The arcB gene of Escherichia coli encodes a sensor-regulator protein for anaerobic repression of the arc modulon

The arcA (dye) and arcB genes of Escherichia coli are responsible for anaerobic repression of target operons and regulons of aerobic function (the arc modulon). The amino acid sequence of ArcA (Dye) indicated that it is the regulator protein of a two-component control system. Here we show that ArcB is a membrane sensor protein on the basis of its deduced amino acid sequence (778 residues), hydropathicity profile, and cellular distribution. On the carboxyl end of the ArcB sequence there is an additional domain showing homology with conserved regions of regulator proteins. Deletion into this domain destroyed ArcB function. ArcB conserved a histidine residue for autophosphorylation of the sensor proteins, and aspartic residues important for the regulator proteins.     

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.     

Modification of regression of virally xenogenized tumor cells by cyclophosphamide and busulfan

Summary Rat fibrosarcoma cells infected with Friend leukemia virus (FV-KMT-17) grow for a short time and then regress spontaneously in syngeneic hosts. This regression mechanism was examined by analyzing the immunomodulating action of the antitumor drugs busulfan (BU) and cyclophosphamide (CY). In preliminary experiments, the optimum dosages of BU and CY for the enhancement of DTH responses to SRBC were 10 mg/kg and 40 mg/kg respectively. Treatment of rats with BU (10 mg/kg) on day 5 induced the regression of KMT-17 cells, while in contrast, the same drug delayed the spontaneous regression of FV-KMT-17 cells. Pretreatment with CY (40 mg/kg) on day 5 did not affect the growth of KMT-17 or FV-KMT-17 cells. After the same treatment schedule, BU inhibited humoral antibody formation against SRBC and against virus-associated antigen (VAA), NK cell activity, and ADCC effector cell activity. On the other hand, CY did not affect the activities of NK cells or ADCC effector cells, although it significantly augmented the DTH responses to SRBC and the production of antibody to VAA but had no effect on production of antibodies to SRBC. These results suggest that NK cells and ADCC may play an important role in the initial stage of the spontaneous regression of FV-KMT-17 cells.Supported in part by a Grant-in-Aid for Cancer Research from the Japanese Ministry of Education Abbreviations used: BU, busulfan; CY, cyclophosphamide; PFC assay, plaque forming cell assay; VAA, virus-associated antigen; NK cell, natural killer cell; ADCC, antibody dependent cellular cytotoxicity; MuLV, murine leukemia virus; DTH, delayed type hypersensitivity; SRBC, sheep red blood cells; C.I., cytotoxic index; CRBC, chicken red blood cells; IL-1, interleukin 1; IL-2, interleukin 2; IFN, interferon     

Isolation of a cAMP-requiring mutant in Escherichia coli K-12: evidence of growth regulation via N-acetylglucosamine metabolism controlled by cAMP

Abstract An adenylate cyclase gene ( cya ) mutant was mutagenized and an adenosine 3,5-cyclic monophosphate (cAMP)-requiring mutant (KM8161) was obtained on Davis minimal medium containing glucose in the presence or absence of cAMP. KM8161 also required N -acetylglucosamine for its growth instead of cAMP. Furthermore, the mutant could use neither glucosamine nor N -acetylglucosamine as the carbon source. These results indicate that the cAMP-requiring property is due to multiple mutations of a few genes involved in amino sugar metabolism in addition to cya . By genetic analysis of KM8161, one gene, which was tentatively termed cidA and located near 2 min on the chromosomal map, proved to be defective. Reversion of cidA mutation in KM8161 resulted in recovery of not only the cAMP-requiring phenotype but also non-utilization of amino sugars. When both cAMP and N -acetylglucosamine or glucosamine were added to the culture medium for KM8161, only N -acetylglucosamine could be utilized as the carbon source. These studies s strongly suggest that the cidA or cya mutation in KM8161 causes deficiency in different stages of amino sugar metabolism and the regulatory circuit of growth by cAMP is mediated via control of N -acetylglucosamine metabolism.     

Surface Charge Density Estimation of Vigna mungo Protoplasts Using a Fluorescent Dye, 9-Aminoacridine

We showed that the surface charge density of protoplasts canbe estimated by the 9-aminoacridine method. The estimated surfacecharge density of the protoplasts isolated from elongating regionsof Vigna mungo root was – 39 ? 8 mC/m². The negative surfacecharge density increased when protoplasts were treated withglutaraldehyde or when EDTA was added to the protoplast suspensionmedium. These results support the validity of our estimationof the surface charge density of protoplasts by the 9-aminoacridinemethod. The concentration of amino groups at the surface ofthe protoplasts was estimated to be 34 mC/m². (Received June 19, 1987; Accepted April 11, 1988)     

A Comparative Study of the Structures of {alpha}-Glucans from Suspension-Cultured Nonglutinous and Glutinous Rice Cells

-Glucans (average mol wt, 1.3 ? 10⁴) extracted with perchloricacid from 8-day-old suspension-cultured nonglutinous (var. Sasanishiki)and glutinous rice (var. Miyakogane) cells were compared. Theresults of hydrolysis by alpha;-, ß- and iso-amylasesand methylation analysis of the -glucans suggested that theirbasic structures are almost the same. These -glucans are highly-branchedpolysaccharides with an average chain length of about 9–10,with exterior and interior chain lengths of about 6–7and 2–3, respectively. ¹Current address: Laboratory of Food Science, Faculty of Education,Hirosaki University, Hirosaki, Aomori 036, Japan. (Received April 27, 1987; Accepted March 2, 1988)     

Application of 39K NMR Spectroscopy to Potassium Transport in Mung Bean Root Tips

The intracellular K⁺ concentration and its change in mung bean[Vigna mungo (L.) Hepper] root tips were investigated non-invasivelywith ³⁹K nuclear magnetic resonance spectroscopy using a membraneimpermeable shift reagent, dysprosium (III) tripolyphosphate[Dy(PPP_i)^7–₂]. The K⁺ resonance was shifted to highermagnetic field in proportion to the concentration of the shiftreagent. In addition to a reference capillary peak for measuringthe K⁺ concentration, two well-resolved peaks (intra- and extracellularK⁺ resonances) were observed in the 39K NMR spectra of mungbean root tips. The intracellular K⁺ concentration was determinedto be 41 mM, which was similar to the value obtained by flamephotometry. When 20 mM KCl was added to the external medium,the intensity of the intracellular K⁺ resonance gradually increasedand the net K⁺ uptake rate was calculated to be 4.1 micromolesper gram fresh weight per hour. After removal of KCl from theperfusion medium, the intracellular K⁺ concentration considerablydecreased. With ³¹P NMR method, 2.5 mM Dy(PPP_j)^7–1₂ and20 mM KCl had little effect on the ATP level in the cells. Wehave indicated that the ³⁹K NMR method can be used to determinethe K⁺ levels and net fluxes of the K⁺ transport in perfusedroot tips successively. (Received April 6, 1988; Accepted September 29, 1988)     

Formation of UDP-Xylose and Xyloglucan in Soybean Golgi Membranes

Soybean (Glycine max) membranes co-equilibrating with Golgi vesicles in linear sucrose gradients contained UDP-glucuronate carboxy-lyase and xyloglucan synthase activities. Digitonin solubilized and increased the activity of the membrane-bound UDP-glucuronate carboxy-lyase. UDP-xylose did not inhibit the transport of UDP-glucuronate into the lumen of Golgi vesicles but repressed the decarboxylation of the translocated UDP-glucuronate. The results suggest that UDP-glucuronate is transported into the vesicles by a specific carrier and decarboxylated to UDP-xylose within the lumen. On incubation of UDP-[¹⁴C]glucuronate with Golgi membranes in the presence of UDP-glucose, [¹⁴C]xylose-labeled xyloglucan was formed. Although the K_m value of UDP-glucuronate for the decarboxylation was 240 micromolar, the affinity of UDP-glucuronate for xyloglucan formation (31 micromolar) was similar to that of UDP-xylose (28 micromolar), suggesting a high turnover of UDP-xylose. The biosynthesis of UDP-xylose from UDP-glucuronate probably occurs in Golgi membranes, where xyloglucan subsequently forms from UDP-xylose and UDP-glucose.