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Biomedical subjects

T Iino

Publications and source records attributed to T Iino.

At least 127 records · Page 7Linked to original sources
PubMed1983

A new class of site-specific endodeoxyribonucleases. Endo.Sce I isolated from a eukaryote, Saccharomyces cerevisiae.

We had found that yeasts had intracellular endodeoxyribonucleases that cut phage DNA into a set of double-stranded fragments with discrete chain lengths. We purified one of them to apparent homogeneity from Saccharomyces cerevisiae and designated it Endo.Sce I. Sequence analysis around 5 cleavage sites in plasmid DNA and phage DNA revealed that Endo.Sce I cuts a defined phosphodiester bond in each strand of double helix at the cleavage sites and produces free cohesive ends consisting of 4 nucleotides protruding at 3'-termini. However, unlike in the case of prokaryotic type II-restriction endonucleases, (i) Endo.Sce I seems to consist of two nonidentical subunits, (ii) no common palindrome or consensus sequence including more than 5 base pairs is detected at or near these cleavage sites, and (iii) Endo.Sce I can cut the DNA isolated from the cells that produced Endo.Sce I. All of the 5 cleavage sites are included in inverted repeats, but these inverted repeats are variable in size, nucleotide sequence, and distance between repeating units. An inverted repeat itself is not a structure recognized by Endo.Sce I. This study shows that Endo.Sce I is the first example of eukaryotic site-specific endonuclease and has properties, as described above, which distinguish it from prokaryotic restriction endonucleases.

Base Sequence↗
PubMed1983

Transfer inhibition of RP4 by F factor.

When RP4 and F factors were brought together into one E. coli cell, the F factor reduced 500-1000-fold the frequency of transfer of RP4. However, F had almost no effect on the surface exclusion and pilus formation by RP4. In contrast, RP4 did not affect the transfer of F. Using in vitro recombinant DNA techniques, a gene of F responsible for the above-mentioned transfer inhibition of RP4 was located within the BamHI fragment (40.4-42.8 kb) of the mini-F sequence on F. From the result of product analysis using minicells, the responsible gene in the BamHI fragment was inferred to encode the 33 K protein.

Chromosome Mapping↗
PubMed1983

Characterization of the mgl operon of Escherichia coli by transposon mutagenesis and molecular cloning.

We used transposon insertion mutagenesis, molecular cloning, and a novel procedure for in vitro construction of polar and nonpolar insertion mutations to characterize the genetic organization and gene products of the beta-methylgalactoside (Mgl) transport system, which utilizes the galactose-binding protein. The data indicate that the mgl operon contained three genes, which were transcribed in the order mglB, mglA, and mglC. The first gene coded for the 31,000 Mr galactose-binding protein, which was synthesized as a 3,000-dalton-larger precursor form. The mglA product was a 50,000 Mr protein which was tightly associated with the membrane, and the mglC product was a 38,000 Mr protein which was apparently loosely associated with the membrane and was probably located on the internal face of the cytoplasmic membrane. Identification of gene products was facilitated by in vitro insertion of a fragment of Tn5 containing the gene conferring kanamycin resistance into a restriction site in the operon. The fragment proved to have a polar effect on the expression of promoter-distal genes only when inserted in one of the two possible orientations. The three identified gene products were necessary and apparently sufficient for transport activity, but only the binding protein was required for chemotaxis towards galactose. The transport system appeared to contain the minimum number of components for a binding protein-related system: a periplasmic recognition component, a transmembrane protein, and a peripheral membrane protein that may be involved in energy linkage.

Bacterial Proteins↗
PubMed1983

Ordering of the flagellar genes in Pseudomonas aeruginosa by insertions of mercury transposon Tn501.

The flagellar genes of Pseudomonas aeruginosa PAO cluster on the chromosome at two distinct regions, region I and region II. The order of the flagellar cistrons in this organism was established by using transducing phage G101 and plasmids FP5 and R68.45. A method to insert transposon Tn501 near the fla genes was devised. We obtained two strains in which Tn501 was inserted at sites close to the flagellar cistrons in region II. We isolated Fla mutants in which the chromosomal segment between the two Tn501 insertion sites was deleted. Using Tn501-encoded mercury resistance as an outside marker, we determined the order of 9 of the 11 flagellar cistrons in region II as follows: puuF-region I-flaG-flaC-flaI-flaH-flaD-flaB-flaA-flaF-flaE-pur-67. By using phage G101-mediated transduction, the mutation converting monoflagellated bacteria into the multiflagellated (mfl) form was closely linked to the five fla cistrons in region I. Using mfl as an outside marker, we determined the order of the five cistrons as follows: puuF-flaV-flaZ-flaW-flaX-flaY-region II. The mfl mutation was shown to be either located within the flaV cistron or linked very closely to this cistron. No linkage was observed in transductions between any of the fla cistrons in region I and any of the fla cistrons in region II.

Chromosome Mapping↗
PubMed1983

Transductional analysis of the flagellar genes in Pseudomonas aeruginosa.

Complementation in bacteriophage E79 tv-l-mediated transduction and the phenotypic properties of the flagellar genes in Pseudomonas aeruginosa PAO were investigated by using 195 flagellar mutants of this organism. A total of 15 fla. 1 mot, and 2 che cistrons were identified. At least 5 fla cistrons (fla V to flaZ) and one mot cistron resided in one region, and at least 10 fla cistrons (flaA to flaJ) and two che cistrons (cheA and cheB) resided in another. The flaC mutants exhibited cistron-specific leakiness on motility agar plates. The flaE cistron may be the structural gene for the component protein of the flagellar filament. The cheA mutations, which resulted in pleiotropic phenotypes for flagellar formation, motility, and taxis, belonged to the same complementation group as the flaF mutations; that is, we inferred that cheA and flaF are synonymous.

Flagella↗
PubMed1983

Genetic mapping of bra genes affecting branched-chain amino acid transport in Pseudomonas aeruginosa.

Pseudomonas aeruginosa PAO mutants defective in the transport systems for branched-chain amino acids were isolated and characterized. Two mutations in strains selected for trifluoroleucine resistance, braA300 and braB307, were mapped in the met-9020-dcu-9108 and the nar-9011-puuC10 region, respectively. The mutation loci in strains selected for azaleucine resistance, braC310 and bra-311 through bra-314, were all located near the fla genes, with an order of region I fla-bra-region II fla. Strains with braA300 showed a marked reduction in the high-affinity branched-chain amino acid transport system (LIV-I) and a considerable decrease in the lower-affinity system (LIV-II). Strains with braB307 were found to be defective in the LIV-II system. Strains selected for azaleucine resistance were all defective only in the LIV-I system and fell into three phenotypically distinct classes. Strains with braC310 produced a binding protein for leucine, isoleucine, valine, alanine, and threonine (LIVAT-BP) altered in binding ability, indicating that the braC gene is the structural one for the LIVAT-BP. Strains with bra-311 or bra-312 showed a complete loss of production of the LIVAT-BP. Strains with bra-313 or bra-314 produced normal levels of functional LIVAT-BP, suggesting that these mutations are located in a gene(s) other than braC.

Amino Acids, Branched-Chain↗
PubMed1983

Unstable angina--clinical manifestations and prognosis of 74 medically-treated patients.

Seventy-four patients with unstable angina are reviewed. A rather high incidence (65%) of a changing pattern syndrome was observed in the study group. A history of prior myocardial infarction was obtained in 22 patients (30%). Fifty-seven patients (77%) were investigated using coronary angiography: 15 did not have any significant occlusive disease but are included in this study; single vessel disease was present in 20, double in 8 and triple in 14. Ten patients (13.5%) had myocardial infarcts which had occurred between 48 hours and 3 months after admission, and no patients died during this period. Patients who on admission had resting angina of new onset were more likely to have a worse short-term prognosis. Four patients underwent coronary bypass surgery in the first year of the follow-up period because of unsuccessful medical treatment. Six patients died within the first 4 years. Of the 6 deaths, 5 had triple vessel disease and one did not have any significant coronary stenosis. Mortality rate at 2 years was 2.7% and at 4 years 10.3%. This low mortality is probably attributable to a low incidence of 2- or 3-vessel disease.

Adult↗
PubMed1982

Genetic block of outer plaque morphogenesis at the second meiotic division in an hfd1-1 mutant of Saccharomyces cerevisiae.

An hfd1-1 mutant of Saccharomyces cerevisiae, SOS4, characterized by predominant production of two-spored asci at 29 degrees C, undergoes normal meiotic nuclear divisions and produces four haploid nuclei, but only two non-sister nuclei among them are incorporated into mature ascospores. Spindle pole bodies and prospore wall membranes at the second meiotic division at 29 degrees C were observed in this mutant by electron microscopy. The spindle pole body at one pole of each spindle had a normal outer plaque which was larger than the inner plaque. At the other pole, the outer plaque was entirely absent and a normal prospore wall membrane was not detected. It was concluded that at 29 degrees C the hfd1-1 mutation blocks the morphogenesis of outer plaques and prospore wall membranes at the two non-sister poles in the second meiotic division, and consequently only non-sister nuclei in the resulting meiotic cell are incorporated into ascospores.

Meiosis↗
PubMed1982

Cloning of trg, a gene for a sensory transducer in Escherichia coli.

Clones of trg, a gene which codes for a chemotactic transducer, were isolated linked to ColE1 and pBR322 vectors. Studies with the hybrid plasmids demonstrated unequivocally that trg is the structural gene for methyl-accepting chemotaxis protein III. The Trg protein was found to be structurally complex, electrophoresing as a series of seven bands on high-resolution sodium dodecyl sulfate-polyacrylamide gels. The multiplicity of bands is a function of the activity of cheR, which codes for a methyltransferase, and of cheB, which codes for a demethylase. It appears that Trg, a quantitatively minor transducer, resembles the two major transducer proteins, Tsr and Tar, in that all three are multiply methylated and also multiply modified in a second way which requires an active cheB gene. However, preliminary analysis of the Trg protein indicated that it is significantly less related structurally to the Tsr or Tar protein than those two transducers are to each other. This implies that the features of multiple methylation and cheB-dependent modification are likely to be critical for the common physiological functions in chemotactic excitation and adaptation performed by all three transducers.

Bacterial Proteins↗
PubMed1982

Acute toxicity study of KB-944, a new calcium antagonist.

The acute toxicity of diethyl 4-(benzothiazol-2-yl)benzylphosphonate (KB-944) was studied in mice, rats and dogs: In mice and rats, toxic symptoms such as deceleration of spontaneous movement, ataxic gait, disappearance of righting reflex, reduction in body temperature and suppression of breathing appeared following administration of a single dose of KB-944. The symptoms developed in dogs were depression, bradycardia and labored breathing. The LD50 values determined in male and female rats were 1622 and 1555 mg/kg, respectively, for oral administration, 2216 and 2820 mg/kg for subcutaneous injection, and 762 and 479 mg/kg for intraperitoneal injection. The LD50 values determined in male and female mice were 2807 and 3856 mg/kg, respectively, for oral administration, greater than 4000 mg/kg for subcutaneous injection, and 815 and 777 mg/kg for intraperitoneal injection. As shown above, there was no evident difference in LD50 between sexes in either species of animals. Since many of the dogs given an oral dose larger than 1000 mg/kg vomited the drug, oral LD50 value could not be determined.

Animals↗
PubMed1982

1-month subacute oral toxicity study of KB-944, a new calcium antagonist, in rats.

Diethyl 4-(benzothiazol-2-yl)benzylphosphonate (KB-944), a new Ca-antagonist, in the dose range of 25--200 mg/kg/day, was orally administered to Jcl:SD rats for five consecutive weeks and the following results were obtained. Neither death nor inhibition of body weight gain nor any toxic symptoms of the drug were noted in rats but an increase in water intake was found in the rats treated with over 100 mg/kg/day of KB-944. In plasma there was a decrease in alkaline phosphatase activity, creatinine level and cholinesterase activity and an increase in GPT activity and total cholesterol level in rats given a higher dosage of the drug. Weight gain of the liver, kidneys, heart, adrenals and ovaries, and degeneration in the epithelium of the renal proximal tubule were observed. However, none of these changes were serious and the maximum non-toxic dose of KB-944 was 25 mg/kg/day.

Animals↗
PubMed1982

1-month subacute oral toxicity study of KB-944, a new calcium antagonist, in beagles.

Using male and female Beagles, we studied the subacute toxicity of diethyl 4-(benzothiazol-2-yl)benzylphosphonate (KB-944) administered orally and the following results were obtained. No dogs died during the administration period. Both male and female dogs given 200 mg/kg and those given 100 mg/kg underwent body weight decrease and inhibition of body weight increase, respectively. Enlargement of liver cells occurred in male dogs given 200 mg/kg and female dogs given 100 or 200 mg/kg. There were no further findings, even histopathologically, which showed any toxic effect of the drug. The maximum non-toxic dose of this drug was considered to be 30 mg/kg.

Animals↗
PubMed1981

Tn1 insertion mutagenesis in Escherichia coli K-12 using a temperature-sensitive mutant of plasmid RP4.

A method for Tn1 insertion mutagenesis in Escherichia coli has been developed using pTH10, a mutant plasmid of RP4 temperature-sensitive for maintenance. The mutagenesis involves three steps. Firstly, from strains carrying pTH10 showing resistance to the antibiotics kanamycin, tetracycline, and ampicillin at 30 degrees C but not at 42 degrees C, clones are isolated resistant to kanamycin at 42 degrees C. Such temperature-independent, drug resistant clones probably carry pTH10 integrated into the host chromosome. Secondly, they are cultivated at 30 degrees C. At this temperature segregants carrying pTH10, which has been excised from the host chromosome, accumulate. Thirdly, to cure such segregants of autonomous pTH10, they are cultivated at 42 degrees C. By these procedures, clones free of pTH10, but carrying Tn1 insertions on the host chromosome, were obtained. About 3% of the clones carrying Tn1 insertions were auxotrophic. Distribution of auxotrophic mutations was not random, indicating the existence of preferential integration sites of Tn1 on the host chromosome. The frequency of precise excision of Tn1 was less than 10(-10). The pTH10 plasmid has a wide host range among Gram-negative bacteria and thus may serve as a excellent vector for insertion mutagenesis of Tn1 in many Gram-negative bacterial species.

DNA Transposable Elements↗
PubMed1981

An immunization method using antigen entrapped in erythrocyte ghosts.

Intraperitoneal injection into mice of dinitrophenol-conjugated ovalbumin entrapped in autologous erythrocyte ghosts gave rise to an increase of anti-dinitrophenol antibody production without use of artificial adjuvants. This technique could be useful for vaccination or other immuno-therapeutic purposes.

Animals↗
PubMed1981

Selective abortion of two nonsister nuclei in a developing ascus of the hfd-1 mutant in Saccharomyces cerevisiae.

A recessive mutation, hfd1-1, in strain SOS4 of Saccharomyces cerevisiae leads the mutant cells to produce predominantly two-spored asci. Light microscopical examination of Giemsa-stained cells revealed no significant differences in the meiotic figures between mutant and wild-type strains. However, only two of the four meiotic products in a developing ascus matured to ascospores in SOS4. Dyad analysis was carried out on an hfd1-1 mutant strain heterozygous for three markers, asp5, gal1, and arg4, which are closely linked to their centromeres, and for his4, which is loosely linked to its centromere. The two-spored asci produced by the hfd1-1 mutant segregated dominant (+) and recessive (-) alleles of each marker in a 1:1 ratio; they generally contained one + and one - spore for any given marker. The occurrence of rare dyads with two + or two - spores can be explained quantitatively by recombination between the marker and its centromere. From the results of these cytological and genetical analyses, we infer that, in the mutant strain, one genome set is partitioned to each of the four second-meiotic division poles, but only two nonsister genomes are incorporated into mature spores. Thus, the hfd1-1 mutation in SOS4 blocks incorporation of two nonsister nuclei into mature ascospores, but does not block enclosure of the remaining two nonsister nuclei.

Genes, Recessive↗
PubMed1981

A simple method for the isolation of flagellar shape mutants in Salmonella.

During prolonged static cultivation of liquid cultures of Salmonella strains with normal flagella, flagellar shape mutants occurred spontaneously and their proportion in the cultures increased so that they were easily isolated by spreading dilutions of the cultures on semisolid medium. By this method various types of flagellar shape mutants, including curly, straight and polymorphous mutants, were obtained.

Flagella↗