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Biomedical subjects

S Iida

Publications and source records attributed to S Iida.

At least 487 records · Page 27Linked to original sources
PubMed1981

Cointegrates between bacteriophage P1 DNA and plasmid pBR322 derivatives suggest molecular mechanisms for P1-mediated transduction of small plasmids.

We characterized cointegrates formed in an Escherichia coli rec+ strain between bacteriophage P1 genomes and small plasmids related to pBR322. The partners were, on the one hand, either phage P1 DNA, which carries one copy of IS1, or phage P1-15 DNA, a derivative which lacks the IS1, and, on the other hand, plasmids containing either a split IS1 or no. In the presence of IS1 sequences on both partners, cointegrates were usually formed by reciprocal recombination between SI1 sequences. Cointegrates between P1 and a plasmid carrying no IS1 sequence were formed by transpositional cointegration mediated by IS1 of P1. Cointegrates between P1-15 and small plasmid containing a split IS1 were formed by one of three ways: (a) acquisition of an IS1 by P1-15 followed by reciprocal recombination between IS1 sequences, (b) transpositional cointegration mediated by the split IS1 element, Tn2657, or (c) involvement of the invertible segment carried on P1-15 DNA. Most cointegrates segregated into the small plasmids and phage P1 derivatives. A comparison of the phenomenon studied and of their frequencies allowed us to conclude that cointegrate formation is a molecular mechanism involved in the transduction of plasmids smaller than those packageable into P1 virions, although it does not seem to be the only process used.

Bacteriophages↗
PubMed1981

The plasmid cloning vector pBR325 contains a 482 base-pair-long inverted duplication.

The plasmid pBR325 is a cloning vector constructed in vitro by addition of the chloramphenicol resistance (Cmr) gene of an IS1-flanked transposon to pBR322 (Bolivar, 1978). It is a 5 995 bp plasmid carrying no sequence originating from IS1. DNA-sequence data suggest that its Cmr segment was derived from a Cm transposon longer than Tn9. The plasmid pBR325 carries between the Cmr and Tcr genes a 482 bp sequence which duplicates, in the opposite orientation, a section pf pBR322 located at the end of the tcr gene. The same structure was found in pBR328, a deletion derivative of pBR325 (Soberon et al., 1980). The possible implications of this inverted duplication on cloning experiments are discussed.

Base Sequence↗
PubMed1981

Is the IS1-flanked r-determinant of the R plasmid NR1 a transposon?

The 23 kilobase multiple drug resistance r-determinant (r-det) of the R plasmid NR1 is an IS1-mediated transposon, Tn2671. Drug-resistant Escherichia coli transductants isolated after infection with bacteriophage P1::Tn2671 derivatives carry the intact r-det in their chromosomes. Independently isolated transductants carry the r-det at different locations on the chromosome. From the E. coli chromosome, Tn2671 can transpose to various locations on the phage P7 genome. Throughout these processes, r-det is maintained as a stable unit. Various possible molecular mechanisms, which all might contribute with characteristic frequencies to the transposition of Tn2671, are discussed. The results presented are relevant to the understanding of mechanisms for a wide spreading of drug resistance genes.

Chloramphenicol↗
PubMed1981

Genesis and natural history of IS-mediated transposons.

The natural genesis of IS1-mediated transposons containing the genetic determinant cat for chloramphenicol resistance is documented. First, the small plasmid pBR325 containing the cat gene served as a target in IS1-mediated transpositional cointegration with the genome of bacteriophage P1, which was the source of the IS1. From the resulting pBR325:P1 plasmids, pBR325::IS1 segregants were isolated. Upon growth of a phage lambda derivative in the presence of this plasmid, rare plaque-forming lambda Cmr specialized transducing phages were formed. In each of six independent lambda Cmr isolates studied, the cat gene was carried between flanking IS1 elements. In one case, these IS1 elements were in the same orientation; in the other five cases, they were in opposite orientation. All of these IS1-cat-IS1 structures transposed as units to the genome of phage P1-15, pointing to stable maintenance of the transposon. However, appropriate selection allowed us to follow the decay of these transposons. Models to explain the genesis of transposons with directly and inversely repeated IS elements are discussed, as well as the evolutionary implications of these mechanisms.

Bacteriophage lambda↗
PubMed1981

Protein synthesis in the isolated forespores from sporulating cells of Bacillus subtilis.

Developing forespores were isolated from Bacillus subtilis at different stages of sporulation and protein synthesis in the forespore compartment was examined. Pulse-labeling experiments indicated that [14C]phenylalanine was continuously incorporated into the sporangium throughout sporulation, and at t5 (early stage V of sporulation) 58% of the radioactivity was located in the forespore compartment. Significantly high incorporation of [14C]phenylalanine was observed when the isolated forespores at t5 were incubated with the corresponding mother-cell cytoplasmic fraction or an amino acid mixture. About 73% of the radioactivity incorporated into the isolated forespore at t5 was found in the cytoplasmic fraction and 26% in the membranous fraction. Analysis by sodium dodecyl sulfate-gel electrophoresis showed that the 14C-labeled cytoplasmic protein had a molecular weight of about 20,000, and that a protein having the same molecular weight was present in the t5 forespore as a slight protein band and also in the mature spore as a clear protein band. Gel electrophoresis also revealed that the 14C-labeled membranous-soluble protein (prepared by solubilization with detergents) had broad peaks with molecular weights of about 74,000, 33,000, 20,000, and 12,000.

Bacillus subtilis↗
PubMed1981

Presence of ACTH-potentiating factors in rat anterior pituitary glands.

High molecular weight ACTH fractions, obtained through gel filtration of boiled rat anterior pituitary extract, induced a marked increase in corticosterone production from isolated rat adrenal cells in the presence of low concentrations of ACTH-(1-24). This indicates the presence of heat-stable factors augmenting the steroidogenic action of ACTH in the rat anterior pituitary. We also noted that these factors potentiated the activity not only of ACTH-1(1-24) but also of ACTH-(1-8). The ACTH-potentiating factors in rat anterior pituitary extract are possibly present in heterogeneous forms according to their molecular weights (8,000, 10,000 and 15,000), their mobility in ion-exchange chromatography and their content in RIA-ACTH activity. Of these three forms, the former comigrated with biological ACTH activity. The remaining two forms were free of it. Since the effect of potentiating factors on modified ACTH-(1-9), shown to be less susceptible to proteolytic degradation from ACTH-(1-24), was similar to the effect on ACTH-(1-24), it is suggested that potentiation was not due to an inhibition of ACTH proteolysis.

Adrenal Glands↗
PubMed1981

Changes in distribution of molecular weight forms of biologically active and immunoreactive adrenocorticotropic hormone after adrenalectomy in rat anterior pituitary.

The concentration of ACTH in extracts of rat anterior pituitary was measured by both radioimmunoassay and bioassay at different stages following adrenalectomy. Both types of ACTH activity decreased the day immediately following adrenalectomy but increased gradually afterwards. Immunological ACTH activity increased to 250% of the control value and biological ACTH activity increased to 490% of control value 3 weeks after adrenalectomy. The increase in biological ACTH activity occurred earlier, and the rate of increase was greater, than that of the immunological ACTH activity. The distributions of molecular weight forms of ACTH in extracts of anterior pituitary lobes was determined by gel filtration. Three molecular weight forms of immunoassayable ACTH were detected. Biological ACTH activity appeared in the 2nd and the 3rd peaks. A striking change was observed after adrenalectomy in the distribution of biologically active forms of ACTH. The ratio of biological ACTH activity to immunological ACTH activity in each peak changed at various stages after adrenalectomy. This indicated the heterogenous nature of the ACTH included in each peak. At 2 and again at 3 weeks, biological activity markedly increased until it exceeded the immunological ACTH activity in the 2nd peak. Dexamethasone had little influence on the elution profile of either immunoassayable and biologically active ACTH in gel filtration. Adrenalectomy may possibly have an effect on the intracellular posttranslational processing of ACTH precursors which leads to the development of biological ACTH activity.

Adrenalectomy↗