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G Mosig

Publications and source records attributed to G Mosig.

At least 37 records · Page 2Linked to original sources
PubMed1989

Impaired expression of certain prereplicative bacteriophage T4 genes explains impaired T4 DNA synthesis in Escherichia coli rho (nusD) mutants.

The Escherichia coli rho 026 mutation that alters the transcription termination protein Rho prevents growth of wild-type bacteriophage T4. Among the consequences of this mutation are delayed and reduced T4 DNA replication. We show that these defects can be explained by defective synthesis of certain T4 replication-recombination proteins. Expression of T4 gene 41 (DNA helicase/primase) is drastically reduced, and expression of T4 genes 43 (DNA polymerase), 30 (DNA ligase), 46 (recombination nuclease), and probably 44 (DNA polymerase-associated ATPase) is reduced to a lesser extent. The compensating T4 mutation goF1 partially restores the synthesis of these proteins and, concomitantly, the synthesis of T4 DNA in the E. coli rho mutant. From analyzing DNA synthesis in wild-type and various multiply mutant T4 strains, we infer that defective or reduced synthesis of these proteins in rho 026-infected cells has several major effects on DNA replication. It impairs lagging-strand synthesis during the primary mode of DNA replication; it delays and depresses recombination-dependent (secondary mode) initiation; and it inhibits the use of tertiary origins. All three T4 genes whose expression is reduced in rho 026 cells and whose upstream sequences are known have a palindrome containing a CUUCGG sequence between the promoter(s) and ribosome-binding site. We speculate that these palindromes might be important for factor-dependent transcription termination-antitermination during normal T4 development. Our results are consistent with previous proposals that the altered Rho factor of rho 026 may cause excessive termination because the transcription complex does not interact normally with a T4 antiterminator encoded by the wild-type goF gene and that the T4 goF1 mutation restores this interaction.

Bacterial Proteins↗
PubMed1989

Functional relationships and structural determinants of two bacteriophage T4 lysozymes: a soluble (gene e) and a baseplate-associated (gene 5) protein.

Lysozymes have proved useful for analyzing the relation between protein structure and function and evolution. In bacteriophage T4, the major soluble lysozyme is the product of the e gene, gpe (gene product = gp). This lysozyme destroys the wall of its host, Escherichia coli, at the end of infection to release progeny particles. Phage T4 contains two additional lysozymes that facilitate penetration of the baseplates into host cell walls during adsorption. At least one of these, a 44-kD protein, is encoded by gene 5. We show here that a segment of the gp5 lysozyme amino acid sequence, deduced from the DNA sequence of gene 5, is remarkably similar to that of the T4 gene e lysozyme. Both T4 lysozymes are somewhat similar to the lysozyme of the Salmonella phage P22, but there is little significant DNA sequence homology among the two T4 lysozyme genes and the P22 lysozyme gene. We speculate that these lysozymes are adapted to differences in the composition of the cell walls of E. coli and S. typhimurium. The cloned gene 5 of the phage T4 directs synthesis of a 63-kD precursor protein that is approximately 19 kD larger than the gene 5 protein isolated from baseplates. Gp5 first associates with gp26 to form the central hub of this structure.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1988

Integration host factor (IHF) represses a Chlamydomonas chloroplast promoter in E. coli.

We show that in E. coli, a Chlamydomonas chloroplast promoter, PA, is repressed by Integration Host Factor (IHF). The himA 42 mutation, altering the alpha-subunit of E. coli IHF, leads to over-accumulation of PA transcripts in vivo. This effect requires upstream chloroplast DNA sequences. DNAase I and methylation protection experiments show that IHF binds in vitro to a site within PA and band-retardation shows that IHF inhibits formation of PA-E. coli RNA polymerase open complexes. We interpret these results, together with our previous deletion analyses, to mean that in E. coli, repression of PA by IHF minimally requires both binding of IHF to a site overlapping PA and binding of one or more additional proteins, perhaps including IHF itself, to sequences upstream of PA.

Bacterial Proteins↗
PubMed1988

Regulation of two nested proteins from gene 49 (recombination endonuclease VII) and of a lambda RexA-like protein of bacteriophage T4.

Phage T4 gene 49, encoding recombination endonuclease VII, specifies, by initiation from an AUG and an internal GUG codon, two in-frame overlapping peptides (of 18 and 12 kD). The gene is transcribed early and late, albeit from different promoters. The sequence predicts that in long early transcripts, initiated far upstream of the coding sequence, the Shine-Dalgarno sequence of the first ribosome binding site can be sequestered in a hairpin and/or cleaved. These processes might reduce initiation from the first AUG and facilitate initiation of the 12-kD peptide from the internal GUG. The potential of this hairpin to participate in Y structures or cruciforms suggests possible autoregulation. Shorter, more stable late transcripts initiated from a late promoter immediately upstream of the first ribosome binding site cannot form this hairpin. More efficient translation of the longer 18-kD gene 49 peptide from these late transcripts accounts for the strong dependence of endonuclease VII activity on late gene expression. An ORF downstream from gene 49 can be translated from a motA-dependent transcript that starts inside gene 49 as well as from the gene 49 transcripts. Its initiation codon overlaps the stop codon of gene 49, suggesting some coupling of translation. The deduced protein resembles, among others, the RexA protein of phage lambda. Possible implications for T4 recombination and for the interference of lambda lysogens with T4 gene 49 and rII mutants are discussed.

Amino Acid Sequence↗
PubMed1988

Roles of the Escherichia coli heat shock sigma factor 32 in early and late gene expression of bacteriophage T4.

We have analyzed early and late T4 gene expression at the levels of transcription and translation in rpoH+ (sigma 32+) and rpoH mutant cells infected under heat shock conditions. We found, as expected, that Escherichia coli cells must be adapted before infection to high temperature by the heat shock response to allow early T4 transcription, subsequent late gene expression, and progeny production at 42 degrees C. Unexpectedly, we found in addition that when rpoH mutant (sigma 32 mutant) cells were shifted from 30 to 42 degrees C 10 min after infection, late T4 genes were not expressed, even though DNA synthesis appeared to be normal.

Escherichia coli↗
PubMed1987

Stimulation of a Chlamydomonas chloroplast promoter by novobiocin in situ and in E. coli implies regulation by torsional stress in the chloroplast DNA.

We have characterized regulation of a complex Chlamydomonas reinhardtii chloroplast (PA) whose activity is stimulated by the DNA gyrase inhibitor novobiocin, both in the alga itself and in a heterologous Escherichia coli plasmid system. Since novobiocin is known to reduce torsional stress in E. coli DNA, we interpret our results to mean that PA is regulated by torsional stress in the chloroplast DNA. In E. coli, where we could readily manipulate PA, we found that this regulation depends on sequences upstream of PA. These sequences contain at least two different kinds of silencing elements that inhibit PA in the absence of novobiocin. Novobiocin stimulates PA only when the promoter-distal silencing element is present.

Base Sequence↗
PubMed1986

A new membrane-associated DNA replication protein, the gene 69 product of bacteriophage T4, shares a patch of homology with the Escherichia coli dnaA protein.

A new phage T4 DNA replication protein, gp69, is found to be associated with membrane fractions, as predicted by the translated base sequence of gene 69. In addition, gp69 shares a patch of homology with a segment of the Escherichia coli dnaA initiation protein. The patchy homology of dnaA protein and gp69 suggests that they may serve some similar functions, such as interactions with the same E. coli components in bacterial and viral DNA replication. We have shown before that gene 69 spans an origin of T4 DNA replication, and that this origin is preferentially associated with membrane fractions. We suggest the possibility that gp69 is involved in the attachment of this origin to the bacterial envelope.

Amino Acid Sequence↗
PubMed1986

Sequence and transcripts of the bacteriophage T4 DNA repair gene uvsY.

We have cloned, sequenced and analyzed transcription of the phage T4 uvsY gene. This gene is transcribed from a single gp MotA-dependent middle promoter to give a major transcript of approximately 930 nucleotides and a minor transcript of approximately 620 nucleotides. All in vivo and in vitro uvsY transcripts show anomalous migration in agarose gels. The uvsY transcript contains an open reading frame coding for an 137 amino acid [15.8 kilodaltons (kD)] UvsY protein and two unidentified open reading frames, ORF UvsY.-1 (9.0 kD) and ORF UvsY.-2 (6.0 kD). Our DNA sequence differs in only three places from that published by TAKAHASHI et al. However, one of these changes alters the predicted carboxy terminus of the UvsY protein. Marker rescue experiments map gene 25 to the region upstream of uvsY. Gene 25 is likely, although not certain, to correspond to an ORF that is found upstream from uvsY and is translated in the same direction.

Amino Acid Sequence↗
PubMed1985

An ATP-dependent supercoiling topoisomerase of Chlamydomonas reinhardtii affects accumulation of specific chloroplast transcripts.

We have found that Chlamydomonas reinhardtii cells contain an ATP-dependent topoisomerase activity that supercoils circular DNA in vitro. Subsequent addition of a type I topoisomerase eliminates the supercoils. Like bacterial gyrase, this activity is inhibited by low concentrations of novobiocin (less than 0.1 microM) and by nalidixic acid (less than 0.1 microM). We have examined the effects of these topoisomerase inhibitors on accumulation of various chloroplast transcripts in vivo. Novobiocin differentially affected such transcripts; some transcripts became more abundant while many others were reduced in the presence of this drug. Nalidixic acid on the other hand caused many transcripts to become more abundant albeit to varying degrees. Inhibitors of this algal topoisomerase specifically stimulate a family of related transcripts which we have previously shown to be under light-dark control. We discuss how the inhibitors of this topoisomerase might exert their in vivo effects.

Chlamydomonas↗
PubMed1985

Bacteriophage T4 gene 32 participates in excision repair as well as recombinational repair of UV damages.

Gene 32 of phage T4 has been shown previously to be involved in recombinational repair of UV damages but, based on a mutant study, was thought not to be required for excision repair. However, a comparison of UV-inactivation curves of several gene 32 mutants grown under conditions permissive for progeny production in wild-type or polA- hosts demonstrates that gene 32 participates in both kinds of repair. Different gene 32 mutations differentially inactivate these repair functions. Under conditions permissive for DNA replication and progeny production, all gene 32 mutants investigated here are partially defective in recombinational repair, whereas only two of them, P7 and P401, are also defective in excision repair. P401 is the only mutant whose final slope of the inactivation curve is significantly steeper than that of wild-type T4. These results are discussed in terms of interactions of gp32, a single-stranded DNA-binding protein, with DNA and with other proteins.

DNA Repair↗
PubMed1985

;Dark-Lethality' of Certain Chlamydomonas reinhardtii Strains Is Prevented by Dim Blue Light.

Cells of certain Chlamydomonas reinhardtii strains dramatically swell and ultimately disintegrate when grown in the dark but not in the light. This effect of darkness is delayed: cells undergo about two divisions before swelling. Measurements on O(2) uptake and ATP content indicate that the cells grown in the dark are not deficient in energy supply. Dim blue light not effective in photosynthesis prevents ;dark-lethality'. Inhibition of protein synthesis during the latent period with cycloheximide also prevents dark-lethality. We propose that blue light stimulates synthesis of one or more proteins which are part of or are important in assembly of an osmoregulatory complex.

Journal Article↗
PubMed1985

Common evolutionary origin of the phage T4 dam and host Escherichia coli dam DNA-adenine methyltransferase genes.

We compared the known DNA nucleotide and encoded amino acid sequences of the Escherichia coli and bacteriophage T4 dam (DNA-adenine methyltransferase) genes. Despite the absence of any DNA sequence homology, there were four regions (11 to 33 residues long) of amino acid sequence homology containing 45 to 64% identity. These results suggest that the genes for these two enzymes have a common evolutionary origin.

Adenine↗
PubMed1984

Regulation of a new bacteriophage T4 gene, 69, that spans an origin of DNA replication.

We have determined the DNA sequence and transcription patterns in a 3-kb segment (between 15 and 18 kb on the standard phage T4 map) spanning an origin of DNA replication. A new gene, 69, spans this origin. Gene 69 codes for two overlapping proteins that share a common C-terminal segment. Defective DNA replication in an appropriate amber mutant shows that at least the larger of the two proteins is required for efficient T4 DNA replication. The two proteins coded by gene 69 are expressed from different transcripts that are under different regulation. The smaller protein, gp69*, can be expressed immediately from an Escherichia coli-like promoter, whereas expression of the larger protein, gp69, must be delayed since its middle promoter requires T4 coded proteins, most likely gp mot, for activation. We discuss the possible significance of two overlapping proteins in the assembly of replisomes. Gene 69 is bracketed by the non-essential early gene dam (DNA adenine methylase) and the late gene soc (small outer capsid protein). Transcripts through this region are interdigitated in a complex pattern, which reveals all elements that are thought to be important in regulation of pre-replicative and post-replicative T4 genes.

Amino Acid Sequence↗
PubMed1984

Cloning and physical mapping of an early region of the bacteriophage T4 genome.

We have cloned DNA restriction fragments from the largely nonessential region of bacteriophage T4 located between genes 39 and 56. The cloned DNA fragments were used to construct a precise map of the sites in this region recognized by eight restriction endonucleases. This restriction map allowed us to compare the cytosine-containing T4 DNA used for cloning with the hydroxymethylcytosine-containing DNA of wild-type T4; there were no detectable rearrangements in the region tested. We were also able to determine the physical locations of several deletion end points and of several genes.

5-Methylcytosine↗
PubMed1984

Regulation of a bacteriophage T4 late gene, soc, which maps in an early region.

We have sequenced and analyzed the expression of an early region of the bacteriophage T4 genome that surprisingly contains a late gene, soc. soc is oriented in the same direction as early genes, like the T4 lysozyme gene. Northern hybridization of early and late T4 RNA, using cloned T4 restriction fragments as probes, identified two long early transcripts and a short late transcript, all containing the soc-coding sequence. Thus, soc is transcribed both early and late. It is, however, translated only late. The inhibition of soc translation from the long early transcripts can be explained by formation of a hairpin in the RNA that sequesters the soc ribosome-binding site. The transcript initiated at the late promoter cannot form this hairpin and is, therefore, translated.

Base Sequence↗