Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Pro-opiomelanocortin (POMC) is the common precursor of several pituitary hormones including alpha-melanotropic hormone, adrenocorticotropic hormone, beta-lipotropin and beta-endorphin. The porcine POMC cDNA was inserted downstream from the late promoter of an SV40-derived expression vector and co-transfected in NIH 3T3 cells with a marker plasmid carrying the neomycin resistance gene. Colonies resistant to the neomycin analog G418 were selected and analyzed for the production of POMC-related peptides by radioimmunoassay. Three clones were found to produce from 350 to 1750 pg of POMC-related peptides per 10(6) cells in 16 h and selected for further analysis. The number of POMC cDNA copies integrated in the host cell genome was determined and the levels of transcription were compared. POMC-related material released in the culture medium by the best producing clone (NJP 4-4) was further analyzed by gel filtration and reversed-phase high-pressure liquid chromatography combined with radioimmunoassays. POMC was found to be synthesized and secreted without further processing or degradation. Negligible amounts of POMC-immunoreactive species were found in cellular extracts indicating that the prohormone is secreted from the NIH 3T3 cells without storage, presumably through a constitutive pathway. Our results suggest that NIH 3T3 fibroblasts do not contain the enzymatic machinery to process complex precursors such as POMC.
Mol Cell Endocrinol 1988 Jul
PMID:Expression of porcine pro-opiomelanocortin cDNA in an established fibroblastic cell line: constitutive secretion of the precursor without proteolytic processing. 246 90

p53 anti-sense sequences were introduced into normal NIH3T3 and transformed CMS 4 cells by infection with the recombinant retrovirus carrying a repeat of the 5'-terminal fragment of p53 cDNA. Clones selected for G418 resistance showed a marked inhibition of proliferative capacity and a reduced ability to enter DNA replication after stimulation of quiescent cells with serum. Clones showing moderate inhibition of proliferation were shown to contain truncated anti-sense DNA integrated into the genome. The anti-sense DNA was transcribed and it correlated with the reduction of the p53 protein level in the cell clones studied. We conclude that the appropriate expression of p53 appears to be required for cell proliferation.
Mol Biol (Mosk)
PMID:[Antisense RNA p53 inhibits proliferation of normal and transformed cells]. 246 39

Murine papilloma cell lines 308 and SP-1 have been used as recipients for transfected oncogenes to investigate malignant conversion. These cell lines express an activated c-rasHa gene with a codon 61 mutation and produce squamous papillomas when transplanted as skin grafts onto nude mice. They are not tumorigenic by subcutaneous injection. Both papilloma cell lines were stably transfected with plasmid DNA containing either a rearranged murine plasmacytoma-derived c-myc (minus exon 1), adenovirus 5 E1A, FBJ v-fos or a human c-fos/FBJ v-fos chimera, using cotransfection with the neomycin resistance gene contained in pSV2neo to select for transformants. Southern and northern blotting analysis confirmed the uptake and expression of exogenous DNA in both G418-selected cell lines and in the derived tumors. Unlike the E1A- and myc-containing plasmids, both fos constructs caused malignant conversion in either cell line, as defined by the squamous cell carcinoma histology of tumors from grafted cells and the development of carcinomas after subcutaneous injection into athymic nude mice. Immunofluorescence analysis for specific keratin gene expression indicated that tumors derived by introduction of either of the fos oncogenes were devoid of staining for K1, a 67 kDa epidermal keratin that is expressed in papillomas but not in squamous carcinomas. Tumors from E1A, myc, or pSV2neo transfectants expressed K1, although in a focal distribution. The malignant phenotype induced by the fos oncogene constructs was not associated with the ability to form agar colonies in vitro or to express gamma-glutamyl transpeptidase in the tumors. Since both 308 and SP-1 were sensitive to the fos oncogene for malignant conversion and insensitive to E1A or myc, it is possible that fos may cooperate with the endogenous-activated c-rasHa gene to convert these cells to malignancy. However, since gamma-glutamyl transpeptidase activity is found in the majority of chemically induced mouse skin carcinomas that possess an activated c-rasHa gene, fos activation may not be a common pathway for spontaneous malignant conversion.
Mol Carcinog 1988
PMID:Malignant conversion of murine squamous papilloma cell lines by transfection with the fos oncogene. 247 37

We have compared the suppression of nonsense mutations by aminoglycoside antibiotics in Escherichia coli and in human 293 cells. Six nonsense alleles of the chloramphenicol acetyl transferase (cat) gene, in the vector pRSVcat, were suppressed by growth in G418 and paromomycin. Readthrough at UAG, UAA and UGA codons was monitored with enzyme assays for chloramphenicol acetyl transferase (CAT), in stably transformed bacteria and during transient expression from the same plasmid in human 293 tissue culture cells. We have found significant differences in the degree of suppression amongst three UAG codons and two UAA codons in different mRNA contexts. However, the pattern of these effects are not the same in the two organisms. Our data suggest that context effects of nonsense suppression may operate under different rules in E. coli and human cells.
Mol Gen Genet 1989 Jun
PMID:Aminoglycoside suppression at UAG, UAA and UGA codons in Escherichia coli and human tissue culture cells. 247 56

Sequences of the plant-pathogenic Ti-plasmid were found to be constitutively expressed in LTK- and in HeLa-cells. Activity of the nopaline-synthase (nos) promoter in these cells was demonstrated by directing expression of G418 resistance from a connected neomycin-phosphotransferase II (NPT II) gene. Control transfections with the widely used thymidine-kinase (TK) promoter gave comparable transfection rates as found for the nos-promoter with NPT II. The function of the nos-promoter was also confirmed by assaying neomycin-phosphotransferase synthesized in cells containing a plasmid with the NPT II-gene under control of this promoter. Several LTK+ clones stably transfected with Ti-plasmid propagated the total Ti-plasmid DNA in a colinear state presumably as an episomal unit. Dot blot analysis and polymerase chain reaction showed predominant transcription of Ti-sequences from the T-DNA area reflecting transcriptional activity of this region not only in plant cells but also in animal cells. These results provide new information about promoter functions in systems unrelated to their natural environment.
Mol Cell Biochem 1989 Oct 05
PMID:Promoter activity and expression of sequences from Ti-plasmid stably maintained in mammalian cells. 248 9

Ataxia-telangiectasia (A-T) is a multisystem hereditary disease featuring neurodegeneration, immunodeficiency, extreme cancer proneness, chromosomal instability, and radiosensitivity. A-T is found in many ethnic groups, and is genetically heterogeneous: four complementation groups have been identified in A-T so far. Attempts to isolate the A-T gene are based in part on gene transfer experiments, using permanent A-T fibroblast lines, obtained by transformation with SV40. "Immortalization" of A-T primary diploid fibroblasts using SV40 is difficult, possibly because of the chromosomal instability of these cells. The number of currently available permanent A-T fibroblast lines is small, and not all of them have been assigned to specific complementation groups. Using the assay of X-ray induced inhibition of DNA synthesis, we have assigned the A-T strain AT22IJE to complementation group AB. Origin-defective SV40 was used to transfect these cells, and one transformant (AT22IJE-T), which survived crisis, was found to have the typical characteristics of permanent cell lines obtained in this way. "In-gel renaturation" analysis did not show any DNA amplification of high degree in AT22IJE-T. Cytogenetic analysis showed considerable chromosomal instability in the new cell line, and medium conditioned by these cells contained the clastogenic activity which is characteristic of the parental strain as well. Other parameters of the "cellular A-T phenotype" have also been retained in the immortalized cells: hypersensitivity to the lethal effects of X-rays and neocarzinostatin, as well as "radioresistant" DNA synthesis. However, the sensitivity of AT22IJE-T to both DNA-damaging agents is less pronounced than that of the parental cells. The capacity of the cells for uptake of foreign DNA was tested by introducing into them the plasmid pRSVneo, using three different transfection methods. Satisfactory frequency of G418-resistant transfectants (0.66%) was achieved using a protocol recently published by Chen and Okayama (Mol. Cell Biol., 7: 2745-2752, 1987), which was found to be superior to the traditional calcium phosphate transfection method and to the polybrene-based method.
 ...
PMID:Cellular and molecular characteristics of an immortalized ataxia-telangiectasia (group AB) cell line. 253 4

Complement-mediated cytolysis of the mouse mammary tumor virus (MMTV)-infected rat hepatoma (HTC) cell line, M1.54, resulted in recovery of a mutant derivative, designated CR5, in which the magnitude of both basal and dexamethasone-induced proviral MMTV RNA expression was selectively reduced. Variant CR5 cells were transfected with a plasmid containing the glucocorticoid-regulated MMTV promoter linked to the neomycin resistance gene (pLNL). Half-maximal resistance to G418 killing was glucocorticoid inducible in both pLNL-transfected CR5 and M1.54 cells and was dependent on glucocorticoid receptor occupancy. The down-transcription of MMTV provirus sequences cannot be conferred to transfected genes driven by the same viral promoter suggesting that CR5 cells are defective in cis acting factors. Consistent with this notion, indirect immunofluorescence of transient heterokaryons revealed that uninfected wild-type HTC cells failed to complement the defect in CR5 while CR5 cells did not suppress the wild-type phenotype of M1.54 cells.
Mol Cell Endocrinol 1989 Jan
PMID:Glucocorticoid responsiveness of mouse mammary tumor virus (MMTV) promoters in a down-transcription hepatoma tissue culture (HTC) variant. 254 81

We have constructed an expression cartridge which has the bacterial hygromycin resistance gene (hph) fused to the Dictyostelium discoideum actin 15 promoter, with a segment of 3'-flanking DNA from the actin 15 locus placed downstream of the hph gene to serve as a transcription terminator. The plasmid pDE109, which contained this cartridge and a Dictyostelium origin of replication, transformed D. discoideum with high efficiency under hygromycin selection. The availability of this selectable marker circumvents the previous limitation of having G418 resistance as the only selectable marker for this organism; secondary transformation can now be used to introduce DNA into previously transformed cell lines.
Mol Cell Biol 1989 May
PMID:Hygromycin resistance as a selectable marker in Dictyostelium discoideum. 254 56

A system is presented for transformation of the fission yeast Schizosaccharomyces pombe to resistance against the antibiotic G418. The bacterial resistance gene of the transposon Tn5 is expressed under the control of promoters and transcription terminators from cauliflower mosaic virus (CaMV). The promoter of the S. pombe alcohol dehydrogenase gene has also been used. Transformants can be selected directly on medium containing G418 (up to 1 mg/ml) due to inactivation of G418 by the Tn5 gene product, the aminoglycoside 3'-phosphotransferase (II). The plant viral promoter 35S confers higher resistance to G418 than the 19S promoter. This corresponds to the relative strengths of these promoters in plant cells. The strong plant promoter 35S yields resistance comparable to that obtained with the strong S. pombe promoter from the alcohol dehydrogenase gene. The constructions with the two plant promoters have been used on multicopy shuttle plasmids that replicate autonomously in S. pombe and Escherichia coli. In addition the 35S and the 19S constructions have been inserted into the S. pombe genome where they confer G418 resistance as single copy genes. Since vector sequences are excluded in this case, all the necessary signals for expression of G418 resistance are contained within the DNA fragments containing the plant promoters, the resistance gene and the plant terminators. This transformation system is independent of S. pombe mutants. It may be useful for the transformation of other lower eukaryotes. The activity of the CaMV promoters in S. pombe may be exploited for the expression of plant genes in fission yeast.
Mol Gen Genet 1989 Dec
PMID:Cauliflower mosaic virus promoters direct efficient expression of a bacterial G418 resistance gene in Schizosaccharomyces pombe. 255 89

A cDNA clone of a rat cytochrome P450b gene was used to construct an expression vector driven by an SV40 promoter and containing a G418-resistance selectable marker. This bifunctional plasmid (pJRSL100) was transfected into the C3H 10T1/2CL8 mouse embryo fibroblast cell line. G418-resistant clones were selected and tested for enhanced sensitivity to the carcinogen 2-acetylaminofluorene (2-AAF), a compound that does not normally induce cytotoxicity or morphological transformation in these cells. One subclone, 19P450b-4, exhibited an increased cytotoxic response to 2-AAF compared to the parental C3H10T1/2CL8 cells. DNA analyses of this subclone showed increased number of copies of the cytochrome P450b and the appearance of unique restriction fragment bands relative to parental and control transfected cells. This subclone also exhibited increased levels of mRNA complementary to the P450b cDNA. Metabolism studies of 2-AAF in this subclone demonstrated an increase in the C-hydroxylated metabolites 1-, 3-, 5/9-, and 7-hydroxy-AAF compared with parental C3H 10T1/2CL8 cells. The results indicate that C3H 10T1/2CL8 cells can be transfected with gene/cDNAs to increase their metabolic competency and that such transfection may enhance the usefulness of the C3H 10T1/2CL8 cells in studies on chemically induced cytotoxicity and morphological transformation.
Mol Carcinog 1989
PMID:Transfection of a rat cytochrome P450b cDNA into C3H10T1/2CL8 mouse embryo fibroblasts. 260 64


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