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Biomedical subjects

Y Jinno

Publications and source records attributed to Y Jinno.

At least 109 records · Page 6Linked to original sources
PubMed1988

Mutational analysis of domain I of Pseudomonas exotoxin. Mutations in domain I of Pseudomonas exotoxin which reduce cell binding and animal toxicity.

Pseudomonas exotoxin (PE) is a single polypeptide chain that contains 613 amino acids and is arranged into three structural domains. Domain I is responsible for cell recognition, II for translocation of PE across membranes and III for ADP ribosylation of elongation factor 2. Treatment of PE with reagents that react with lysine residues has been shown to lead to a reduction in cytotoxic activity apparently due to a modification of domain I (Pirker, R., FitzGerald, D. J. P., Hamilton, T. C., Ozols, R. F., Willingham, M. C., and Pastan, S. (1985) Cancer Res. 45, 751-757). To determine which lysine residues are important in cell recognition, all 12 lysines in domain I were converted to glutamates by site-directed mutagenesis. Also, two deletion mutants encompassing almost all of domain I (amino acids 4-252) or most of domain I (amino acids 4-224) were studied. The mutant proteins were produced in Escherichia coli, purified, and tested for their cytotoxic activity against Swiss 3T3 cells and in mice. The data indicate that conversion of lysine 57 to glutamate reduces cytotoxic activity towards 3T3 cells 50-100-fold and in mice about 5-fold. Deletion of amino acids 4-224 causes a similar reduction in toxicity towards cells and mice. Deletion of most of the rest of domain I (amino acids 4-252) causes a further reduction in toxicity toward cells and mice indicating this second region between amino acids 225 and 252 of domain I is also important in the toxicity of PE. Competition assays indicated that the ability of PEGlu57 to bind to 3T3 cells was greatly diminished, accounting for its diminished cytotoxic activity.

ADP Ribose Transferases↗
PubMed1988

A novel effect of EGF on mRNA stability.

Expression of the epidermal growth factor (EGF) receptor gene is stimulated by EGF and the phorbol ester, 4 beta-phorbol 12-myristate 13-acetate (PMA). PMA elevates EGF receptor mRNA levels in human KB epidermoid carcinoma cells, but does not significantly affect the half-life of this mRNA when its decay is examined after the addition of actinomycin D. In contrast, EGF greatly prolongs the half-life of EGF receptor mRNA suggesting a possible mechanism for the stimulatory effect of EGF on EGF receptor mRNA levels. EGF also stabilizes beta-tubulin and beta-actin mRNAs but has very little effect on the degradation of total mRNA.

Actins↗
PubMed1988

Epidermal growth factor receptor gene promoter. Deletion analysis and identification of nuclear protein binding sites.

To determine the location of sites that may be important for the function of the promoter of the epidermal growth factor (EGF) receptor gene and to characterize the factors that bind to these sites, the promoter region was analyzed by deletion analysis, exonuclease III protection and gel retardation assays with crude and fractionated nuclear extracts and DNase I footprinting using purified Sp1. Transfection of chimeric chloramphenicol acetyltransferase plasmids containing various deletions of the EGF receptor gene promoter into CV-1 cells indicated that the region between -178 and -16 (initiator ATG is +1) is sufficient for promoter activity. Exonuclease III protection assays revealed the presence of eight specific nuclear protein binding sites in the region between -481 and -16. Gel retardation assays confirmed that multiple protein binding sites exist in this region (-481 to -16) and quantitatively agree with exonuclease III protection. DNase I footprinting using purified Sp1 showed that this transcription factor can bind to four sites (-457 to -440, -365 to -286, -214 to -200, and -110 to -84) in the EGF receptor gene promoter and therefore may play a role in its regulation.

Animals↗
PubMed1988

Modulation of epidermal growth factor receptor proto-oncogene transcription by a promoter site sensitive to S1 nuclease.

The epidermal growth factor (EGF) receptor is the functional target of the mitogen EGF and the cellular homolog of the avian erythroblastosis virus erbB oncogene product. Regulation of expression of the proto-oncogene encoding the EGF receptor can be elucidated by studying the structure and function of the gene promoter outside the confines of the cell. Previously, we reported the isolation of the human EGF receptor gene promoter. The promoter is highly GC rich, contains no TATA or CAAT box, and has multiple transcription start sites. An S1 nuclease-sensitive site has now been found 80 to 110 base pairs (bp) upstream from the major in vivo transcription initiation site. Two sets of direct repeat sequences were found in this area; both conform to the motif TCCTCCTCC. When deletion mutations were made in this region of the promoter by using either Bal 31 exonuclease or S1 nuclease, we found that in vivo activity dropped three- to fivefold, on the basis of transient-transfection analysis. Examination of nuclear protein binding to normal and mutated promoter DNAs by gel retardation analysis and DNase I footprinting revealed that two specific factors bind to the direct repeat region but cannot bind to the S1 nuclease-mutated promoter. One of the specific factors is the transcription factor Sp1. The results suggest that these nuclear trans-acting factors interact with the S1 nuclease-sensitive region of the EGF receptor gene promoter and either directly or indirectly stimulate transcription.

Animals↗
PubMed1988

Topical application of cyclosporine on guinea pig allergic contact dermatitis.

Topical cyclosporine applied to the test site substantially inhibited the elicitation reaction of contact sensitivity in the guinea pigs previously sensitized with 2,4-dinitrochlorobenzene (DNCB). This suppressive effect of the drug was short lived and reversible. Cyclosporine was not effective when given six hours or later after antigenic challenge to the test site. Cyclosporine had no effect on the toxic contact reaction in normal animals either to croton oil or to DNCB in high concentration. Cyclosporine applied topically to the challenge site also resulted in a reduction of retest and flare-up reactions of contact sensitivity to DNCB, but did not affect the production of generalized rash in the same animals. These results indicate that in the future local topical application of cyclosporine may make treatment of human cutaneous immune-mediated disorders a possibility without serious side effects.

Administration, Topical↗
PubMed1987

Histological study on the fate of haptenated epidermal cells injected intradermally in guinea pigs.

The percutaneous administration of in vitro haptenated epidermal cells (EC) has become established as a procedure to produce contact sensitivity (CS) in experimental animals for routine use. The cells have also been found to elicit a significant delayed-type skin reaction by intradermal test in the animals sensitized by painting the skin with the hapten. The fate of 2,4-dinitrophenylated (DNP) isogeneic epidermal cell suspensions (EC) injected intradermally was investigated histologically in intact or 2,4-dinitrochlorobenzene (DNCB)-sensitized strain 13 guinea pigs to study the role of the cells in CS. DNP-EC were found to proliferate actively in the dermis and formed EC nests with central keratinization and then elicited inflammatory reaction associated with necrosis of the epidermal structures 7 days after injection in the intact animals. DNP-EC injected intradermally into the animals which had received and reacted against DNCB underwent a suppression of EC proliferation. These findings are discussed in relation to the role of the haptenated EC in CS.

Animals↗
PubMed1987

A histologic study on the fate of intradermally implanted epidermal cells in guinea pigs: a new method for evaluation of skin allograft survival.

The fate of allogeneic (strains 13, 2, and JY-1) and autologous epidermal cell (EC) suspensions injected intradermally was investigated histologically in JY-1 strain guinea pigs. Epidermal cells were found to proliferate actively in the dermis and form EC nests with central keratinization. The significant reject reaction associated with necrosis of the epidermal structures was seen in due time in the animals implanted with allogeneic ECs. We attempted to assess the effect of cyclosporin A (CYA) on skin allograft survival by observing the fate of strain 13 ECs implanted intradermally into the CYA-treated JY-1. Successful prolongation of allograft survival with CYA was clearly demonstrated by this method. This is considered to be a useful experimental way for evaluation of skin allograft survival and to be suitable for routine use.

Animals↗
PubMed1986

Amplification of human argininosuccinate synthetase pseudogenes.

The human genome contains multiple pseudogenes for an argininosuccinate synthetase (AS) gene. To elucidate the molecular mechanisms of generation and dispersion, complete nucleotide sequences of four different AS pseudogenes, psi AS-Y, psi AS-A1, psi AS-A2 and psi AS-A3, have been determined. A comparison of these sequences with those of three reported AS pseudogenes, psi AS-1, psi AS-3 and psi AS-7 revealed that two pairs, psi AS-Y/psi AS-7 and psi AS-A3/psi AS-1, are highly homologous but not identical, thereby suggesting that one of the pairs is generated by a duplication of the other member of the pairs. The psi AS-Y, which is probably located on chromosome Y, and the partially sequenced psi AS-7 are both interrupted by an Alu element at exactly the same site in their 3'-end regions. These two Alu elements are located in an opposite orientation relative to the direction of transcription of the pseudogene, and their possible role on pseudogene dispersion was examined. The psi AS-A1 is also accompanied by an Alu element at its 3' end. In this case, the orientation of the Alu element is the same as that of the pseudogene. The psi AS-A1 and the Alu element are flanked with direct repeats, as if they had been inserted into a chromosomal site, as a single unit.

Argininosuccinate Synthase↗
PubMed1986

Messenger RNA coding for argininosuccinate synthetase in citrullinemia.

Messenger RNA coding for argininosuccinate synthetase (ASS), extracted from the livers of some patients with citrullinemia, was analyzed using a cell-free translation system and dot and Northern blot hybridization with cDNA probe for ASS. In patients with quantitative-type citrullinemia, called type II here, previous studies have demonstrated that the hepatic content of the enzyme was about 10% of the control value, whereas the translatable mRNA level for the enzyme was similar to that of control livers. Here, we confirmed that the type II liver contained an almost normal amount of mRNA coding for ASS, judged by the dot-blot hybridization technique with cDNA. Northern blot hybridization of RNA indicated that there was hybridizable mRNA of approximately normal size (about 1.7 kilobase [kb]) in each, suggesting that large structural gene deletions had not occurred. These results indicate that in type II citrullinemia, the decrease in the enzyme protein is due either to increased degradation of the enzyme or to decreased or inhibited translation in the liver. Another type of citrullinemia was found and classified as type III. It is characterized by no detectable enzyme activity for ASS or translation activity for ASS mRNA. However, a smaller amount of RNA molecule hybridized for ASS cDNA was detected.

Adult↗