Mu gem2ts DNA integration is not necessary for induction of synchrony of cell division in Escherichia coli K12.

The gem2ts mutant of bacteriophage Mu induced synchrony of cell division on bacteria surviving infection. Induction of synchronous growth could also be observed as a response to the entire infected bacterial population, as in the case of infection of hic mutants, a peculiar class of gyrB alleles. After Mu wild-type or Mu gem2ts infection of hic mutants, there was a lack of viral DNA integration and replication, while phage gene expression (including that of A gene, coding for the transposase) seemed to be quite normal.

A method for DNA extraction from the desert cyanobacterium chroococcidiopsis and its application to identification of ftsZ.

A method was developed for extraction of DNA from Chroococcidiopsis that overcomes obstacles posed by bacterial contamination and the presence of a thick envelope surrounding the cyanobacterial cells. The method is based on the resistance of Chroococcidiopsis to lysozyme and consists of a lysozyme treatment followed by osmotic shock that reduces the bacterial contamination by 3 orders of magnitude. Then DNase treatment is performed to eliminate DNA from the bacterial lysate.

Regulation of the bacteriophage Mu gem operon.

The gem operon of bacteriophage Mu, responsible for the complex phenomenon of phage conversion, is included in the so called "semiessential early" region of phage DNA. Unlike the other early genes of the phage which are transcribed from the pe promoter, expression of the gem operon is driven by its own promoter, which escapes the control of the repressor. In fact, the transcript corresponding to gem was detected in immune lysogens by using a combined reverse transcription and a subsequent amplification of the resulting cDNA.

A spontaneous Escherichia coli K12 mutant which inhibits the excision-reintegration process of Mu gem2ts.

Escherichia coli K12 strains lysogenic for Mu gem2ts with the prophage inserted in a target gene (i.e., lacZ::Mu gem2ts lysogenic strains) revert to Lac+ by prophage precise excision with a relatively high frequency (about 1 x 10(-6)).

Knee joint space width measurement: an experimental study of the influence of radiographic procedure and joint positioning.

We studied the influence of the radiographic procedure and joint positioning on knee joint space width (JSW) in 10 healthy volunteers, and the intrareader reproducibility of JSW measurements on radiographs performed 2 weeks apart using a standardized procedure. Results show that a 5 or 10 downward inclination of the X-ray beam and 15 or 30 of induced external foot rotation significantly reduced JSW. In contrast, knee flexion increased JSW.

Reversal of Mu gem2ts-induced mutations.

Mutations induced by the integration of a Mu gem2ts mutant prophage can revert at frequencies around 1 x 10(-6), more than 10(4)-fold higher than that obtained with Mu wild-type. Several aspects characterize Mu gem2ts precise excision: (i) the phage transposase is not involved; (ii) the RecA protein is not necessary; and (iii) revertants remain lysogenic with the prophage inserted elsewhere in the host genome. In addition, prophage re-integration seems to be non-randomly distributed, whereas Mu insertion into the host genome is a transposition event without any sequence specificity.

A novel illegitimate recombination event: precise excision and reintegration with the Mu gem mutant prophage.

The bacteriophage Mu is known to insert its DNA more or less randomly within the Escherichia coli chromosome, as do transposable elements, but unlike the latter, precise excision of the prophage, thereby restoring the original sequence, is not observed with wild-type Mu, although it has been reported with certain defective mutants. We show here that the mutant prophage Mu gem2ts can excise precisely from at least three separate loci -- malT, lac and thyA (selected as Mal+, Lac+ and Thy+, respectively). This excision occurs under permissive conditions for phage development, is observed in fully immune (c+) lysogens, and is independent of RecA and of Mu transposase.

A possible linkage of HLA-DRB haplotypes with Tiopronin intolerance in rheumatoid arthritis.

To investigate the relationship between HLA class II genotypes and toxic intolerance during treatment with Tiopronin, a slow-acting drug used in the treatment of rheumatoid arthritis (RA), we studied 40 patients who were divided into two groups: a group of 22 patients without side effects and a group of 18 patients with intolerance to Tiopronin. The PCR-RFLP method was used to determine the HLA-DR, DQ and DP genotypes. The patients in the two groups had similar genetic backgrounds with an expected high frequency of DRI and DR4 alleles.

The Mu gem operon: its role in gene expression, recombination and cell cycle.

Two genes, gemA and gemB, belong to the gem operon located in the semi-essential early region of bacteriophage Mu. The product of gemA modulates the expression of various host genes, including cell division and DNA replication genes. In addition, GemA is also responsible for decreasing host DNA gyrase activity and for DNA relaxation.

[A comparative controlled trial of 2 administration modalities of tiopronin in rheumatoid arthritis].

Thiopronin is a second line drug for rheumatoid arthritis with proven efficacy in controlled trials versus a placebo, D-penicillamine, or gold salts. This 4-month study was aimed mainly at comparing the efficacy and safety of two thiopronin regimens, i.e.

Interleukin-1 and naproxen down-regulate the expression of IL-1 receptors in cultured human rheumatoid synovial cells (HRSC).

Using affinity binding of [125I]-IL-1 alpha and Scatchard analysis, we demonstrate here that exposure of cultured synovial cells (HRSC) to NPX (10(-4) M) for 96 h decreased the binding of IL-1 by 20 to 35%. This effect results from a down-regulation of the IL-1 receptors without change in the apparent binding affinity (kD: 770 pM). Pretreatment of cultures with IL-1 alpha (500 pg/ml) reduced the total binding of [125I]-IL-1 alpha on HRSC by 65%, indicating that IL-1 decreases the expression of its own receptors.

[Rheumatoid arthritis. Changes in lymphocyte subsets under the effect of tiopronin].

Lymphocytes from 12 rheumatoid arthritis patients were phenotyped before and after a 2-month treatment with tiopronin. Originally reduced, CD4 CD45RA-T lymphocytes were shown to augment significantly. Abnormal activation (evaluated on HLA-DR and CD25 expression) of each cell population and sub-population was partially amended.

Tiopronine-induced reduction of rheumatoid factor functional affinity and asialylated IgG in patients with rheumatoid arthritis.

Serum and synovial fluid (SF) from 16 rheumatoid arthritis patients were evaluated before and after a 2-month treatment with tiopronine (TP). The levels of rheumatoid factor (RF) declined, as did the functional affinity of the remaining RF (p less than 0.01 in serum and p less than 0.

A class of gyrB mutants, substantially unaffected in DNA topology, suppresses the Escherichia coli K12 ftsZ84 mutation.

Previous work in our laboratory suggested that DNA topology could be implicated in the regulation of the division gene ftsZ. To settle this question, we have selected and characterized mutants in the gyrB gene able to phenotypically suppress the defects of the ftsZ84 mutation. No strict correlation was found between the degree of plasmid DNA relaxation and the level of suppression of the thermosensitivity of the ftsZ84 strain.

[Decrease of the functional affinity of rheumatoid factors under the effect of tiopronin].

Sixteen rheumatoid arthritis patients were serologically examined, before and after a 2-month treatment with tiopronine (TP). The titer of agglutinating rheumatoid factor (RF) as well as the level of non-agglutinating RF were significantly reduced in serum and synovial fluid (SF). The functional affinity of the remaining RF was attenuated in serum (p less than 0.

Synchronous division induced in Escherichia coli K12 by phage Mu: analysis of DNA topology and gene expression during the cell cycle.

Bacteriophage Mu mutants in gene gem (Mu gemts2) induce cycles of synchronous divisions after infection of a bacterial population in steady-state conditions. In this paper, two classes of gyrB mutants, synchronizable and non-synchronizable, are described. The existence of the non-synchronizable class suggests that the gyrase B subunit is involved with Gem in the process of synchronization.

Global changes in gene expression in Escherichia coli K12 induced by bacteriophage Mu Gem protein.

We have studied the growth properties of some Mu lysogens with respect to the non-lysogenic strain and have observed that the division time in minimal medium was increased over 4-fold when the bacteria carried the prophage mutated in the gem gene (Mu gem3). Since this phage gene has previously been shown to be involved in modulation of expression of host genes, we have analysed the proteins extracted from lysogens and non-lysogens as a rapid assay of global gene expression. The pattern of proteins extracted showed marked quantitative variations between non-lysogens, lysogens for wild-type Mu and lysogens for phage Mu gem3.

The bacteriophage Mu gem gene: a positive regulator of the C operon required for normal levels of late transcription.

The gem product of bacteriophage Mu modulates synthesis of various host proteins and alters the host chromosome topology. To elucidate the role of the gem gene in Mu development, we analyzed the behavior of several mutants in this gene. The results, obtained with two Mu gem- phages, show that (1) phage growth is significantly delayed and inhibited, (2) early transcription is normal but late transcription is delayed and reduced, (3) DNA replication appears normal, and (4) the Mu C gene, whose product positively regulates Mu late genes, is one of the gem target sites.

Mu gem3 as a tool to investigate the influence of chromosome supercoiling on gene expression in Escherichia coli K12.

We have developed a rapid method to investigate the influence of chromosome supercoiling on gene expression in Escherichia coli K12. This method exploits the ability of the gem3 mutant of the bacteriophage Mu, even in the prophagic state in immune cells, to induce relaxation of the host chromosome. The experiments can thus be performed under physiological conditions, and without the use of the drugs.

Synchronous division induced in Escherichia coli K12 by gemts mutants of phage Mu.

Infection with the bacteriophage mutant Mu c+ gemts2 at 42 degrees C induces synchrony in cell division in cultures of Escherichia coli K12. This synchrony may last for several cycles and is not only due to selection since synchronization is observed even when bacterial survival to the infection is over 80% as in lysogens for Mu c+ gemts2. The mechanism by which synchrony is induced is not known, but since the product of Mu gene gem (previously called lig) has been shown to interact with the enzymatic system in the bacteria controlling the degree of DNA supercoiling, the phenomenon could be a consequence of this interaction.

The expression of the DNA ligase gene of Escherichia coli is stimulated by relaxation of chromosomal supercoiling.

Many genes of Escherichia coli have been shown to be sensitive to DNA superhelicity. The superhelicity of the chromosome is itself also supercoiling-dependent. We have developed a general strategy for investigating how a particular gene responds to changes in DNA topology.

Suppression of the thermosensitive DNA ligase mutations in Escherichia coli K12 through modulation of gene expression induced by phage Mu.

We have previously shown that Mu can sustain the growth at non-permissive temperature of an Escherichia coli strain harbouring a thermosensitive mutation in the DNA ligase structural gene. This "complementation" reaches a maximal level with the Mu lig3 mutant which restores the viability of a ligts7 strain to the level of the wild type (Ghelardini et al. 1980; Paolozzi et al.