Studies on detection of Candida antigen in the sera of mice inoculated orally with Candida albicans

Summary The authors succeeded in establishing a murine model of systemic candidiasis being disseminated from the primary gastrointestinal lesions caused by oral inoculation of Candida albicans. Using this model, an attempt was made for detecting the Candida antigen by enzyme-linked immunosorbent assay using avidin-biotin (AB-ELISA) from the serum of infected mice.Gastrointestinal candidiasis was formed in all of the 20 mice treated with the drugs (antibiotics, antineoplastic agents, hydrocortisone, etc.) and inoculated orally with C. albicans. Fourteen of these mice suffered from submucosal candidiasis, and C. albicans was cultured from the visceral organs in 12 of them. The assay by AB-ELISA was able to detect 1.0 ng/ml Candida mannan in the mouse serum. The Candida antigen was detected in the sera of 11 of the 14 mice with submucosal candidiasis. However, the antigen could not be detected in the sera of the 6 mice with intramucosal candidiasis.The assay by AB-ELISA is more sensitive and specific for the diagnosis of systemic candidiasis than other serological assays.     

Fumarate reduction systems in members of the family Rhodospirillaceae with different quinone types

Nineteen established and one undesignated species of the Rhodospirillaceae were examined for fumarate reduction in connection with their quinone systems. The fumarate reductase activity with reduced methyl viologen (MVH) or FMNH₂ as electron donor was found in membrane (chromatophore) preparations from phototrophically grown cells of all species containing menaquinone (MK) and/or rhodoquinone. The species having ubiquinone as the sole quinone contained no fumarate reductase activity, except some Rhodobacter species showing the FMNH₂-dependent activity. The MVH-fumarate reductase activity of the MK-type species was not inhibited by Triton X-100 or acetone treatment, suggesting the presence of a fumarate reductase reacting directly with MVH, while such an enzyme was absent in the MK-lacking strains, with few exceptions. The FMNH₂-fumarate reduction system was abolished by a detergent or acetone extraction in all bacteria but differed much among species with different quinone types as to the response to respiratory inhibitors. These differences in fumarate-reducing properties and quinone systems among the phototrophic bacteria are discussed from evolutionary and taxonomic viewpoints.Non-standard abbreviations RQ rhodoquinone - MK menaquinone - MVH reduced methyl viologen - HOQNO 2-n-heptyl-4-hydroxyquinoline-N-oxide - TTFA 2-thenoyltrifluoroacetone     

A monoclonal antibody to the carbohydrate chain on human hepatocellular carcinoma-associated antigen which suppressed tumor growth in nude mice

Summary There have been few reports stating that monoclonal antibody alone inhibits human solid tumor growth in vivo. The present study demonstrated that monoclonal antibody S1 (IgG2a), which recognized the antigenic determinant of the carbohydrate moiety, showed antibody-dependent cell (or macrophage)-mediated cytotoxicity (ADCC or ADMC) in conjunction with murine splenocytes of both BALB/c and athymic mice. In vivo experiments demonstrated that the antibody S1 clearly prolonged the survival of athymic mice which had been inoculated with a human liver carcinoma cell line. In addition, the antibody S1 significantly suppressed the human hepatoma line transplanted s.c. into nude mice. ¹²⁵I-Labeled monoclonal antibody S1 revealed that the antibody accumulated significantly in the tumor mass. Many mononuclear cells were observed surrounding tumor cells when the antibody was given. This model system might be useful for analyzing the ADCC (or ADMC) mechanism in vivo.     

Cloning and sequencing of a ligninase gene from a lignindegrading basidiomycete,Phanerochaete chrysosporium

Summary A ligninase gene has been cloned from a Phanerochaete chrysosporium genomic DNA library. Nucleotide sequencing of the gene has revealed that the ligninase structural gene contains 1116 bp of the protein-encoding sequence, of which 84 bp encode the signal peptide. The protein-encoding sequence is interrupted by eight introns which conform to the universal G-T/A-G splicing rule observed for the 3 and 5 intron boundaries. The putative eukaryotic regulatory sequences, i.e. CAAT and TATA box-like sequences, are present in the 5 flanking region.     

Solubilization and Characterization of Calcitonin Gene-Related Peptide Binding Site from Porcine Spinal Cord

The binding site for calcitonin gene-related peptide (CGRP) was solubilized with 3-[(3-cholamidopropyl)dimethylammonio]-1-propane sulfonate (CHAPS) in an active form from porcine spinal cord. 125I-labeled human alpha-CGRP (125I-CGRP) binding to the solubilized protein was determined by filtration using a GF/B glass filter. The maximal binding activity (approximately 60% of the crude membrane fraction) was obtained with 5 mM CHAPS. 125I-CGRP binding to the solubilized protein was of high affinity, saturability, and high specificity, having KD and Bmax values of 3.69 pM and 338 fmol/mg of protein, respectively. The binding activity was eluted in a single peak with a molecular mass of 400,000 daltons by gel filtration on TSK gel G4000SW. These results suggest that the solubilized protein may be responsible for the specific binding site.     

Change in Gene Expression in Radish Cotyledons during Dark-Induced Senescence

Cotyledons detached from light-grown radish (Raphanus sativusL. cv. Comet) seedlings were used as a model system to studythe changes in nuclear gene expression during dark-induced senescenceof green leaves. Polyadenylated RNA was prepared from the cotyledonsat different times and then translated in a wheat germ system.Approximately 1,000 different polypeptides of the translationproducts were separated from each other by two-dimensional gelelectrophoresis. As judged from the density of autoradiographicspots of the translation products, the induction of senescenceby dark treatment involved an increase in 26 species, a decreasein 11 species, and a temporary increase and subsequent decreasein 8 species of translatable mRNA. A similar pattern of changein protein synthesis was also observed in the dark-treated cotyledonswhen the cotyledons were pulse-labeled with ³⁵S-methionine andthe soluble proteins separated by two-dimensional gel electrophoresis,though the polypeptide pattern on the gel did not coincide exactlywith those of the cell-free translation products. These findingsstrongly suggest that the process of leaf senescence is notsimply a passive and gradual death of the tissue, but involvesa drastic and sequential response of the cells to environmentalstimuli with respect to the gene expression of the cells. (Received July 21, 1987; Accepted September 30, 1987)     

ent-Kaurene Synthesis and Endogenous Levels of Gibberellins in a Shoot Forming Tobacco Crown Gall Tissue

The in vitro ent-Mcaurene synthesizing capacity, as well asthe endogenous GA content of shoot-forming tobacco crown gallsinduced by a nopaline-type Ti plasmid, was studied. For determinationof the ent-kaurene synthesizing capacity, an HPLC procedurepreceded by sample clean-up was used and the GA content wasexamined by GC-SIM. Kaurene synthesis reached a maximum at thebeginning of the logarithmic phase of growth. There was a clearcorrelation between the ent-kaurene synthesizing capacity andthe content of C₂₀-GAs. It seems that gibberellin synthesisis related to growth and development of the tissue. The natureof the GAs identified suggests, that the GA metabolism mightbe an unusual one. (Received October 12, 1987; Accepted April 11, 1988)     

Levels of IAA, Cytokinins, ABA and Ethylene in Rice Plants as Affected by a Gibberellin Biosynthesis Inhibitor, Uniconazole-P.

Effects of uniconazole-P, a triazole-type growth retardant,on endogenous levels of IAA, cytokinins, ABA and ethylene inrice seedlings were investigated. Endogenous levels of IAA andABA were similar between control and uniconazole-P-treated riceshoots. Evolution of ethylene was promoted slightly, being 1.8times greater under 0.3 ppm uniconazole-P treatment than thatof control. The most obvious effect was the increase of trans-Zand trans-RZ in shoots. Shoots treated with uniconazole-P (10mg/m² nursery box) contained 3.4 times and 3 times more trans-Zand trans-RZ than control, respectively. No significant differencesof cytokinin levels were recognized in roots except for cis-RZ.The increase of ethylene and active forms of cytokinins, andthe decrease of gibberellin in the shoots may be the basis forphysiological phenomena caused by uniconazole-P, namely thepromotion of flowering in woody plants and the enhancement offemaleness in cucumber. (Received September 9, 1987; Accepted October 20, 1987)     

Uptake of Gibberellin Methyl Esters by Spores and Induction of Dark Germination in Lygodium japonicum

In the fern Lygodium japonicum, the effect of the exogenousapplication of two gibberellin methyl esters, gibberellin A₄methyl ester (GA₄Me) and gibberellin A₂₀ methyl ester (GA₂₀Me)on spore germination in the dark and uptake of GA₄Me and GA₂₀Meby spores was investigated. Tritiated GA₄Me and GA₂₀Me wereprepared and used as radioactive tracers. The activity of GA₄Mewas more than 100-fold that of GA₂₀Me for the induction of sporegermination. When treated for 24 h, the activity for inducingspore germination remained after removal of the gibberellinmethyl esters from the medium. The amount of GA₄Me taken upby spores was more than three times that of GA₂₀Me throughoutthe 24 h time course of treatment. The uptake of both gibberellinmethyl esters was proportional to the external concentrationfor the range of concentrations between 10^–9 M and 10^–6M. When treated with the tritiated gibberellin methyl estersat 10^–6 M and 10^–7 M for 24 h, most of the gibberellinmethyl esters taken up by the spores were not metabolized. Althoughthe uptake of the two gibberellin methyl esters differed by3- to 5- fold, their abilities to induce spore germination differedby more than 100-fold. Therefore, the difference in the activityof the two gibberellin methyl esters regarding the inductionof spore germination could not be explained solely by the differencein their uptake. (Received January 11, 1988; Accepted May 26, 1988)