Abstract:
:Heteroduplexes were prepared from two plasmids, pRH4-14/TK and pRH5-8/TK, containing different amber mutations in the neomycin resistance gene (Neor). The Neor gene was engineered to be expressed in both bacterial and mammalian cells. A functional Neor gene conferred kanamycin resistance to bacteria and resistance to the drug G418 to mammalian cells. In addition, the plasmids contained restriction site polymorphisms which did not confer a selectable phenotype but were used to follow the pattern of correction of mismatched bases in the heteroduplexes. In a direct comparison of the efficiency of transforming mouse LMtk- cells to G418r, the injection of heteroduplexes of pRH4-14/TK-pRH5-8/TK was 10-fold more efficient than the coinjection of pRH4-14/TK and pRH5-8/TK linear plasmid DNA. In fact, injection of 5 to 10 molecules of heteroduplex DNA per cell was as efficient in transforming LMtk- cells to G418r as the injection of 5 to 10 molecules of linear plasmid DNA per cell containing a wild-type Neor gene. To determine the pattern of mismatch repair of the injected heteroduplexes, plasmids were "rescued" from the G418r cell lines. From this analysis we conclude that the generation of wild-type Neor genes from heteroduplex DNA proceeds directly by correction of the mismatched bases, rather than by alternative mechanisms such as recombination between the injected heteroduplexes. Our finding that a cell can efficiently correct mismatched bases when confronted with preformed heteroduplexes suggests that this experimental protocol could be used to study a wide range of DNA repair mechanisms in cultured mammalian cells.
journal_name
Mol Cell Bioljournal_title
Molecular and cellular biologyauthors
Folger KR,Thomas K,Capecchi MRdoi
10.1128/mcb.5.1.70subject
Has Abstractpub_date
1985-01-01 00:00:00pages
70-4issue
1eissn
0270-7306issn
1098-5549journal_volume
5pub_type
杂志文章abstract::We demonstrate that ATF3, a member of the ATF/CREB family of transcription factors, is induced in a variety of stressed tissues: mechanically injured liver, toxin-injured liver, blood-deprived heart, and postseizure brain. We also demonstrate that an ATF3-interacting protein, gadd153/Chop10, forms a nonfunctional hete...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/mcb.16.3.1157
更新日期:1996-03-01 00:00:00
abstract::We have previously identified an 11-base DNA sequence, 5'-G-G-T-A-C-C-T-A-A-C-C-3' (simian virus 40 [SV40] map position 294 to 304), which is important in the control of SV40 late RNA expression in vitro and in vivo (Brady et al., Cell 31:625-633, 1982). We report here the identification of another domain of the SV40 ...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/mcb.4.1.133
更新日期:1984-01-01 00:00:00
abstract::The mouse Surfeit locus, which contains a cluster of at least four genes (Surf-1 to Surf-4), is unusual in that adjacent genes are separated by no more than 73 base pairs (bp). The heterogeneous 5' ends of Surf-1 and Surf-2 are separated by only 15 to 73 bp, the 3' ends of Surf-1 and Surf-3 are only 70 bp apart, and t...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/mcb.9.1.224
更新日期:1989-01-01 00:00:00
abstract::Retinoblastoma (RB) tumors develop when both alleles of a gene (RB1) are mutated and unable to function normally. Recently, Friend et al. [S. H. Friend, R. Bernards, S. Rogelj, R. A. Weinberg, J. M. Rapaport, D. M. Albert, and T. P. Dryja, Nature (London) 32:643-646, 1986] reported the cloning of a gene, 4.7R, with so...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/mcb.8.5.2082
更新日期:1988-05-01 00:00:00
abstract::We report results showing that several gamma gene promoter elements participate in the developmental control of gamma-globin genes. Four gamma gene constructs with 5' truncated at -141, -201, -382, and -730 of the A gamma gene promoter linked to a micro locus control region (microLCR) cassette were used for production...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/mcb.13.12.7636
更新日期:1993-12-01 00:00:00
abstract::After flagellar detachment in Chlamydomonas reinhardi, there is a rapid synthesis and accumulation of mRNAs for tubulin and other flagellar proteins. Maximum levels of these mRNAs (flagellar RNAs) are reached within 1 h after deflagellation, after which they are rapidly degraded to their predeflagellation levels. The ...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/mcb.6.1.54
更新日期:1986-01-01 00:00:00
abstract::Smad proteins play a key role in the intracellular signaling of the transforming growth factor beta (TGF-beta) superfamily of extracellular polypeptides that initiate signaling from the cell surface through serine/threonine kinase receptors. A subclass of Smad proteins, including Smad6 and Smad7, has been shown to fun...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/mcb.20.9.3157-3167.2000
更新日期:2000-05-01 00:00:00
abstract::To identify and characterize the proteins encoded by the erbA proto-oncogene, we expressed the C-terminal region of v-erbA in a bacterial trpE expression vector system and used the fusion protein to prepare antiserum. The anti-trp-erbA serum recognized the P75gag-erbA protein encoded by avian erythroblastosis virus an...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/mcb.8.10.4155
更新日期:1988-10-01 00:00:00
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journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/MCB.26.4.1259-1271.2006
更新日期:2006-02-01 00:00:00
abstract::The expression of c-ets2 is rapidly induced in a variety of myelomonocytic cell lines as they differentiate into macrophages. We find that constitutive expression of c-ets2 in the M1D+ myeloblast leukemic cell line (M1ets2) is sufficient to push these cells to a more differentiated state. The expression of several dif...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/mcb.16.12.6851
更新日期:1996-12-01 00:00:00
abstract::The extensive array of basic helix-loop-helix (bHLH) transcription factors and their combinations as dimers underpin the diversity of molecular function required for cell type specification during embryogenesis. The bHLH factor TWIST1 plays pleiotropic roles during development. However, which combinations of TWIST1 di...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/MCB.00663-19
更新日期:2020-05-14 00:00:00
abstract::Glucose-6-phosphate dehydrogenase (G6PD) produces cellular NADPH, which is required for the biosynthesis of fatty acids and cholesterol. Although G6PD is required for lipogenesis, it is poorly understood whether G6PD in adipocytes is involved in energy homeostasis, such as lipid and glucose metabolism. We report here ...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/MCB.25.12.5146-5157.2005
更新日期:2005-06-01 00:00:00
abstract::In Saccharomyces cerevisiae, the intracellular concentration of trehalose increases rapidly in response to many environmental stresses, including heat shock. These high trehalose levels have been correlated with tolerance to adverse conditions and led to the model that trehalose functions as a chemical cochaperone. He...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/MCB.01158-06
更新日期:2007-02-01 00:00:00
abstract::The cellular response to DNA damage is an intricate mechanism that involves the interplay among several pathways. In this study, we provide evidence of the roles of the polyadenylation factor cleavage stimulation factor 50 (CstF-50) and the ubiquitin (Ub) escort factor p97 as cofactors of BRCA1/BARD1 E3 Ub ligase, fac...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/MCB.00364-17
更新日期:2018-01-29 00:00:00
abstract::The interleukin enhancer binding factors ILF2 (NF45) and ILF3 (NF90/NF110) have been implicated in various cellular pathways, such as transcription, microRNA (miRNA) processing, DNA repair, and translation, in mammalian cells. Using tandem affinity purification, we identified human NF45 and NF90 as components of precu...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/MCB.00306-15
更新日期:2015-10-01 00:00:00
abstract::The baker's yeast Saccharomyces cerevisiae possesses a single gene encoding heat shock transcription factor (HSF), which is required for the activation of genes that participate in stress protection as well as normal growth and viability. Yeast HSF (yHSF) contains two distinct transcriptional activation regions locate...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/mcb.18.11.6340
更新日期:1998-11-01 00:00:00
abstract::During Xenopus development, the synthesis of ribosomal proteins is regulated at the translational level. To identify the region of the ribosomal protein mRNAs responsible for their typical translational behavior, we constructed a fused gene in which the upstream sequences (promoter) and the 5' untranslated sequence (f...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/mcb.10.2.816
更新日期:1990-02-01 00:00:00
abstract::Telomerase promotes telomere maintenance by copying a template within its integral RNA subunit to elongate chromosome ends with new telomeric repeats. Motifs have been defined within the telomerase RNA that contribute to mature RNA accumulation, holoenzyme catalytic activity, or enzyme recruitment to telomeres. Here, ...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/MCB.01033-10
更新日期:2011-02-01 00:00:00
abstract::The biosynthesis, phosphorylation, and degradation of the epidermal growth factor (EGF) receptor were examined in normal human fibroblasts. The receptor was initially synthesized as an Mr = 160,000 immature form which matured to an Mr = 170,000 form in a monensin-sensitive manner. Tunicamycin treatment led to the accu...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/mcb.4.9.1718
更新日期:1984-09-01 00:00:00
abstract::Oncostatin M is a polypeptide cytokine, produced by normal and malignant hematopoietic cells, that has several in vitro activities, including the ability to inhibit growth of cultured carcinoma cells. Here we present a structural and functional comparison of two oncostatin M-related proteins (Mr 36,000 and 32,000) sec...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/mcb.10.5.1882
更新日期:1990-05-01 00:00:00
abstract::In an effort to identify widely active positive regulatory elements, we have examined the action of the cytomegalovirus enhancer-promoter in transgenic mice. These elements activated expression in 24 of 28 tissues tested. The greatest expression was observed in the heart, kidney, brain, and testis. Maximum expression ...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/mcb.10.8.4406
更新日期:1990-08-01 00:00:00
abstract::Enhancer elements modulate promoter activity over vast chromosomal distances, and mechanisms that ensure restrictive interactions between promoters and enhancers are critical for proper control of gene expression. The human beta-globin locus control region (LCR) activates expression of five genes in erythroid cells, i...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/MCB.00602-07
更新日期:2007-08-01 00:00:00
abstract::GCN4 encodes a transcriptional activator of amino acid biosynthetic genes in Saccharomyces cerevisiae. The GCN3 product is a positive regulator required for increased synthesis of GCN4 protein in amino acid-starved cells. GCN3 appears to act indirectly by antagonizing GCD-encoded negative regulators of GCN4 expression...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/mcb.8.11.4808
更新日期:1988-11-01 00:00:00
abstract::Sphingomyelin is a major lipid in the bilayer of subcellular membranes of eukaryotic cells. Different sphingomyelinases catalyze the initial step in the catabolism of sphingomyelin, the hydrolysis to phosphocholine and ceramide. Sphingomyelinases have been postulated to generate ceramide as a lipophilic second messeng...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/mcb.22.11.3633-3638.2002
更新日期:2002-06-01 00:00:00
abstract::We have designed and constructed integration-defective retroviral vectors to explore their potential for gene targeting in mammalian cells. Two nonoverlapping deletion mutants of the bacterial neomycin resistance (neo) gene were used to detect homologous recombination events between viral and chromosomal sequences. St...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/mcb.9.4.1621
更新日期:1989-04-01 00:00:00
abstract::By using a retrovirus-derived vector system, we generated derivatives of the human colon cancer cell line HT29 that stably overexpress a full-length cDNA encoding the beta 1 isoform of rat protein kinase C (PKC). Two of these cell lines, PKC6 and PKC7, displayed an 11- to 15-fold increase in PKC activity when compared...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/mcb.10.9.4650
更新日期:1990-09-01 00:00:00
abstract::The Saccharomyces cerevisiae Trf4 and Trf5 proteins are members of a distinct family of eukaryotic DNA polymerase beta-like nucleotidyltransferases, and a template-dependent DNA polymerase activity has been reported for Trf4. To define the nucleotidyltransferase activities associated with Trf4 and Tr5, we purified the...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/MCB.25.22.10183-10189.2005
更新日期:2005-11-01 00:00:00
abstract::Cooperation between STAT3 and c-Jun in driving transcription during transfection of reporter constructs is well established, and both proteins are present on some interleukin-6 (IL-6) STAT3-dependent promoters on chromosomal loci. We report that small interfering RNA knockdown of c-Jun or c-Fos diminishes IL-6 inducti...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/MCB.00613-07
更新日期:2007-09-01 00:00:00
abstract::Heat shock proteins (HSPs) are highly conserved among all organisms from prokaryotes to eukaryotes. In mice, the HSP genes Hsp70.1 and Hsp70.3 are induced by both endogenous and exogenous stressors, such as heat and toxicants. In order to determine whether such proteins specifically influence genomic instability, mice...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/mcb.24.2.899-911.2004
更新日期:2004-01-01 00:00:00
abstract::The transcriptional coactivators p300 and CREB binding protein (CBP) are important regulators of the cell cycle, differentiation, and tumorigenesis. Both p300 and CBP are targeted by viral oncoproteins, are mutated in certain forms of cancer, are phosphorylated in a cell cycle-dependent manner, interact with transcrip...
journal_title:Molecular and cellular biology
pub_type: 杂志文章
doi:10.1128/mcb.20.8.2676-2686.2000
更新日期:2000-04-01 00:00:00