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Query: UNIPROT:P06889 (Mol)
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It is reported here that the rpr DNA repair gene of Serratia marcescens does not complement an Escherichia coli xth nfo AP endonuclease mutation for resistance to methyl methanesulphonate (MMS). Rather, rpr sensitized Escherichia coli wild-type, xth, and nfo strains to MMS. Also, it was found that rpr could not complement a triple tag alkA recA mutation in E. coli, indicating that there are limits to rpr complementing capabilities. It was determined that rpr gene dosage was not a factor in recA complementation. MMS sensitization of an E. coli wild-type strain, however, was directly related to rpr copy number. These data indicate that Rpr does not have an associated AP endonuclease activity, and that it is incapable of substituting for Tag I, Tag II, and RecA in a tag alkA recA background.
Mol Microbiol 1990 Apr
PMID:Serratia marcescens rpr gene sensitizes Escherichia coli wild-type, xth, and nfo strains to methyl methanesulphonate. 169 47

We report here the molecular isolation of a DNA fragment which encodes Tag-like activity from the Gram-negative bacterium Serratia marcescens. A recombinant plasmid encoding Tag-like activity was isolated from a S. marcescens plasmid gene library by complementation of an Escherichia coli tag mutant, which is deficient in 3-methyladenine DNA glycosylase I. The clone complements E. coli tag, recA, alkA, but not alkB, mutants for resistance to the DNA-damaging agent methyl methanesulphonate (MMS). The coding region of the Tag activity, initially isolated on a 6.5kb BamHI fragment, was defined to a 1.8kb BglII-SmaI fragment. Labelling of plasmid-encoded proteins using maxicells revealed that the 1.8kb fragment encodes two proteins of molecular weights 42,000 and 16,000. Data presented here suggest that the cloned fragment encodes a DNA repair protein(s) that has similar activity to the 3-methyladenine DNA glycosylase I of E. coli.
Mol Microbiol 1989 Feb
PMID:Molecular cloning and characterization of a genetic region from Serratia marcescens involved in DNA repair. 266 89

A transgenic mouse line (EGF/Tag) has been established in which expression of SV40 T-antigen is directed by a 5.5 kb fragment of the 5'-flanking region of the mouse epidermal growth factor (EGF) gene. Of the two principal sites of EGF expression in mice, submaxillary gland and kidney, T-antigen mRNA and protein were detected in the former but not in the latter tissue of the EGF/Tag animals. T-antigen expression in the submaxillary gland was restricted to the EGF-producing cells of the granular convoluted tubules, and the oncoprotein induced hyperplasia of these cells. T-antigen levels were markedly higher in the submaxillary glands of male compared with female transgenic mice, suggesting that expression of the transgene was androgen-regulated, like the endogenous EGF gene. These results indicate that the 5.5 kb fragment upstream of the mouse EGF gene contains the DNA enhancer elements required for hormonally regulated expression in the submaxillary gland. Since the hyperplastic submaxillary glands of the EGF/Tag mice continue to synthesize EGF, these glands provide a tissue source from which it may prove possible to establish EGF-secreting cell lines for further in vitro studies of the mechanisms regulating expression of the EGF gene.
J Mol Endocrinol 1994 Jun
PMID:Directed expression of simian virus 40 T-antigen in transgenic mice using the epidermal growth factor gene promoter. 791 70

Ran/TC4, a member of the RAS gene superfamily, encodes an abundant nuclear protein that binds and hydrolyzes GTP. Transient expression of a Ran/TC4 mutant protein deficient in GTP hydrolysis blocked DNA replication, suggesting a role for Ran/TC4 in the regulation of cell cycle progression. To test this possibility, we exploited an efficient transfection system, involving the introduction of cDNAs in the pMT2 vector into 293/Tag cells, to analyze phenotypes associated with mutant and wild-type Ran/TC4 expression. Expression of a Ran/TC4 mutant protein deficient in GTP hydrolysis inhibited proliferation of transfected cells by arresting them predominantly in the G2, but also in the G1, phase of the cell cycle. Deletion of an acidic carboxy-terminal hexapeptide from the Ran/TC4 mutant did not alter its nuclear localization but did block its inhibitory effect on cell cycle progression. These data suggest that normal progression of the cell cycle is coupled to the operation of a Ran/TC4 GTPase cycle. Mediators of this coupling are likely to include the nuclear regulator of chromosome condensation 1 protein and the mitosis-promoting factor complex.
Mol Cell Biol 1994 Jun
PMID:Effects of mutant Ran/TC4 proteins on cell cycle progression. 819 59

A critical stage in pollen development is the dissolution of the four products of meiosis, the tetrads, into free microspores. The tetrads are surrounded by a thick callose wall composed of beta-1,3-glucan. At the completion of meiosis, the tetrads are released into the anther locule after hydrolysis of the callose by a beta-1,3-glucanase. Using the polymerase chain reaction, we have amplified and subsequently cloned a cDNA corresponding to a beta-1,3-glucanase, tobacco (Nicotiana tabacum cv. Samsun) anther glucanase (Tag 1), which is expressed exclusively in anthers from meiosis to the free microspore stage of pollen development. The identity of the clone was determined by DNA and deduced protein sequence similarity to other known beta-1,3-glucanases. Several regions strictly conserved among four classes of glucanases are also conserved in the Tag 1 protein. Tag 1 represents a novel class of beta-1,3-glucanase based on phylogenetic analysis and RNA expression pattern. Tag 1 RNA was detected in situ only in the tapetum, with maximal expression just prior to tetrad dissolution. Due to its expression pattern and sequence similarity to other beta-1,3-glucanases, we believe Tag 1 may be involved in tetrad dissolution.
Plant Mol Biol 1994 Mar
PMID:Cloning and characterization of Tag 1, a tobacco anther beta-1,3-glucanase expressed during tetrad dissolution. 820 27

We describe a novel technique for isolation of sequences that are present in one genome (tracer), but absent in another (driver). Tracer DNA, cleaved with Sau 3A and capped with a single stranded PCR adapter, is allowed to hybridize with an excess of sheared biotinylated driver; biotinylated DNA and its hybrids with the tracer are removed by phenol/chloroform extraction after incubation with streptavidin. After several rounds of subtraction the ends of self-annealed tracer molecules from the nonextractable fraction are filled-in with Tag polymerase and amplified, using the single stranded PCR adapter as a primer. The method has been applied to purification of fragments from a 2.9 kb plasmid added to E. coli DNA at equimolar quantity. Plasmid derived fragments (250-1000 bp), initially comprising 1/1400th part of tracer DNA, were purified to homogeneity after two rounds of subtraction followed by PCR.
Mol Gen Mikrobiol Virusol
PMID:A method for isolation of sequences missing in one of two related genomes. 835 Aug 79

Testicular tumorigenesis was observed in transgenic mice expressing the 6-kb mouse inhibin alpha-subunit promoter/Simian virus 40 T-antigen (SV40 Tag) fusion gene. The tumors were confined to Leydig cells using immunohistochemistry with anti-Tag antibody, specific binding of biotinylated hCG and histochemistry for 3 beta-hydroxysteroid dehydrogenase. Leydig cell hyperplasia and presence of Tag protein in the testicular interstitial tissue were already evident at 5 and 6.5 days of age, respectively. An immortalized cell line, BLT-1, was established from one testicular tumor. These cells expressed the LH receptor and P450scc mRNAs, and displayed LH-responsive cAMP and progesterone production, and low testosterone production. The cells also specifically bound 125I-labeled recombinant human LH with high affinity (36000 binding sites/cell), and the binding was regulated by 8Br-cAMP and hCG. This gonadal tumor model is valuable for further studies on endocrine functions of Leydig cells and their tumorigenesis in vivo and in vitro.
Mol Cell Endocrinol 1996 May 31
PMID:The mouse inhibin alpha-subunit promoter directs SV40 T-antigen to Leydig cells in transgenic mice. 880 33

In this study we have investigated the subcellular localization in transfected COS-1 cells of the two major forms of the hepatitis C virus core protein: the immature protein of 191 residues (p21) and its proteolytically cleaved product of 173 residues (p19). In this study, and unlike previous investigations, we have been able to distinguish separately the localization of p21 from p19. This was achieved by the addition of a C-terminal HSV Tag to the p21 full coding sequence, and exploiting the fact that it is subsequently lost in the p19 product. In order to obtain an accurate localization of both p21 and p19 we used a mouse anti-HSV Tag MAb together with a human anti-core MAb (B12.F8) to perform double immunofluorescence studies. The results have shown that p21 is always localized around the nuclear envelope. On the other hand, p19 can be found diffused in the cytoplasm to different degrees. These in vivo results reinforce the proposed links between the regulated processing of the hepatitis C virus core protein and the possibility that this may contribute towards the regulation of its diverse biological functions.
Cell Mol Biol (Noisy-le-grand) 1998 May
PMID:Localization of the different hepatitis C virus core gene products expressed in COS-1 cells. 962 Apr 47

The availability of large EST (Expressed Sequence Tag) databases has led to a revolution in the way new genes are cloned. Difficulties arise, however, due to high error rates and redundancy of raw EST data. For these reasons, one of the first tasks performed by a scientist investigating any EST of interest is to gather contiguous ESTs and assemble them into a larger virtual cDNA. The REX (Recursive EST eXtender) algorithm described in this paper completely automates this process by finding ESTs that can be clustered on the basis of overlapping bases, and then assembling the contigs into a consensus sequence. By combining the clustering and assembly steps, REX can quickly generate assemblies from EST databases that are frequently updated without having to preprocess the data. A consensus assembly method is used to correct miscalled bases and remove indel errors. A unique feature of this method is that it addresses the issues of splice variants and unspliced cDNA data. Since REX is a fast greedy algorithm, it can address the problem of generating a database of assembled sequences from very large collections of EST data. A procedure is described for creating and maintaining an Assembled Consensus EST database (ACE) that is useful for characterizing the large body of data that exists in EST databases.
Proc Int Conf Intell Syst Mol Biol 1998
PMID:Automated clustering and assembly of large EST collections. 978 26

The Schistosoma mansoni gene sequence encoding the breast basic conserved protein 1/ribosomal protein L13 has been isolated from an adult worm cDNA library using the Expressed Sequence Tag strategy. The cDNA codes for a putative protein of 184 amino acids which is approximately 55% identical to other eukaryotic L13 ribosomal proteins. A PCR amplified genomic fragment containing the coding region of the gene was seen to possess only a single large intron interrupting the open reading frame. Studies of gene expression by RT-PCR showed the transcript is expressed in distinct stages of the parasite life cycle. The cDNA was also hybridized with an ordered cosmid library of S. mansoni and the identified cosmids were mapped to chromosomes 3 and W by chromosomal in situ suppression hybridization.
Comp Biochem Physiol B Biochem Mol Biol 1998 Aug
PMID:Characterization of a Schistosoma mansoni homologue of the gene encoding the breast basic conserved protein 1/L13 ribosomal protein. 985 18


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