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C Dickson

Publications and source records attributed to C Dickson.

At least 91 records · Page 5Linked to original sources
PubMed1990

Subcellular fate of the int-2 oncoprotein is determined by choice of initiation codon.

Fibroblast growth factors (FGFs) have been implicated in many aspects of cell growth and differentiation both in normal and neoplastic settings. For example, the mouse int-2 gene, which encodes an FGF-related product, is a frequent target of proviral activation in carcinomas induced by mouse mammary tumour virus, but apparently functions at discrete stages of normal embryonic development. Six classes of int-2 messenger RNA have been identified in embryonic cells, each of which is predicted to encode the same 245-amino-acid protein. But all known int-2 transcripts include sequences upstream of the AUG codon presumed to be the initiation codon. Here we report an additional N-terminally extended int-2 gene product initiated at an in-frame CUG codon. In COS-1 cells transiently transfected with appropriate int-2 complementary DNAs, the AUG-initiated product is found predominantly in the secretory pathway, whereas the CUG-initiated form is localized to the nucleus. These data indicate that the Int-2 oncoprotein could influence cellular behaviour by two distinct mechanisms.

Amino Acid Sequence↗
PubMed1990

The int-2 gene product acts as an epithelial growth factor in transgenic mice.

The induction of mammary tumors by mouse mammary tumor virus (MMTV) is thought to occur through proviral activation of one or more cellular genes. One of these, int-2, encodes a 27 kd protein which exhibits striking homology to the basic fibroblast growth factor family. To assess directly the role of the int-2 protein in cell proliferation, we have established transgenic mice which carry the int-2 gene driven by the MMTV promoter/enhancer. Expression of the int-2 gene in female transgenic mice results in pronounced mammary gland hyperplasia. Interestingly, expression of the MMTV-int-2 transgene in the prostate gland of male carriers results in a benign, but dramatic, epithelial hyperplasia similar to benign prostatic hypertrophy (BPH), a common but poorly understood disorder in human populations. Together, these results indicate that the int-2 product can act as a potent growth factor in these epithelial tissues.

Animals↗
PubMed1990

Tetravalent vanadium mediated oxidation of low density lipoprotein.

1. Tetravalent vanadium causes oxidation of low density lipoprotein (LDL) as manifest by protein degradation and lipid peroxidation. 2. Oxidative modification of the apolipoprotein B-100 is paralleled by the formation of thiobarbituric acid reactive substance and fluorescent chromolipid production. 3. The metal chelators ethylenediamine tetracetic acid and desferrioxamine, and the alcohols, ethanol and isopropanol inhibit the oxidation of LDL by tetravalent vanadium. No inhibition is observed with superoxide dismutase, catalase or mannitol. 4. The data suggest that aldehydes formed during the process of lipid peroxidation induced by tetravalent vanadium react with the proteins in LDL to form fluorescent chromolipids and that the oxidative process originates within the hydrophobic domain of LDL.

1-Propanol↗
PubMed1990

Gene amplification on chromosome band 11q13 and oestrogen receptor status in breast cancer.

We have analysed DNA from 183 primary breast cancers for amplification or rearrangement of a number of cellular proto-oncogenes, focusing primarily on a cluster of markers on the long arm of chromosome 11. Two of these oncogenes, INT2 and HST1, both of which encode members of the fibroblast growth factor family, are implicated in the generation of virally induced mammary tumours in mice. Here we confirm earlier reports that the q13 region of chromosome 11, in which these genes are tandemly linked, is modestly amplified in approximately 15% of primary human breast cancers. This amplification is confined, with one exception, to cases in which the oestrogen receptor (ER) levels are in excess of 20 fmol/mg protein (P = 0.001). However, DNA amplification does not usually result in detectable expression of either the INT2 or HST1 gene. The data imply that some other gene in the vicinity must contribute to the development of a subset of ER-positive tumours and that assessing the amplification of this region of DNA may be of value in defining a separate category of ER-positive tumour.

Adult↗
PubMed1990

Proviral insertions within the int-2 gene can generate multiple anomalous transcripts but leave the protein-coding domain intact.

We examined the effects of mouse mammary tumor virus integration on the multiple RNA transcripts expressed from the int-2 proto-oncogene in virally induced breast tumors. Proviral insertion either upstream or downstream of the gene could simultaneously activate transcription from three dissimilar int-2 promoters. In some tumors, the activating provirus lies within the transcription unit and disrupts the structures of the various RNAs. Insertions in the 5' region of the gene had complex effects depending on the orientation and position of the provirus relative to the three promoters and intron-exon boundaries. RNase protection experiments identified transcripts initiated in the viral long terminal repeat, at normal and cryptic sites in the int-2 sequences, and from cryptic promoters in an inverted provirus. AT the 3' end, insertions occurred within the untranslated trailer and provided alternative termination signals that substituted for one or both of the normal the poly(A) addition sites. However, in no instance, of the 20 tumors analyzed in detail, did a provirus perturb the presumed open reading frame of the gene. These data strongly implicate the normal product of the int-2 gene, which is related to the fibroblast growth factor family, as a contributory factor in virally induced mammary tumors.

Animals↗
PubMed1990

Int-2: a member of the fibroblast growth factor family has different subcellular fates depending on the choice of initiation codon.

The int-2 gene, which encodes a member of the fibroblast growth factor family, was discovered as a protooncogene transcriptionally activated following proviral insertion into adjacent chromosomal DNA. Analyses of the synthesis and processing of the int-2 protein, using an SV40-based vector to express cloned cDNA, showed four major products in the size range 27.5-31.5 kd that were associated with the secretory pathway. Further experiments using a cell-free translation system programmed with int-2 cRNA revealed a larger N-terminally extended protein. Site-directed mutagenesis of possible initiation codons confirmed that the first in-frame AUG codon would specify the start of a protein that includes a signal peptide for transport into the endoplasmic reticulum. However, protein synthesis also initiates at an upstream CUG codon to yield a polypeptide extended at the N-terminus by 29 amino acids. Immunofluorescent staining showed that a substantial proportion of the CUG-initiated protein resides in the cell nucleus, while a truncated int-2, lacking both the N-terminal extension and the signal peptide, was exclusively nuclear. These observations suggest that a nuclear localisation signal occurs in the body of the int-2 molecule, but is only accessible to the nuclear transport system if entry to the secretory pathway is compromised. Thus, the choice of initiation codon changes the subcellular fate of the int-2 protein and provides the potential for a duality of function through alternative transport pathways.

Animals↗
PubMed1990

Characterization of int-2: a member of the fibroblast growth factor family.

int-2 was discovered as a proto-oncogene transcriptionally activated by MMTV proviral insertion during mammary tumorigenesis in the mouse. Sequence analysis showed int-2 to be a member of the fibroblast growth factor family of genes. In normal breast and most other adult mouse tissues, int-2 expression was not detected except for low levels in brain and testis. However, using in situ hybridization, expression was found at a number of sites during embryonic development, from day 7 until birth. An analysis of the int-2 transcripts found in embryonal carcinoma cells revealed six major classes of RNA initiating at three promoters and terminating at either of two polyadenylation sites. Despite the transcriptional complexities, all size classes of RNA encompass the same open reading frame. Using an SV40 early promoter to drive transcription of an int-2 cDNA in COS-1 cells, several proteins were observed. These were shown to be generated by initiation from either of two codons: One, a CUG, leads to a product which localizes extensively to the cell nucleus and partially to the secretory pathway. In contrast, initiation at a downstream AUG codon results in quantitative translocation across the endoplasmic reticulum and the accumulation of products ranging in size from 27.5 x 10(3) Mr to 31.5 x 10(3) Mr in organelles of the secretory pathway. These proteins represented glycosylated and non-glycosylated forms of the same primary product with or without the signal peptide removed. These findings suggest the potential for a dual role of int-2; an autocrine function acting at the cell nucleus, and a possible paracrine action through a secreted product.

Amino Acid Sequence↗
PubMed1990

Absence of activating transmembrane mutations in the c-erbB-2 proto-oncogene in human breast cancer.

The rat neu proto-oncogene, which is a putative growth factor receptor closely related to the epidermal growth factor receptor, can be activated in vivo by a single point mutation in the sequence encoding its transmembrane region. The human homologue of neu, c-erbB-2, can be activated in vitro to an oncogenic form by point mutations in the same relative position in the gene. We have sought the presence of such activating mutations in a series of 100 cases of human breast cancer by polymerase chain reaction amplification and sequence-specific oligonucleotide hybridization, and also by a designed restriction fragment length polymorphism strategy in cases with Southern blot evidence of c-erbB-2 amplification. No evidence of activating point mutations in the c-erbB-2 protooncogene was found in any of these cases.

Base Sequence↗
PubMed1990

The mouse int-2 gene exhibits basic fibroblast growth factor activity in a basic fibroblast growth factor-responsive cell line.

The int-2 protein is related to basic fibroblast growth factor (bFGF) by amino acid sequence homology. To assess its biological activity, we constructed retroviral vectors containing four variants of mouse int-2 complementary DNA under the transcriptional control of the beta-actin promoter and tested their effects on human SW13 adrenal cortical tumor cells. This cell line specifically requires bFGF, interleukin 1, or transforming growth factor e for anchorage-independent growth in soft agar. Despite encoding a signal sequence that should direct the protein to the secretory pathway, vectors containing unmodified int-2 complementary DNA, or a form optimized for translation initiation at the AUG codon, were incapable of inducing SW13 growth in soft agar. However, SW13 transfectants expressing a construct (pSP1), in which a mouse immunoglobulin signal peptide sequence is linked to the int-2 coding sequences, grew well in soft agar. The concentrated conditioned medium from these pSP1-transfected cells supported anchorage-independent growth of SW13 indicator cells and competed with bFGF for binding to receptors. Western blot analysis with an int-2-specific antiserum detected Mr 30,000-32,000 int-2 products in cell extracts and conditioned medium from pSP1-transfected clones, whereas the conditioned medium from these and other SW13 clones contained only low levels of bFGF as measured in a specific radioimmunoassay. These data suggest that the product of the int-2 gene can functionally replace bFGF in modulating the anchorage-independent growth of SW13 cells.

Amino Acid Sequence↗
PubMed1989

The mouse homologue of hst/k-FGF: sequence, genome organization and location relative to int-2.

We have located and sequenced the murine homologue of the hst/k-FGF oncogene in genomic DNA clones that extend 3' from int-2 on mouse chromosome 7. The two genes are in the same transcriptional orientation, less than 20 kilobase pairs (kb) apart, and presumably evolved by tandem duplication of a common ancestral gene. RNase mapping and primer extension analyses indicated that the major 3.2 kb hst transcript expressed in undifferentiated embryonal carcinoma (EC) cell lines initiates at a unique cap site downstream from an obvious TATA-box. The 3' end of the transcript as identified in multiple cDNA clones occurs at an appropriate distance from a variant polyadenylation signal, ATTAAA. Translation of the major open reading frame would yield a 202 amino acid protein that is 82% homologous to human HST but lacking 4 residues at the presumed signal peptide cleavage site.

Amino Acid Sequence↗
PubMed1989

The structure and function of the int-2 oncogene.

The classification of int-2 as a growth factor is based primarily on the similarities between the predicted amino acid sequence and that of basic fibroblast growth factor (bFGF), as well as other members of this expanding family of related proteins. In this review, we summarise the background to the identification of int-2 as a proto-oncogene in virally induced mouse mammary tumours and describe key features of the structure and expression of both the mouse and human homologues. The normal sites of int-2 expression include specific embryonic cell types suggesting multiple inductive or morphogenetic roles. Recent progress in the characterisation of the int-2 product will be discussed in relation to the similarities and differences between int-2 and other FGFs.

Amino Acid Sequence↗
PubMed1989

The mouse homolog of the hst/k-FGF gene is adjacent to int-2 and is activated by proviral insertion in some virally induced mammary tumors.

The fibroblast growth factor-related protooncogenes, int-2 and hst/k-FGF, are within 17 kilobase pairs of one another on mouse chromosome 7 and are in the same transcriptional orientation. Approximately 70% of tumors induced in BR6 mice by mouse mammary tumor virus have proviral insertions adjacent to the int-2 gene. We find that the murine homolog of the hst/k-FGF gene can also be transcriptionally activated by the insertion of mouse mammary tumor virus DNA either upstream or downstream of the gene. In most tumors, only one of these adjacent genes is activated, but in some cases both genes are expressed. One of the hst-expressing tumors also has a virally activated int-3 gene. At least five distinct cellular genes (int-1, -2, -3, -4, and hst/k-FGF) can therefore contribute, either singly or in concert, to the development of histologically indistinguishable mammary tumors in mice infected by mouse mammary tumor virus.

Animals↗
PubMed1989

A putative int domain for mouse mammary tumor virus on mouse chromosome 7 is a 5' extension of int-2.

We extended the physical map of the mouse int-2 locus by demonstrating that the site of insertion for mouse mammary tumor virus DNA in plaque-type mammary tumors of GR mice is directly linked to int-2. An additional example of proviral integration is described in which a provirus in a presumed enhancer-insertion mode 15 kilobases upstream of the int-2 promoters is capable of activating expression of the gene at levels typical of other virally induced mammary tumors.

Animals↗
PubMed1989

Detection and characterization of the fibroblast growth factor-related oncoprotein INT-2.

Products of the fibroblast growth factor-related proto-oncogene int-2 have been detected by using a monoclonal antibody and polyclonal antisera raised against synthetic peptides predicted from the DNA sequence. COS-1 monkey cells transfected with int-2 DNA linked to the simian virus 40 early promoter contained at least four int-2-specific proteins, presumably representing modified forms of the expected 27-kilodalton primary translation product. The level of expression was increased approximately six- to eightfold by mutation of sequences around the presumed initiation codon, negating their capacity to encode a short oligopeptide in the +1 reading frame. Both tunicamycin inhibition and in vitro translation experiments indicated that some of the modifications correspond to asparagine-linked glycosylation, for which the sequence predicts a single site. In line with the similarities between INT-2 and other fibroblast growth factors, the in vitro translation products functioned as weak mitogens for mammary epithelial cells.

Amino Acid Sequence↗
PubMed1989

Sequence organization of the human int-2 gene and its expression in teratocarcinoma cells.

We report the genomic organization and DNA sequence of the human homologue of int-2, a proto-oncogene implicated in virally induced mammary tumours in the mouse, and expressed at specific sites and times during embryogenesis. Direct comparisons with the coding domains of mouse int-2 allowed us to delineate the intron-exon boundaries of the human gene. These boundaries were subsequently confirmed by ribonuclease protection analyses of the single 1.7 kilobase (kb) int-2 transcript detectable in the human teratocarcinoma cell line, Tera-2. The data suggest that human int-2 may also function in embryonic lineages but that its transcription may be less complex than in the mouse. The predicted human protein comprises 239 amino acids and is 89% homologous to its murine counterpart, except at the carboxy terminus. This divergence occurs distal to the region of int-2 that shows homology to other members of the FGF family of growth modulators and oncogenes.

Amino Acid Sequence↗
PubMed1988

Expression of the FGF-related proto-oncogene int-2 during gastrulation and neurulation in the mouse.

The proto-oncogene int-2 has been implicated in the formation of mouse mammary-tumour-virus-induced mammary tumours. Analysis of the predicted coding sequence indicates that int-2 is a member of the fibroblast growth factor family. Previous studies using Northern blot analysis suggested that normal expression of int-2 may be confined to extra-embryonic endoderm lineages of embryonic stages of mouse development. We have used in situ hybridization and Northern blot analysis to examine directly int-2 expression in embryo stem cells and in the developing embryo from early gastrulation to midsomite stages. Complex patterns of accumulation of int-2 RNA were observed in embryonic and extra-embryonic tissues. The data suggest multiple roles for int-2 in development which may include migration of early mesoderm cells and induction of the otocyst.

Animals↗