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J F Viret

Publications and source records attributed to J F Viret.

16 recordsLinked to original sources

Transient expression from cab-m1 and rbcS-m3 promoter sequences is different in mesophyll and bundle sheath cells in maize leaves.

Cell-specific and light-regulated expression of the beta-glucuronidase (GUS) reporter gene from maize cab-m1 and rbcS-m3 promoter sequences was studied in maize leaf segments by using an in situ transient expression microprojectile bombardment assay. The cab-m1 gene is known to be strongly photoregulated and to be expressed almost exclusively in mesophyll cells (MC) but not in bundle sheath cells (BSC). Expression of GUS from a 1026-base-pair 5' promoter fragment of cab-m1 is very low in dark-grown leaves; GUS expression is increased about 10-fold upon illumination of dark-grown leaves. In illuminated leaves, the ratio of GUS expression in MC vs. BSC is about 10:1. The cab-m1 region between 868 and 1026 base pairs 5' to the translation start confers strong MC-preferred expression on the remainder of the chimeric gene in illuminated leaves, but a region between -39 and -359 from the translation start is required for photoregulated expression. Transcripts of rbcS-m3 are found in BSC but not in MC and are about double in BSC of greening dark-grown seedlings. In contrast to the behavior of the cab-m1-GUS construct, GUS expression driven by 2.1 kilobase pairs of the rbcS-m3 5' region was about twice as high in MC as in BSC of unilluminated dark-grown maize leaves. The number of BSC, but not MC, expressing GUS nearly doubled upon greening of bombarded etiolated leaves. These data suggest that the 5' region of rbcS-m3 used here could be responsible for most of the light-dependent increase in rbcS-m3 transcripts observed in BSC of greening leaves and that transcriptional or posttranscriptional mechanisms are responsible for the lack of rbcS-m3 transcripts in MC.

Base Sequence

The cab-m7 gene: a light-inducible, mesophyll-specific gene of maize.

Southern blot analysis has revealed the existence in maize of perhaps 12 members of the nuclear cab multigene family encoding the chlorophyll a- and b-binding proteins of the photosystem II light-harvesting complex. Hybridization with 3' probes derived from unsequenced cDNA clones showed that six members of this family differ from one another with respect to expression in mesophyll and/or bundle sheath cells and regulation by light. An additional member of this family, designated cab-m7, that encodes a 28 kDa primary translation product has now been identified. It has been cloned from a maize genomic library and sequenced to begin to define the bases for differences in the expression of these genes. This cab gene is shown to be strongly preferentially expressed in the mesophyll (vs. bundle sheath) cells of maize. Furthermore, the gene is photo-responsive; although small amounts of cab-m7 mRNA are present in etiolated leaves, the mRNA pool is 8-fold larger after six hours of illumination. DNA sequences upstream of the cab-m7 gene resemble those found in the 5'-flanking regions of some other plant genes.

Amino Acid Sequence

Characterization of the Shigella serotype D (S. sonnei) O polysaccharide and the enterobacterial R1 lipopolysaccharide core by use of mouse monoclonal antibodies.

In the course of developing a live vaccine, we generated three murine monoclonal antibodies (MAb) specific for Shigella sonnei. The specificities of these MAb were determined by enzyme-linked immunosorbent assay and immunoblot analyses with whole cells or purified lipopolysaccharides (LPSs) as antigens. Two of them are specific for the Shigella serotype D O-polysaccharide determinant, whereas one specifically binds to the core hexose region of R1-type LPSs. With these MAb, it was possible to analyze clinical isolates and a hybrid Salmonella typhi strain for their expression of the corresponding LPS moieties. In addition to their use in the screening of candidate vaccine strains, the new MAb provide a powerful tool for epidemiological and phylogenetic studies of natural enterobacterial populations.

Animals

Recombination-dependent concatemeric plasmid replication.

The replication of covalently closed circular supercoiled (form I) DNA in prokaryotes is generally controlled at the initiation level by a rate-limiting effector. Once initiated, replication proceeds via one of two possible modes (theta or sigma replication) which do not rely on functions involved in DNA repair and general recombination. Recently, a novel plasmid replication mode, leading to the accumulation of linear multigenome-length plasmid concatemers in both gram-positive and gram-negative bacteria, has been described. Unlike form I DNA replication, an intermediate recombination step is most probably involved in the initiation of concatemeric plasmid DNA replication. On the basis of structural and functional studies, we infer that recombination-dependent plasmid replication shares important features with phage late replication modes and, in several aspects, parallels the synthesis of plasmid concatemers in phage-infected cells. The characterization of the concatemeric plasmid replication mode has allowed new insights into the mechanisms of DNA replication and recombination in prokaryotes.

Bacteria

Cloning and nucleotide sequence of the phosphoenolpyruvate carboxylase-coding gene of Corynebacterium glutamicum ATCC13032.

As a first step in determining the importance of the anaplerotic function of phosphoenolpyruvate carboxylase (PEPC) in amino acid biosynthesis, the ppc gene coding for PEPC of Corynebacterium glutamicum ATCC13032 has been cloned by complementation of an Escherichia coli ppc mutant strain. PEPC activity encoded by the cloned gene is not affected by acetyl-CoA under conditions where the E. coli enzyme is strongly activated, whereas acetyl-CoA is able to relieve inhibition by L-aspartate used singly or in combination with alpha-ketoglutarate. Amplification of the ppc gene in a C. glutamicum lysine-excreting strain resulted in increased PEPC-specific activity and lysine productivity. The nucleotide sequence of a DNA fragment of 4885 bp encompassing the ppc gene has been determined. At the amino acid level, PEPC from C. glutamicum presents overall a high degree of similarity with corresponding enzymes from three different organisms. The location of some strictly conserved regions may have important implications for PEPC activity and allostery.

Amino Acid Sequence

A DNA sequence outside the pUB110 minimal replicon is required for normal replication in Bacillus subtilis.

The origin of lagging strand synthesis in pUB110, oriL, has been localized within 140 bases outside the pUB110 minimal replicon. The oriL DNA sequence is a cis-acting and orientation dependent determinant required for normal plasmid replication. Rearrangements affecting oriL cause plasmid instability, lead to the accumulation of replication intermediates and result in a marked reduction of the plasmid copy number in some recombination deficient mutant strains. In addition, deletion of oriL triggers a dnaB-dependent mode of replication. Insertion of the functionally asymmetric oriL region in the proper orientation into pC194 reduces the accumulation of single-stranded DNA during the replication of this plasmid.

Bacillus subtilis

Functional analysis of the dna (Ts) mutants of Bacillus subtilis: plasmid pUB110 replication as a model system.

We determined the effect of various Bacillus subtilis dna(Ts) mutations on pUB110 and chromosomal replication. Leading strand DNA synthesis of pUB110, starting by a nick at the plasmid replication origin (oriU), is performed by DNA polymerase III, since replication is blocked at non-permissive temperature in thermosensitive mutants dnaD, dnaF, dnaH and dnaN known to cause thermosensitivity of the various subunits of DNA polymerase III. When the lagging strand origin (oriL) is exposed, the DnaG protein (DNA primase) alone, or in association with unknown protein(s) binds asymmetrically to oriL to form a primer that is also extended by DNA polymerase III. In oriL- plasmids like pBT32, leading and lagging strand DNA syntheses are decoupled from each other. The DnaB protein, that is not required for pUB110 replication, may be associated with priming at a second unidentified lagging strand origin on pBT32. At non-permissive temperature, the dnaC30 and dnaI2 mutations affect both pUB110 and chromosomal DNA synthesis.

Bacillus subtilis

Replication and incompatibility properties of plasmid pUB110 in Bacillus subtilis.

Within plasmid pUB110 we have identified a 1.2 kb segment necessary and sufficient for driving autonomous replication in Rec+ cells at a wild-type copy number. This region can be divided into three functionally discrete segments: a 24 base pair (bp) region that acts as an origin, a 949 bp determinant of an essential replication protein, repU, and a 358 bp incompatibility region, incA, overlapping with the repU gene. The synthesis of the IncA determinant/s proceeds in the direction opposite to that of RepU. The positively (RepU) and negatively (IncA) trans-acting products seem to be involved in the control of plasmid replication. The RepU product has an Mr of 39 kDa, could be overproduced in Escherichia coli, and binds to the pUB110 origin region. Outside the minimal replicon a cis-acting, orientation dependent, 516 bp determinant is required (i) to compete with a coexisting incompatible plasmid and (ii) for segregational stability.

Bacillus subtilis

Generation of linear multigenome-length plasmid molecules in Bacillus subtilis.

Linear multigenome-length double and single stranded plasmid DNA was identified in a Bacillus subtilis ATP-dependent DNAase mutant strain (addA5) bearing plasmids pC194 or pBD95ts. Plasmid pBC30, a seg mutant of pC194, as well as some pUB110 derivatives with rearrangements external to the minimal replicon, produce high amounts of such a concatemeric DNA, even in Rec+ cells. The synthesis of this type of plasmid DNA occurs in the absence of an active plasmid-encoded Rep protein and is markedly affected in polA5 and recE4 genetic backgrounds. To account for these observations, we propose that the AddAB complex serves to prevent a sigma-type replication of plasmid DNA.

Bacillus subtilis

Plasmid maintenance in Bacillus subtilis recombination-deficient mutants.

An isogenic set of 11 recombination-deficient mutant strains of Bacillus subtilis has been constructed. Whereas plasmid pUB110 is stably maintained in such Rec- cells, the high copy number plasmid pC194 is unstable. Instability in Rec- strains could be mostly attributed to the deleterious effect of the presence of the plasmid on the Rec- cells' growth capability. In part, instability of pC194 derivatives could also be correlated with the presence of an unusually high amount of multimeric DNA molecules.

Bacillus subtilis

A new mutator strain of Bacillus subtilis.

Bacillus subtilis strain SB1207, widely used in our laboratory, was found to be highly temperature-sensitive and to exhibit a strong SOS-independent mutator phenotype at elevated temperatures. Both chromosomal and plasmid-borne genes were affected by the mutator. Lethality and mutator phenotype could not be attributed to a replication shut off or to thymine starvation. Due to the high frequency of base misincorporation, the mutator phenotype probably results from an editing defect rather than from a post-replication defect (mismatch repair).

Bacillus subtilis

Cloning of the recA gene of Bordetella pertussis and characterization of its product.

A recA gene of Bordetella pertussis was identified in a plasmid library by complementation of a recA mutation in E coli and subcloned as a 2.1-kb Sph I DNA fragment. Southern hybridization experiments showed no similarity to the E coli recA gene, but very strong similarity to other Bordetella species. E coli recA mutant cells containing the B pertussis recA gene at high gene dosage were resistant to DNA-damaging agents such as methyl methane sulfonate or 4-nitroquinoline-N-oxide, displayed induction of SOS functions, and were able to promote DNA recombination, but not induction of phage lambda. The latter phenotype distinguishes the B pertussis recA gene product from the corresponding proteins from most other Gram-negative organisms. Amino acid sequence comparisons revealed a high degree of structural conservation between prokaryotic RecA proteins.

Amino Acid Sequence

Morphological and cell wall alterations in thermosensitive DNA mutants of Bacillus subtilis.

Incubation of thermosensitive dna mutants of Bacillus subtilis at the nonpermissive temperature results, at the cellular level, in the appearance of swellings. The study of one particular type of swelling, named "terminal balloon", reveals that the occurrence of the latter was correlated with completion of rounds of DNA replication. Morphological and autoradiographic observations reveal that (a) cell wall consists of two layers, (b) the outer layer splits at a fixed distance from the cell pole and allows the formation of a balloon contained within a single wall layer and (c) the bulk of active synthesis of cell wall is localized in the balloon area leading to the formation of a near spherical monolayer. Implications of these results for cell wall morphogenesis are discussed.

Bacillus subtilis