNA sequence families account for approximately 90% of these clones One of the clones hybridized to an actin probe; none hybridized to any of 13 oncogene probes tested Five different RNAs that appear to be previously uncharacterized have been further analyzed These RNAs accumulate and decay rapidly following stimulation by serum or purified growth factors, or by a tumor promoter, and they are superinduced by serum in the presence of cycloheximide Three of the RNAs could be enriched by hybridization to cDNAs and translated in vitro, yielding proteins of approximately 43, 40 and 35 kd, respectively

High level expression of introduced chimaeric genes in regenerated transformed plants

Abstract: Promoter DNA sequences from a petunia chlorophyll a/b binding protein gene were fused to octopine synthase DNA sequences and the resulting chimaeric genes were introduced into petunia and tobacco cells. Populations of transformed regenerated petunia plants containing the chimaeric genes were examined so that the expression of any particular construction could be compared between independent transformants. Substantial variation was observed between transformants in the level of chimaeric gene expression. In general, transcriptional fusions in which a linker sequence interrupted the 5'-untranslated region gave rise to less chimaeric mRNA accumulation than a translational fusion. In the most actively expressing transformants the amount of mRNA from the introduced chimaeric genes was half that of the endogenous wild-type gene. Transcription initiated at the same place in the chimaeric and endogenous genes. Construction of the translational cab/ocs fusion caused three amino acid changes in the octopine synthase protein and functional octopine synthase enzyme was absent from plants in which mRNA for the chimaeric gene was abundantly expressed.

Developmental regulation of T-cell receptor gene expression.

Abstract: In contrast to B cells or their antibody products, T lymphocytes have a dual specificity, for both the eliciting foreign antigen and for polymorphic determinants on cell surface glycoproteins encoded in the major histocompatibility complex (MHC restriction)1–4. The recent identification of T-cell receptor glycoproteins5–7 as well as the genes encoding T-cell receptor subunits will help to elucidate whether MHC proteins and foreign antigens are recognized by two T-cell receptors or by a single receptor. An important feature of MHC restriction is that it appears to be largely acquired by a differentiating T-cell population under the influence of MHC antigens expressed in the thymus8–10, suggesting that precursor T cells are selected on the basis of their reactivity with MHC determinants expressed in the host thymus9–11. To understand this process of ‘thymus education’, knowledge of the developmental regulation of T-cell receptor gene expression is necessary. Here we report that whereas messenger RNAs encoding the β-and γ-subunits are relatively abundant in immature thymocytes, α mRNA levels are very low. Interestingly, whereas α mRNA levels increase during further development and β mRNA levels stay roughly constant, γ mRNA falls to very low levels in mature T cells, suggesting a role for the γ gene in T-cell differentiation.

Herpes simplex virus type 1 ICP27 is an essential regulatory protein.

Abstract: The five immediate-early genes of herpes simplex virus are expressed during the initial stages of the infectious cycle, and certain immediate-early proteins have been shown to play a regulatory role in subsequent viral gene expression. Until recently, the functional properties of only one immediate-early protein, ICP4, had been examined in any detail, primarily because mutants had been isolated only in the gene for ICP4. We report herein the genetic and phenotypic characterization of four temperature-sensitive mutants of herpes simplex virus type 1 (tsY46, tsE5, tsE6, and tsLG4) that have begun to elucidate the function(s) of a second immediate-early protein, ICP27. The four mutants complemented each other inefficiently or not at all, indicating that they are defective in the same function. Marker rescue tests placed the mutations in tsY46 and tsE5 in sequences that encode the transcript for ICP27; the mutations in tsE6 and tsLG4 lie in or near these sequences. The ability of wild-type ICP27 expressed from a cloned gene to complement tsY46 and tsLG4 constitutes additional evidence that these mutants are defective in an ICP27-associated function. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of mutant-infected cell polypeptides showed that certain immediate-early (alpha) polypeptides were overproduced, whereas significant levels of early (beta) and drastically reduced levels of several late (gamma) proteins were synthesized at the nonpermissive temperature. Interestingly, the mutants were observed to form a spectrum with regard to their relative abilities to induce the expression of a number of polypeptides, especially those of the delayed-early (beta gamma) class. Consistent with their ability to induce expression of early polypeptides, all of the mutants induced the synthesis of substantial levels of viral DNA at the nonpermissive temperature. Taken together, the results of these studies demonstrate that ICP27 plays an essential regulatory function in virus replication, that this function is required after the onset of early gene expression and viral DNA synthesis, and that the inability of the mutants to induce the synthesis of late proteins is independent of viral DNA synthesis.

Expression of the Rous Sarcoma Virus pol Gene by Ribosomal Frameshifting

Abstract: The pol gene of Rous sarcoma virus is positioned downstream of the gag gene in a different, briefly overlapping reading frame; nevertheless, the primary translation product of pol is a gag-pol fusion protein. Two mechanisms, ribosomal frameshifting and RNA splicing, have been considered to explain this phenomenon. The frameshifting model is supported by synthesis of both gag protein and gag-pol fusion protein in a cell-free mammalian translation system programmed by a single RNA species that was synthesized from cloned viral DNA with a bacteriophage RNA polymerase. Under these conditions, the ratio of the gag protein to the fusion protein (about 20 to 1) is similar to that previously observed in infected cells, the frameshifting is specific for the gag-pol junction, and it is unaffected by large deletions in gag. In addition, synthesis of the fusion protein is ten times less efficient in an Escherichia coli cell-free translation system and cannot be explained by transcriptional errors or in vitro modification of the RNA. Ribosomal frameshifting may affect production of other proteins in higher eukaryotes, including proteins encoded by several retroviruses and transposable elements.

Superinduction of c-fos by nerve growth factor in the presence of peripherally active benzodiazepines.

Abstract: Alterations in proto-oncogene expression after stimulation of rat pheochromocytoma (PC12) cells by nerve growth factor (NGF) have been investigated. A specific stimulation of c-fos messenger RNA and protein was detected 30 minutes after treatment. This induction was enhanced more than 100-fold in the presence of peripherally active benzodiazepines. The effect was specific as very little change was observed in the levels of c-rasHa, c-rasKi, c-myc, and N-myc messenger RNA's. Under the conditions used here, NGF treatment ultimately results in neurite outgrowth, with a reduction or cessation of cell division. Thus, stimulation of the c-fos gene in this system appeared to be associated with differentiation and not with cellular proliferation. The effect of benzodiazepines was stereospecific and represents a novel action of these compounds at the level of gene expression.

Cloning and expression in Escherichia coli of the gene for human tumour necrosis factor

Abstract: Tumour necrosis factor (TNF) was found originally in mouse serum after intravenous injection of bacterial endotoxin into mice primed with viable Mycobacterium bovis, strain Bacillus Calmette-Guerin (BCG). TNF-containing serum from mice is cytotoxic or cytostatic to a number of mouse and human transformed cell lines, but less or not toxic to normal cells in vitro. It causes necrosis of transplantable tumours in mice. TNF also occurs in serum of rat, rabbit and guinea pig. Rabbit TNF has been purified recently to give a single band on SDS-polyacrylamide gel electrophoresis (PAGE). The purified TNF had a relative molecular mass (Mr) 40,000 +/- 5,000 measured by gel filtration, and 17,000 by SDS-PAGE. Its isoelectric point is 5.0 +/- 0.3. The necrotic activity in vivo and the cytotoxicity in vitro are produced by the same substance. The gene encoding TNF has been identified in a human genomic DNA library using as a probe a cloned cDNA encoding a portion of rabbit TNF. The regions of this gene encoding an amino-acid sequence corresponding to mature TNF have been expressed in Escherichia coli and the product of this expression isolated in pure form and shown to produce necrosis of murine tumours in vivo.

Abrogation of metastatic properties of tumour cells by de novo expression of H-2K antigens following H-2 gene transfection

Abstract: H–2 gene transfection was used to restore expression of H–2K antigens in metastatic and non-metastatic subclones of a murine fibrosarcoma that lack their major histocompatibility complex-encoded H–2K antigens. De novo expression of H–2K reduced tumorigenicity and abolished the formation of metastasis in syngeneic mice. Expression of H–2K may lead to effective recognition of the disseminating tumour cells by the host immune system.

Pretranslational modulation of acute phase hepatic protein synthesis by murine recombinant interleukin 1 (IL-1) and purified human IL-1.

Abstract: During the acute phase response to tissue injury or inflammation, the concentration of several plasma proteins change. Previous work (29-34) suggested a role for interleukin 1 (IL-1) in the acute phase response. The availability of recombinant-generated mouse IL-1 prompted a study designed to directly test the function of IL-1 and its mechanism of action on hepatic synthesis of two positive acute phase proteins (serum amyloid A [SAA] and complement factor B), and a negative acute phase reactant (albumin). Intravenous injection of purified recombinant-generated murine-IL-1 into C3H/HeJ endotoxin-resistant mice induced a dose-dependent increase in SAA-specific hepatic messenger RNA (mRNA), and an increase in SAA plasma protein concentration. In primary murine hepatocyte cultures, both the recombinant IL-1 and highly purified human IL-1 induced a dose- and time-dependent, reversible increase in expression of the SAA and factor B genes, and a decrease in albumin gene expression. This regulation is pretranslational, since the kinetics and direction of change in specific mRNA for SAA, factor B, and albumin correspond to the changes in synthesis of the respective proteins. Moreover, the effect of IL-1 was specific, since actin gene expression was unaffected, and the IL-1 response was inhibited by antibody specific for IL-1. These data provide direct evidence that a single mediator, IL-1, can effect the positive and negative changes in specific hepatic gene expression characteristic of the acute phase response.

Constitutive and conditional suppression of exogenous and endogenous genes by anti-sense RNA

Abstract: Plasmid DNA directing transcription of the noncoding (anti-sense) DNA strand can specifically inhibit the expression of several test genes as well as normal, endogenous genes. The anti-sense plasmid constructions can be introduced into eukaryotic cells by transfection or microinjection and function in both transient and stable transformation assays. Anti-sense transcripts complementary to as little as 52 bases of 5' untranslated target gene mRNA specifically suppress gene activity as well as, or more efficiently than, anti-sense transcripts directed against the protein coding domain alone. Conditional anti-sense inhibition is accomplished with the use of hormone-inducible promoter sequences. Suppression of endogenous actin gene activity by anti-sense RNA is detected as a decrease in growth rate and as a reduction in the number of actin microfilament cables. These observations suggest that anti-sense RNA may be generally useful for suppressing the expression of specific genes in vivo and may be a potential molecular alternative to classical genetic analysis.

Erythroid-specific expression of human beta-globin genes in transgenic mice.

Abstract: Transgenic mice carrying human beta-globin genes were produced by microinjecting linear DNA molecules containing cloned beta-globin genes with up to 4300 bp of 5'-flanking sequence and 1700 bp of 3'-flanking sequence. Most (15 of 20) of these transgenic mice expressed the human beta-globin genes in blood cells and the level of expression in some mice was comparable with that obtained from endogenous beta-globin genes. Human beta-globin gene expression appeared to be restricted to cells of the erythroid lineage and was first detected between 11 and 14 days of development, in parallel with mouse beta-globin. Constructs with as little as 48 bp of 5'-flanking sequence also appeared to be expressed appropriately. The mRNA transcripts had correct 5' ends and directed human beta-globin synthesis in reticulocyte lysates. Human beta-globin protein was detectable in mature erythrocytes from progeny of one of these mice. The frequency and extent of expression was severely depressed when the procaryotic vector DNA was not removed prior to microinjection.

Insulin-like growth factor-II gene expression in Wilms' tumour and embryonic tissues

Abstract: Wilms' tumour (nephroblastoma) is an embryonal neoplasm occurring in hereditary and spontaneous forms. Both types show rearrangements of the short arm of chromosome 11. The germ line of children with the rare inherited triad of aniridia, genitourinary abnormality and mental retardation carry a chromosome 11 that has a deletion in its short arm (band 11p13) and these children are at increased risk of developing Wilms' tumour1,2. Neonates with the Beckwith–Wiedemann syndrome, in which there may be duplication of the 11p13–11p15 region, are similarly predisposed3. In the spontaneous form of the tumour a deletion of the Hp14 band in tumour cells, but not in normal cells, has been reported4, and the development of homozygosity for recessive mutations in the 11p region is implicated in the aetiology of Wilms' tumour5–8. In view of these chromosomal rearrangements and because Wilms' tumour is historically indistinguishable from the early stages of kidney development9, we have now examined the expression of genes localized to 11p in Wilms' tumour and human embryonic tissue. In 12 sporadic tumours examined, the expression of the gene coding for insulin-like growth factor-II (IGF-II), localized to the 11p15 region, was markedly increased relative to adult tissues, but was comparable to the level of expression in several fetal tissues including kidney, liver, adrenals and striated muscle. This may reflect the stage of tumour differentiation, but could also contribute to the malignant process, as IGF-II is an embryonal mitogen10–13.

Quantitative analysis of in situ hybridization methods for the detection of actin gene expression.

Abstract: We have implemented an efficient, quantitative approach for the optimization of in situ hybridization using double-stranded recombinant DNA probes. The model system studied was actin mRNA expression in chicken embryonic muscle cultures. Actin and control (pBR322) probes were nick-translated with p32 labeled nucleotides, hybridized to cells grown on coverslips, and quantitated in a scintillation counter. Cellular RNA retention was monitored via the incorporation of H3-Uridine into RNA prior to cell fixation. Over a thousand samples were analyzed, and among the technical variables examined were the fixation protocol, proteolytic cell pretreatment, the time course of hybridization, saturation kinetics, hybridization efficiency, and effect of probe size on hybridization and network formation. Results have allowed us to develop a reproducible in situ hybridization methodology which is simpler and less destructive to cellular RNA and morphology than other protocols. Moreover, this technique is highly sensitive and efficient in detection of cellular RNAs. Lastly, the rapid quantitative approach used for this analysis is valuable in itself as a potential alternative to filter or solution hybridizations.

Cloning and expression in Escherichia coli of the cDNA for murine tumor necrosis factor.

Abstract: A murine tumor necrosis factor (MuTNF) cDNA was isolated from a cDNA library prepared by using mRNA from the murine macrophage-like cell line PU5-1.8 induced with 4 beta-phorbol 12 beta-myristate 13 alpha-acetate. The cDNA encodes a polypeptide consisting of a 79 amino acid pre sequence followed by a mature MuTNF sequence of 156 amino acids. The 235 amino acid murine pre-TNF polypeptide is 79% homologous to the human pre-TNF protein. There is one potential N-linked glycosylation site on MuTNF, in contrast to human TNF, which lacks any such site. The MuTNF cDNA, when engineered for expression in Escherichia coli, was found to direct the synthesis of biologically active MuTNF as determined by its cytotoxicity against several transformed cell lines.

Rearrangements of the cellular p53 gene in erythroleukaemic cells transformed by Friend virus

Abstract: There is now good evidence that the cellular protein, p53, is involved in the transformation process, although its precise role is unknown1. It was reported recently that expression of the p53 gene can immortalize cells2 and that the p53 gene can replace the myc oncogene in a myc–ras immortalization/transformation assay3,4. We have investigated whether p53 is involved in the progression towards the neoplastic state in vivo and report here that erythroleukaemic cell lines transformed by different isolates of Friend leukaemia virus show altered expression of the cellular p53 gene. High levels of p53 protein are found in certain lines, but the protein is undetectable in others. This heterogeneity in p53 gene expression is associated with heterogeneity in tumorigenicity. We demonstrate that genomic rearrangements are responsible for p53 gene Jnactivation in these cell lines and that they occur in vivo during the natural progression of Friend virus-induced erythroleukaemia.

Cloning, sequence, and expression of a human granulocyte/macrophage colony-stimulating factor.

Abstract: Human granulocyte/macrophage colony-stimulating factor (GM-CSF) is a glycoprotein that is essential for the in vitro proliferation and differentiation of precursor cells into mature granulocytes and macrophages. In this report we have used a mouse GM-CSF cDNA clone to isolate human GM-CSF clones from libraries made from HUT-102 messenger RNA and mitogen-stimulated T-lymphocyte messenger RNA. The human cDNA clones contained a single open-reading frame encoding a protein of 144 amino acids with a predicted molecular mass of 16,293 daltons and showed 69% nucleotide homology and 54% amino acid homology to mouse GM-CSF. One of these cDNA clones was shown to direct the synthesis of biologically active GM-CSF using a yeast expression system. The gene for human GM-CSF appears to exist as a single-copy gene.