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C A Kraft

Publications and source records attributed to C A Kraft.

14 recordsLinked to original sources
PubMed2001

Agonist-dependent traffic of raft-associated Ras and Raf-1 is required for activation of the mitogen-activated protein kinase cascade.

Stimulation of HIRcB fibroblasts with insulin leads to accumulation of active components of the mitogen-activated protein kinase cascade in endocytic compartments. However, the factors that regulate the mobilization of these components through the endocytic pathway and the relevance of this event to cellular signaling remain unclear. Here we report that Ras proteins are associated with lipid rafts in resting HIRcB fibroblasts. Ras is rapidly internalized into the endocytic compartment following stimulation with insulin. The redistribution of Ras is independent of its activation. Attachment of the C-terminal 20 amino acids of Ha-Ras to green fluorescent protein was sufficient to target this construct to the same loci as the endogenous Ras protein, indicating that Ras distribution is a consequence of the association of its lipid modified C terminus with membranes. Depletion of plasma membrane cholesterol delocalized Ras and blocked insulin-dependent Ras traffic. Cholesterol depletion also blocked insulin-dependent phosphorylation of MEK and mitogen-activated protein kinase (MAPK) but had no effects on the translocation and activation of Raf-1. A second inhibitor of endocytosis, cytochalasin D, also blocked insulin-dependent MAPK phosphorylation. Taken together, these results suggest that mobilization of active Raf-1 through the endocytic compartment is required for completion of the MAPK cascade.

Animals↗
PubMed1987

Restriction enzyme fingerprinting of trimethoprim resistance plasmids.

Restriction enzyme fingerprinting was applied to 72 transferable trimethoprim resistance plasmids to examine aspects of their epidemiology and molecular relatedness. These plasmids had previously been divided into 25 groups according to differences in mol. wts and in antimicrobial resistance determinants. Restriction enzyme fingerprinting allowed the plasmids to be further divided into 44 different groups. The groups based on molecular weight and resistance patterns often, but not invariably, corresponded with those based on restriction enzyme fingerprints. Some plasmids with the same mol. wt and resistance pattern had different digest fingerprints and conversely, although more rarely, plasmids which differed in molecular weight by as much as 10 MDa or in resistance pattern by one resistance marker, had indistinguishable fingerprints. The plasmids were initially divided into three broad categories according to which restriction enzymes gave fingerprints of 6-20 fragments. These categories differed in the molecular weights of the plasmids contained, the numbers of resistance markers, and the proportions of the plasmids which carried transposon Tn7. Some plasmids were more widespread and persistent than others with the same mol. wt and resistance pattern but with a different restriction enzyme fingerprint. Thus, application of this technique has shown the trimethoprim resistance plasmids studied to be more diverse than was indicated by determination of mol. wt and resistance pattern, and has indicated changes in the plasmid pool over the 3 years during which they were collected.

Molecular Weight↗
PubMed1986

Restriction enzyme fingerprinting of enterobacterial plasmids: a simple strategy with wide application.

Restriction enzyme fingerprints were generated from purified plasmid DNA from 324 clinical isolates that belonged to 7 enterobacterial genera and 88 single plasmids in Escherichia coli K 12 according to the following strategy. Purified plasmid DNA was digested with PstI. The number of fragments detected in a 0.8 agarose gel was used to determine which 2 of 6 restriction enzymes including PstI was most likely to provide a fingerprint comprising sufficient fragments to ensure specificity but sufficiently few to allow easy visual assessment and minimize coincidental matching. When PstI produced greater than 20 fragments, EcoRI and HindIII were used; when PstI generated less than 6 fragments Bsp 1286 and AvaII were used and SmaI was employed when between 6 and 20 fragments were obtained from PstI digests. Using a minimum of 12 fragments from a combination of 2 enzymes as the criterion for characterizing a strain/plasmid, satisfactory 2-enzyme fingerprints were obtained from 87% of the strains and plasmids studied using PstI and no more than two additional enzymes per strain. Of the remaining 54 strains, 51 harboured only small plasmids (less than 10 kb) and 3 produced satisfactory fingerprints when digested with a fourth enzyme.

Chromosome Mapping↗
PubMed1986

Distribution and genetic location of Tn7 in trimethoprim-resistant Escherichia coli.

A series of 178 strains of Escherichia coli, highly resistant to trimethoprim, isolated from hospital patients and patients in the community between 1979 and 1983, was examined for the presence of Tn7 on a plasmid or on the chromosome only, by transposition to RP4 and restriction endonuclease digestion with Hind III. Of the isolates, 57% carried Tn7. Comparison of hospital isolates from 1979 to 1980 and 1982 showed that although the proportions that carried Tn7 were similar (63% and 57%) there had been a significant change in the genetic location of the transposon. The proportion of plasmid-mediated Tn7 had fallen from 62% to 30% with a corresponding rise in Tn7 located exclusively on the chromosome from 38% to 70%. This change may be the result of continuing transposition of Tn7 from plasmids to the bacterial chromosome followed by plasmid loss. The consequent reduction in the mobility of trimethoprim-resistance genes may in turn lead to changes in the incidence of resistance.

Chromosomes, Bacterial↗
PubMed1985

Trimethoprim resistance in urinary coliforms from patients in the community: plasmids and R-transfer.

A study of urinary coliform isolates, from patients attending their general practitioner, between 1981 and 1983 showed that 10.2% were resistant to trimethoprim. Forty-four trimethoprim-resistant Escherichia coli were collected for further study and the results compared with those obtained from hospital isolates collected during 1979-80 and 1982. Thirty-two (73%) were highly resistant to trimethoprim (MIC greater than 1024 mg/l) a similar proportion to that in the hospital isolates. The highly resistant isolates bore a close resemblance to the hospital isolates in that 66% carried multiple plasmids, 97% were multiply drug-resistant and all were highly resistant to sulphamethoxazole. The frequency of antimicrobial resistance transfer (59%) was similar to that from the hospital isolates. More than half of the trimethoprim resistance plasmids characterized were indistinguishable in terms of mol.wt and resistance pattern from those found in isolates from the hospital collections. Our results suggest that, whereas trimethoprim-resistant E. coli are less commonly isolated from patients in the community than in hospitals, many of the community isolates may have originated from a hospital source.

Conjugation, Genetic↗
PubMed1984

Antimicrobial resistance and the ecology of Escherichia coli plasmids.

Four hundred and seven clinical isolates of Escherichia coli were examined for the presence of plasmids. These isolates comprised 189 which were collected irrespective of antimicrobial resistance (VP) and 218 which were collected on the basis of high-level trimethoprim resistance (TPR). The VP isolates were divided into drug sensitive (VPS) and drug-resistant (VPR) subpopulations. Plasmids were detected in 88% of VP isolates (81% of VPS and 94% of VPR) and 98% of TPR isolates. The distribution of plasmids in both groups and subpopulations was very similar. However, there were small but statistically significant differences between the plasmid distributions. These showed that more isolates in the resistant groups harboured plasmids than in the sensitive subpopulation (VPS) and that the number of plasmids carried by resistant isolates was greater. Multiple drug resistance was significantly more common among TPR isolates than the VPR subpopulation and this was paralleled by increased numbers of plasmids. Fifty-eight per cent of VPR and 57% of TPR isolates transferred antimicrobial resistance and plasmids to E. coli K12. Of the R+ isolates, 60% carried small plasmids (MW less than 20Md) and 52% of these co-transferred with R-plasmids. These results are discussed.

Ampicillin↗
PubMed1984

Distribution and transferability of plasmids in trimethoprim-resistant urinary strains of Escherichia coli: a comparative study of hospital isolates.

Trimethoprim-resistant urinary isolates of Escherichia coli, collected in 1982, were studied and the results compared with those obtained for isolates collected during the period 1979-1980. Ninety-eight (81%; a 10% increase) were resistant to trimethoprim 1024 micrograms/ml and 93 (95%) were also resistant to sulphamethoxazole 1024 micrograms/ml. The frequency distributions of plasmids were similar in both collections although there was a significant increase in the number of small plasmids (mol. wt less than or equal to 20 X 10(6] in the 1982 collection. Transfer of resistance was associated with isolates that carried larger numbers of plasmids. A significantly smaller proportion of isolates in this, than in the earlier series, transferred trimethoprim resistance to E. coli K12 suggesting continued transposition of trimethoprim resistance on to the bacterial chromosome. Fifteen different trimethoprim resistance plasmids were identified, of which five, including that found most frequently, were common to both collections. Plasmids which transferred trimethoprim resistance without sulphonamide resistance were more common in the 1982 series. Plasmids which transferred linked trimethoprim and streptomycin resistances, in particular those not carrying other resistance markers, were less common in the 1982 series of isolates. Although the majority of E. coli isolates highly resistant to trimethoprim remained resistant to sulphonamide, our results suggested changes in the genetic location and linkage of the resistance markers.

Anti-Bacterial Agents↗
PubMed1983

Distribution and transferability of plasmids in trimethoprim resistant urinary Escherichia coli.

Urinary isolates of Escherichia coli that were resistant to trimethoprim were collected in Glasgow Royal Infirmary during 1979 and 1980. Eighty-eight were resistant to trimethoprim 1024 micrograms/ml and 80 (92%) were also resistant to sulphamethoxazole 1024 micrograms/ml; 73% were multiresistant. Plasmids were detected in 98% of strains and 60% carried two or more. Half of the isolates transferred trimethoprim resistance to E. coli K12 and 70% of these cotransferred resistance to sulphonamide although these markers were often not linked. Trimethoprim resistance was carried on 12 different plasmids, four of which also conferred sulphonamide resistance. All except two carried streptomycin resistance which suggests that Tn7 was probably present. The results are discussed in relation to current prescribing policy.

Anti-Bacterial Agents↗
PubMed1981

The effect of aging on beta-adrenoceptor-stimulated cyclic AMP formation in human lymphocytes.

1. Responsiveness of the beta-adrenoceptor adenylate cyclase system was measured in lymphocytes from healthy young and old subjects by incubating the cells with isoprenaline in the presence of a phosphodiesterase inhibitor and by measuring production of adenosine 3':5'-cyclic monophosphate (cyclic AMP) with a competitive binding assay. 2. The two groups did not differ significantly in the levels of cyclic AMP produced or in the concentration of isoprenaline required to give half-maximal stimulation of the cells (ED50).

Adolescent↗
PubMed1980

Improved hypnotic treatment using chlormethiazole and temazepam.

The effects of a single 384 mg oral dose of chlormethiazole were compared with those of 20 mg of temazepam and placebo in healthy old and young women (mean ages 72.9 and 24.7 years respectively). Both drugs were effective hypnotics and had no detectable pharmacological action the next morning. Even four hours after administration performance of a simple psychomotor test was not impaired and sway (measured by an ataxiameter) was not increased in either age group. Pharmacokinetic studies showed that chlormethiazole was rapidly absorbed, distributed, and eliminated by both groups, so that minimal plasma concentrations existed 11 hours after administration. Temazepam, however, was less quickly absorbed and distributed, especially in the young group, and substantial amounts remained in the plasma 11 hours after administration. No unwanted effects occurred after temazepam, but 17 of the 20 subjects suffered from nasal irritation after taking chlormethiazole. Thus hangover effects may be avoided in elderly subjects after they have taken hypnotic drugs, and temazepam and chlormethiazole allow sleep to be interrupted safely.

Adult↗