Biomedical subjects

M A Cross

Publications and source records attributed to M A Cross.

44 records · Page 3Linked to original sources

PubMed1990

HlyB-dependent secretion of hemolysin by uropathogenic Escherichia coli requires conserved sequences flanking the chromosomal hly determinant.

The synthesis and secretion of hemolysin (HlyA) by Escherichia coli are governed by four contiguous genes (hlyCABD) that are closely conserved on plasmids and, among human pathogenic strains, on the chromosome. We have previously shown that in plasmid pHly152 the coexpressed synthesis and export functions are uncoupled by intraoperon transcription termination, which is in turn alleviated by antitermination dictated in cis by a region upstream of the hly operon. In this study we describe an analogous region of ca. 1,100 base pairs flanking the chromosomal hly determinant of the uropathogenic strain E. coli 2001. This region had no significant effect on intracellular levels of hemolysin but activated strongly, both in cis and in trans, the specific hlyB-hlyD-dependent hemolysin secretion function. The secretion-activating region increased the transcription of the secretion gene hlyB, but the transcription effect was not as pronounced as that seen in the pHly152 determinant and was not evident when the region was present in trans to the hemolysin genes, suggesting that, in addition to transcriptional activation, the region may possibly exert a secondary posttranscriptional influence. Southern hybridizations with the 1,100-base pairs secretion-activating sequence showed low identity to plasmid pHly152 and no identity with total DNA from nonhemolytic uropathogenic E. coli or hemolytic isolates of Proteus vulgaris, Proteus mirabilis, and Morganella morganii. In contrast, hybridization to total DNA from hemolytic E. coli isolates belonging to different serotypes showed strong conservation of the activating sequence, indicating that it is an integral component of the chromosomal hly determinant that is widespread among uropathogenic E. coli.

Blotting, Southern↗

PubMed1990

Sensitivity to Ca2+ and the effects of a calcium channel antagonist in resistance vessels from two strains of genetically hypertensive rat.

Ca sensitivity and sensitivity to diltiazem were studied in two strains of genetically hypertensive rat [spontaneously hypertensive rats (SHRs) and genetically hypertensive (GH) rats] and their normotensive control strains [Wistar-Kyoto (WKY) and normal Wistar (N) rats] at two ages before and after establishment of significant hypertension. Ca sensitivity was equal in both hypertensive/normotensive strain pairs from young rats but significant relative increases in Ca sensitivity were present in older rats. Sensitivity to the vasodilating action of diltiazem in vessels precontracted using K+ was also present in adult rats. These results suggest that the abnormality in Ca sensitivity seen in genetic hypertension in rats is an acquired characteristic not involved in the etiology of hypertension.

Animals↗

PubMed1989

Breast conservation radiation therapy technique for women with large breasts.

A technique is presented for the conservative irradiation of women with huge (size "EE") breasts. Patients are treated in a modified lateral decubitus position with an immobilization device. The technique is easily reproducible and allows favorable cosmetic outcome. The isodose plots for treatment with this technique are compared to treatment in standard fashion. We feel that this reproducible technique offers breast conserving therapy to women with large breasts, without poor cosmesis.

Breast↗

PubMed1986

Analysis of the vegetative replication origin of broad-host-range plasmid RK2 by transposon mutagenesis.

A range of Tn1723 transposon mutants of the oriV region of broad-host-range plasmid RK2 have been isolated, and the internal EcoRI fragment of the transposon has been deleted from each to reduce the insertion size from 9.6 kb (Tn1723) to 35 bp (delta Tn1723). Sequencing from the delta Tn1723-derived EcoRI site has allowed the precise mapping of these insertions to various points dispersed through the origin region. Using these mutants we have determined which regions of oriV RK2 are of functional importance to plasmid establishment following transformation of the host species Escherichia coli, Pseudomonas putida, and P. aeruginosa. Insertions into an A/T-rich region, and a region containing five direct repeat sequences prevented successful transformation of each host species tested, but the continuity of sequences adjacent to the five repeats were essential only in E. coli and P. putida. The establishment and maintenance in E. coli of a mini-RK2 replicon was found to be inhibited by transcription from an inducible promoter positioned to read into oriV RK2 against the direction of replication. Assays of transcription emerging from Tn1723 demonstrated significant levels from one end of the transposon only. Four mutants with insertions downstream of oriV RK2 were unable to become established in E. coli, and contained Tn1723 in the orientation which would supply transcription toward the oriV RK2 region. These results demonstrate both that the sequence requirements for oriV RK2 function differ between host bacterial species, and that origin function may be further influenced by the genetic environment in which it lies.

Base Sequence↗

PubMed1985

Regulation of the trfA and trfB promoters of broad host range plasmid RK2: identification of sequences essential for regulation by trfB/korA/korD.

Using a plasmid containing a transcriptional fusion in which the E.coli galK gene is expressed from the trfB promoter of broad host range plasmid RK2 we show that transcription from the trfB promoter is repressed by the products of both the trfB and korB genes as we have previously predicted from the sequence homology of the trfA and trfB promoters and the fact that the trfA promoter is regulated by trfB and korB. These loci, trfB and korB are normally transcribed from the trfB promoter. Thus the trfB incC korB operon of RK2 is doubly autogeneously regulated. In addition, we describe the isolation and characterization of a mutant trfA promoter which has become insensitive to repression by trfB as a result of a point mutation within the inverted repeat sequence previously predicted to be the trfB protein binding site. These results provide strong evidence for our previously proposed model for control of transcription from the trfA and trfB promoters.

Bacterial Proteins↗

PubMed1985

Regulation of replication and maintenance functions of broad host-range plasmid RK2.

Replication of broad host-range plasmid RK2 depends on a cisacting vegatative replication origin oriVRK2 and the polypeptide product(s) of the trans-acting gene trfA as well as on host-specified products. The trfA gene is the second cistron in a polycistronic unit whose first cistron may be kilD, one of 4 known RK2-specified kil loci (kilA, B, C, and D) which are inhibitory for bacterial host or plasmid vector in the absence of kor functions which suppress in trans the effect of their respective kil genes. Transcription of the operon containing trfA is negatively regulated by the products of both the trfB locus (alias korD and korA) and korB. The loci, trfB and korB, are expressed from a single transcriptional unit which we propose to be negatively autoregulated by the products of both loci, although an additional, weaker and unregulated transcript may also express korB. While deletions in the oriVRK2 region have indicated the presence of copy number control elements adjacent to and possibly overlapping with the minimal oriVRK2 segment, the overriding control of copy number seems to reside in the trfB and korB loci which in conjunction appear to reduce expression of the trfA gene to levels limiting for replication. Coregulation of trfA with kil genes may indicate that kil genes play a role in plasmid maintenance other than replication.

Bacterial Proteins↗

PubMed1984

Analysis of copy number control elements in the region of the vegetative replication origin of the broad host range plasmid RK2.

Broad host-range plasmid RK2 is able to replicate in a controlled manner in most Gram negative bacterial species. To analyze the elements of its control mechanism, we have measured the copy number in Escherichia coli of mini-RK2 replicons isogenic except for defined deletions in regions adjacent to the vegetative replication origin, oriVRK2, which have previously been implicated in copy number control because of their expression of plasmid incompatibility. The results indicate that while the previously defined 700-bp HaeII oriVRK2 fragment carries one copy control element (copA), a second (copB) lies at least partly outside this fragment towards the tetracycline resistance genes of RK2. Deletions affecting both these regions give a mini replicon with a copy number of 35-40 compared with 4-7 for parental RK2. Further incompatibility experiments indicate that targets for both incA (copA) and incB (copB) lie within the 700-bp HaeII oriVRK2 fragment.

Base Sequence↗

PubMedDate not supplied

Prevention of increases in blood pressure and left ventricular mass and remodeling of resistance arteries in young New Zealand genetically hypertensive rats: the effects of chronic treatment with valsartan, enalapril and felodipine.

The relative efficacy of three antihypertensive drugs in the prevention of further elevation of blood pressure (BP) and cardiovascular structural remodeling in 4-week-old genetically hypertensive (GH) rats was studied by means of two complementary methods, stereology and myography. Four to 10-week-old GH rats were treated with valsartan (10 mg/kg/day), enalapril (10 mg/kg/day) or felodipine (30 mg/kg/day). Untreated GH and normotensive control rats of Wistar origin served as controls. Tail-cuff systolic SBP was measured weekly and left ventricular (LV) mass determined at the end of the experiment. Mesenteric resistance arteries (MRA) were either fixed by perfusion, embedded in Technovit and sections stained for stereological analysis, or mounted on a wire myograph for structural and functional measurements. BP and LV mass were significantly reduced by all drugs; decreases in BP and LV mass were smaller after felodipine treatment. Valsartan and enalapril caused a decrease in BP to normotensive control values. Felodipine kept BP at the 4-week level and prevented further rise with age. Valsartan caused hypotrophic outward remodeling of MRA, enalapril eutrophic outward remodeling and felodipine hypotrophic remodeling. Myograph measurements showed remodeling of the same order. While all drugs lowered the media/lumen ratio in GH to normal, the outward remodeling after valsartan and enalapril indicates that valsartan and enalapril might be more effective in reversing the inward remodeling of resistance arteries found in essential hypertension.

Animals↗