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At least 1,387 records · Page 77Linked to original sources
PubMed1996

Efficiency of MucAB and Escherichia coli UmuDC proteins in quinolone and UV mutagenesis in Salmonella typhimurium: effect of MucA and UmuD processing.

The role of MucAB and Escherichia coli UmuDC proteins in mutagenesis by 4-quinolone (4-Q) compared to that in UV mutagenesis has been studied in hisG428 Salmonella typhimurium strains. A low-copy plasmid carrying mucAB genes, but not umuDC, promotes reversion of the hisG428 mutation by the 4-Q ciprofloxacin. In contrast, a umuDC plasmid mediates the reversion of hisG428 by UV, although less efficiently than a mucAB one. In addition, a unique copy of mucAB genes is enough to promote UV mutagenesis, whereas, several copies of them are required to detect ciprofloxacin mutagenesis. Therefore, the mutagenic repair of quinolone damage by MucAB proteins is not a very efficient process. The presence of an umuD'C plasmid but not a mucA'B one, slightly increases the reversion of the hisG428 mutation by ciprofloxacin and this finding is further discussed. In contrast, MucA'B are still more active than UmuD'C proteins in UV mutagenesis. These results suggest that the enhanced processing of MucA compared to UmuD would not explain all functional differences between MucAB and UmuDC proteins in the error-prone DNA repair.

Anti-Infective Agents↗
PubMed2002

Association of tapasin and COPI provides a mechanism for the retrograde transport of major histocompatibility complex (MHC) class I molecules from the Golgi complex to the endoplasmic reticulum.

Tapasin is a subunit of the transporter associated with antigen processing (TAP). It associates with the major histocompatibility complex (MHC) class I. We show that tapasin interacts with beta- and gamma-subunits of COPI coatomer. COPI retrieves membrane proteins from the Golgi network back to the endoplasmic reticulum (ER). The COPI subunit-associated tapasin also interacts with MHC class I molecules suggesting that tapasin acts as the cargo receptor for packing MHC class I molecules as cargo proteins into COPI-coated vesicles. In tapasin mutant cells, neither TAP nor MHC class I are detected in association with the COPI coatomer. Interestingly, tapasin-associated MHC class I molecules are antigenic peptide-receptive and detected in both the ER and the Golgi. Our data suggest that tapasin is required for the COPI vesicle-mediated retrograde transport of immature MHC class I molecules from the Golgi network to the ER.

Antiporters↗
PubMed2004

Mitochondrial transcription factor A regulates mtDNA copy number in mammals.

Mitochondrial DNA (mtDNA) copy number regulation is altered in several human mtDNA-mutation diseases and it is also important in a variety of normal physiological processes. Mitochondrial transcription factor A (TFAM) is essential for human mtDNA transcription and we demonstrate here that it is also a key regulator of mtDNA copy number. We initially performed in vitro transcription studies and determined that the human TFAM protein is a poor activator of mouse mtDNA transcription, despite its high capacity for unspecific DNA binding. Next, we generated P1 artificial chromosome (PAC) transgenic mice ubiquitously expressing human TFAM. The introduced human TFAM gene was regulated in a similar fashion as the endogenous mouse Tfam gene and expression of the human TFAM protein in the mouse did not result in down-regulation of the endogenous expression. The PAC-TFAM mice thus had a net overexpression of TFAM protein and this resulted in a general increase of mtDNA copy number. We used a combination of mice with TFAM overexpression and TFAM knockout and demonstrated that mtDNA copy number is directly proportional to the total TFAM protein levels also in mouse embryos. Interestingly, the expression of human TFAM in the mouse results in up-regulation of mtDNA copy number without increasing respiratory chain capacity or mitochondrial mass. It is thus possible to experimentally dissociate mtDNA copy number regulation from mtDNA expression and mitochondrial biogenesis in mammals in vivo. In conclusion, our results provide genetic evidence for a novel role for TFAM in direct regulation of mtDNA copy number in mammals.

Animals↗
PubMed2000

Are linker histones (histone H1) dispensable for survival?

In the multicelled filamentous ascomycete Ascolobus immersus, the single copy gene for histone H1 can be silenced by methylation in the process known as methylation-induced premeiotically (MIP). The results of a recent paper using this unique system(1) have shown that histone H1 silencing results in an enhanced DNA accessibility to nucleases and an increase in the overall extent of DNA methylation. Interestingly, while none of these effects appear to decrease the immediate viability of this fungus, silencing of histone H1 results in a significant decrease in its overall life span. These results suggest that while linker histones may be dispensable for the relatively short life span of an individual cell, they are most likely indispensable for survival of higher eukaryote organisms.

Animals↗
PubMed1994

Extremely stable transcripts may compensate for the elimination of the gene fert-1 from all Ascaris lumbricoides somatic cells.

The single-copy gene fert-1 becomes eliminated from all somatic cells during the process of chromatin diminution in Ascaris lumbricoides var. suum. By using Northern blot and in situ hybridization techniques, we have analyzed its rather unusual expression pattern. Different splicing and 3' end formation events generate in a developmentally regulated manner various poly(A)+ and poly(A)- fert-1 RNA species. The lack of any significant open reading frame in most of its RNA products indicates that fert-1 may function as structural RNA rather than encoding a protein. Fert-1 transcripts are produced in the precursors of the gametes, but degraded at the time of meiosis and not passed on to the zygote. Embryonic transcription of fert-1 sets in as soon as the female nucleus has completed its meiosis. Our data thus demonstrate that the Ascaris transcription apparatus is active prior to the general onset of zygotic transcription, which we think takes place in the four- to six-cell-stage embryos. Upon elimination of fert-1 gene from the somatic cells, most of its transcripts disappear. Two short fert-1 RNA products, however, are stably maintained throughout development until the second larval stage, which is more than 1 month after the elimination of their coding sequences. Possible functions of fert-1 are discussed.

Amino Acid Sequence↗
PubMed2002

Ancient duplications of the human proglucagon gene.

The human proglucagon gene (GCG) is encoded within a finished 576-kb DNA sequence generated by the Human Genome Project. GCG is flanked by 18 kb and 65 kb of DNA, 5' and 3', respectively, that do not encode genes. The genomic sequence that includes GCG was found to have a long history of gene duplication events. Some members of the glucagon-like family of genes, GCG on chromosome 2 and GIP on chromosome 17, may be products of ancient genome duplications on the early vertebrate lineage. A large genomic tandem duplication event that included DPP4-like and GCG genes occurred before the amphibian-mammal divergence, but one of the duplicated copies of GCG has been lost on the human lineage. Recently, a processed pseudogene of the X-chromosome-linked gene TIMM8A was inserted downstream of GCG. Some ancient duplicates of GCG may retain physiological functions in other vertebrates.

Animals↗
PubMed1990

Relationship of vector insert size to homologous integration during transformation of Neurospora crassa with the cloned am (GDH) gene.

We used lambda and plasmid vectors containing the am+ gene in an insert of from 2.7 to 9.1 kb, to transform am point mutant and deletion strains. A total of 199 transformants were examined with the potential to yield am+ transformants by homologous recombination. When we used vectors that had 9.1 kb of homology with the chromosomal DNA, 30% of the transformants obtained were the result of homologous recombination regardless of whether the vector was a lambda molecule, a circular plasmid, or a plasmid that had been linearized prior to transformation. When vectors with up to 5.1 kb of homology were used, very few transformants (1 of 89 tested) resulted from homologous recombination. Of a sample of 29 ectopic integration events obtained by transformation with the 9.1 kb fragment cloned in a lambda vector, 18 included a major part (usually almost all) of both arms of lambda with the entire Neurospora 9.1 kb insert between them. Four included only lambda long arm sequence together with an adjacent segment of the insert containing the am gene. The remaining seven were the result of multiple integrations. There was no evidence of circularization of the lambda vector prior to integration. All transformants that had multiple copies of the am gene appeared to be subject to the RIP process, which causes multiple mutations in duplicated sequences during the sexual cycle.

Bacteriophage lambda↗
PubMed1988

Duplications and deletions of Vh genes in inbred strains of mice.

The evolution of variable region (Vh) gene family copy number and polymorphism was investigated by the analysis of the immunoglobulin heavy chain variable region (Igh-V) locus in 74 inbred strains and substrains of mice. Several strains were found to have slight differences from Igh-V haplotypes previously identified, usually involving the gain or loss of one or a few members of a single Vh gene family. These results indicate that the evolution of copy number in the mouse Igh-V locus proceeds largely by the accumulation of incremental changes, reflecting the clustered organization of the mouse Igh-V locus. We have found no evidence of very large or frequent duplication or deletion events indicative of rapid expansion or contraction processes. The existence of one or more particularly large Vh gene families most likely reflects random copy number variation, rather than selection for the amplification of their members. The identification of strains with recombinant Vh gene arrays demonstrates that recombination, both within and between haplotypes, appears to be the predominant mechanism generating the high restriction fragment length polymorphism in the Igh-V locus.

Animals↗
PubMed1979

Characterization of 10S RNA: a new stable rna molecule from Escherichia coli.

When cells of Escherichia coli are labeled with 32Pi for long periods of time and the cell content is subjected to electrophoresis in polyacrylamide gels, an RNA band appears which is about 10S in size. This band seems to contain three conformers. After treatment with formamide only a single band appears in this region of the gel, which contains 550 nucleotides as determined from its mobility. The complexity of the fingerprint of this material, after digestion with T1-RNase, is in agreement with the size as determined by the mobility, this confirming that indeed it is a single molecule. Composition of the T1-oligonucleotides was determined by digesting the T1-generated oligonucleotides with pancreatic RNase and T2-RNase. The quantitative and qualitative analysis of these digestions suggest that 10S RNA contains 609 nucleotides. The molecule contains, besides the four regular bases, one copy per molecule of the modified base pseudouridine. 10S RNA cannot be processed by cell extracts to tRNA-sized molecules and does not bind significantly to ribosomes, hence it is unlikely to be a tRNA precursor or an mRNA.

Base Sequence↗
PubMed1991

Structure of rabbit butyrylcholinesterase gene deduced from genomic clones and from cDNA with introns.

1. Three clones were isolated from a rabbit genomic library. They covered the entire coding sequence of the rabbit BChE gene. The positions of splice sites between exons 2, 3, and 4 are identical to those found in the human gene (Arpagaus et al., 1990). Exon 2 covers 83% of the coding sequence. This contrasts with the small size of exon 3 (167 bp) and large size of introns 2 and 3 (greater than 20 kb each). The active-site serine at position 198 is found in a highly conserved region. Aspartic acids in positions 91 and 170 are conserved in human and rabbit, and one of them could be involved in the calytic triad. Aspartic acid 70, present in the anionic site of human BChE, is also conserved in rabbit BChE. The coding sequences of human and rabbit BChE are 89% identical over 744 bp around the active-site serine. 2. In addition to the genomic clones, one cDNA clone (BNY1) was isolated. This cDNA was unusual in that it contained intronic sequences. The insert of 1 kb contained 167 coding bases homologous to the nucleotide sequence 1434 to 1600 of human cDNA and corresponded to exon 3 of the BChE gene. On each side of this coding region, consensus sequences of intron-exon boundaries were found. 3. The presence of large-size transcripts in Northern blots and the existence of a cDNA copy of unprocessed mRNA found in the BNY1 clone suggest a slow processing of transcripts. A genomic sequence unspliced in a cDNA of Torpedo AChE could give a transmembrane domain (Sikorav et al., 1988); the corresponding sequence in rabbit BChE gene, also found in a cDNA, had no homology with Torpedo AChE but could be translated in a hydrophobic C-terminal domain if maintained in mature mRNA.

Amino Acid Sequence↗
PubMed2006

Processed pseudogenes, processed genes, and spontaneous mutations in the Arabidopsis genome.

We identified 411 processed sequences in the Arabidopsis thaliana genome based on the fact that they have lost their intron(s) and have a length that is at least 95% of the length of the gene that gave rise to them. These sequences were generated by 230 different genes and clearly originated from retrotranspositons events because most of them (91%) have a poly(A)-tail. They are composed of 376 sequences with frame shifts and/or premature stop codons (processed pseudogenes) and 35 sequences without disablements (processed genes). Eleven of these processed genes are likely functional retrotransposed genes because they have low Ka/Ks ratios and high Ks values, and their sequences match numerous Arabidopsis ESTs. Processed sequences are mostly randomly distributed in the Arabidopsis genome and their rate of accumulation has steadily been decreasing since it peaked some 50 MYA. In contrast with the situation observed in mammals, the processed sequences found in the Arabidopsis genome originate from genes with high copy numbers and not from highly expressed genes. The patterns of spontaneous mutations in Arabidopsis are slightly different than those of mammals but are similar to those observed in Drosophila. This suggests that methylated cytosine deamination is less frequent in Arabidopsis than in mammals.

Arabidopsis↗
PubMed1985

Electric field mediated transformation: isolation and characterization of a TK+ subclone.

Transformation of mammalian TK- cells by a plasmid carrying the TK gene from Herpes virus simplex 1 (pAGO) was mediated by electroporation. The cells were treated either in suspension or growing in monolayers directly in the petri dish. The yield of transformation was between 8.10(-5) and 2.10(-4) per microgram DNA depending on the experimental conditions. The structure of the integrated DNA was investigated proving the occurrence of a duplication process that affected preferentially the pBR322 part of the pAGO DNA (60 copies per cell). The TK gene that gave the TK+ phenotype to the selected clone was present in less than 6 copies.

Animals↗
PubMed1983

Some stochastic models for plasmid copy number.

Some stochastic models for the copy number of plasmids in a cell line are studied. When considering the behavior of copy number in the whole cell line, the theory of multitype branching processes is appropriate. Attention is paid to the cure rate in the cell line, and the asymptotic fractions of cells containing a given number of plasmids. These quantities are used to compare the models numerically.

DNA Replication↗
PubMed1987

Induced sister chromatid exchange frequency is not increased in homogeneously staining regions that contain amplified genes.

Gene amplification is a process by which cells become resistant to selective agents by increasing gene copy number and overproducing specific enzymes. The molecular mechanism by which gene amplification occurs is unknown, but unequal sister chromatid exchange (SCE) has been suggested as one possibility. Unequal SCE results in one chromatid containing an extra copy of a selected gene that is deleted in the sister chromatid. Two predictions of the unequal SCE model are that agents that increase SCE frequency would increase gene amplification, and that SCE would be more prevalent in arrays of amplified units. We examined SCE frequency in the Chinese hamster ovary cell line MK42, which contains amplified dihydrofolate reductase genes that are stably integrated as a homogeneously staining region in chromosome #2. Under treatment conditions known to increase amplification of the dihydrofolate reductase gene, a number of agents did increase SCE frequency in MK42 cells. The frequency of SCE in the amplified region of chromosome #2, however, was no different from that expected if SCE were induced randomly as a function of chromosome length. These data, therefore, do not support the prediction of unequal SCE as a model for gene amplification. Our experiments, however, do not test the idea that unequal SCE at normal frequencies may lead to gene amplification, nor do they rule out the possibility of transient periods early in amplification, during which the homogeneously staining region may be a "hot spot" for SCE.

Animals↗
PubMed1988

Peptide radioimmunoassays: a BASIC program for data reduction.

A BASIC computer program is listed and described which may be used for calculating large amounts of data generated by radioimmunoassays. The user is given the opportunity to construct standard curves and to calculate mass quantities from raw counts or to incorporate previously determined protein values with mass quantities for final data expression. Calculated data may be viewed onscreen or obtained as printed copy.

Electronic Data Processing↗
PubMed1996

Structural characterization of the mouse ribosomal protein S6-encoding gene.

The gene encoding mouse ribosomal protein (r-protein) S6 is 2.7 kb in length, and is composed of five exons. The intron positions of the mouse S6 (Rps6) coincide exactly to those of the homologous human S6 (RPS6), but the last intron present in the human is absent in the mouse gene. The latter displays higher G + C content than the RPS6, both in the overall sequenced region and at the 3rd codon position. The promoter area is highly conserved between mouse and human, and contains several putative cis-acting elements. Comparison of the intronic sequences of both genes revealed surprisingly a high degree of identity (63%) within 350 bp of the first intron. Besides the single-copy Rsp6 there are up to 15 S6 family members, most likely processed pseudogenes. Characterization of the Rps6 provides a basis to study the functions of the mammalian S6 by gene targeting.

Amino Acid Sequence↗
PubMed2005

A hypomorphic allele of the first N-glycosylation gene, ALG7, causes mitochondrial defects in yeast.

The modification of proteins at asparagine residues with oligosaccharides (N-glycans) plays critical roles in diverse cell functions. N-glycans originate from a common lipid-linked oligosaccharide (LLO) precursor whose synthesis is initiated by the Dol-P-dependent GlcNAc-1-P transferase (GPT) encoded by an essential ALG7 gene. To identify cellular processes affected by ALG7 and N-glycosylation, we replaced the genomic copy of ALG7 with its hypomorphic allele in two genetically distinct haploid yeast cells. We show that ALG7 knockdown gave rise to an unexpected phenotype of mitochondrial dysfunction. The alg7 mutants did not grow on glycerol and DNA arrays revealed the absence of mitochondrial genes' expression. Accordingly, the alg7 mutants displayed no detectable mtDNA and respiratory activity. Both mutants exhibited diminished abundance of LLO and under-glycosylation of carboxypeptidase Y (CPY). Moreover, another N-glycosylation mutant with a LLO defect, alg6, was respiratory deficient. Collectively, our studies provide evidence that the dysregulation of N-glycosylation in haploid yeast cells leads to mitochondrial dysfunction.

3' Untranslated Regions↗
PubMed2004

Homing endonuclease genes: the rise and fall and rise again of a selfish element.

Homing endonuclease genes (HEGs) are selfish genetic elements that spread by first cleaving chromosomes that do not contain them and then getting copied across to the broken chromosome as a byproduct of the repair process. The success of this strategy will depend on the opportunities for homing--in other words, the frequency with which HEG(+) and HEG(-) chromosomes come into contact--which varies widely among host taxa. HEGs are also unusual in that the selection pressure for endonuclease function disappears if they become fixed in a population, which makes them susceptible to degeneration and imposes a need for regular horizontal transmission between species. HEGs will be selected to reduce the harm done to the host organism, and this is expected to influence the evolution of their sequence specificity and maturase functions. HEGs may also be domesticated by their hosts, and are currently being put to human uses.

Endonucleases↗