Mitogen-induced B-cell differentiation in Xenopus laevis

Abstract. Four genes are known to affect pigmentation in the Mexican axolotl. The purpose of this article is to review previous information pertinent to these genes and to reevaluate such information in light of new evidence that demonstrates (in a preliminary way) how pigments, and subsequently phenotypes, are affected by the various pigment genes. Each of the mutant phenotypes – m (melanoid), ax (axanthic), a (albino), and d (white) - is compared to the wild type (D). All of these genes are recessives, all of them affect phenotypic changes during development, and three of the four ( m, a , and d ) also affect specific biochemical (i.e., pigment) changes during development. In the axolotl, color patterns can be directly correlated to the presence(or absence) of a variety of pigments that are normally found in discrete pigment cells. Qualitative and quantitative analyses of the bright-colored pigments (pteridines and flavins in this case) present in axolotl skin demonstrate that these pigments vary significantly among the various phenotypes under consideration. These analyses raise some interesting questions with regard to how each of the pigment genes is believed to act, and numerous possibilities for continued experimentation are suggested.     

Identification and characterization of a brilliant yellow pigment produced by Bordetella pertussis

Culture supernatants of Bordetella pertussis are a brilliant yellow; however, the structure and biological role of the responsible pigment have not been investigated. In this study, a brilliant yellow‐colored fraction was extracted from culture supernatants of B. pertussis and analyzed by HPLC. UV–visible spectral analysis and mass spectrometry identified the brilliant yellow pigment as riboflavin. Riboflavin production was high in lag and early log phases and riboflavin was found to enhance growth of B. pertussis in low‐density cultures. Riboflavin production is not regulated by the BvgAS system. In addition, it was found that other Bordetella species, such as B. parapertussis , B. holmesii and B. bronchiseptica, also release riboflavin into their culture supernatants. This is the first report that B. pertussis secrets riboflavin to the extracellular space and that riboflavin may promote its growth. The mechanism may be associated with pathogenesis of B. pertussis .

     

Genetic and developmental characteristics of a new color variant, axanthic, in the Mexican axolotl, Ambystoma mexicanum Shaw

A new genetic trait, axanthic, is described in the Mexican axolotl with a phenotype completely lacking visible xanthophores and iridophores. Animals received from the colony of Dr. L. E. DeLanney at Ithaca, New York, have been mated among themselves for a total of seven spawnings, in which the axanthic trait was inherited as a simple Mendelian recessive. The symbol for this newly found gene isax. Homozygous recessive animals appear normal except for their lack of visible xanthophores and iridophores. The results of the spawnings also indicate that the axanthic gene is not linked to the melanoid gene (m). Its linkage with other known mutants in the axolotl is being tested.     

Pigment cell differentiation: the relationship between pterin content, allopurinol treatment, and the melanoid gene in axolotls

The effects of allopurinol (an inhibitor of the enzyme xanthine dehydrogenase (XDH] and the melanoid gene on pigment cell differentiation in the axolotl were examined by analyzing pigment components of the xanthophore (pterins). Pterin contents of skin extracts (70% ethanol) from wild type, allopurinol-treated and melanoid axolotls were determined by thin layer chromatography (TLC) and fluorometric scanning of TLC plates. Heights of peaks produced were used as a quantitative measure for pterin content. Results reveal that melanoid animals contain significantly reduced amounts of all seven pterins examined as compared with wild type animals. Allopurinol-treated animals have reduced levels of four pterins (xanthopterin, isoxanthopterin, biopterin and sepiapterin) as compared with the wild type. These findings suggest that the alterations in pterin biosynthetic pathways, either by drug-induced inhibition of XDH activity or by the melanoid gene, produce similar dramatic changes in pigment phenotype which are manifested by alterations in pigment cell differentiation.     

Patterns of purine synthesis related to iridophore development in the wild type, melanoid, and axanthic strains of the Mexican axolotl, Ambystoma mexicanum Shaw

Differentiation of the iridophore in the axolotl is inhibited by each of two nonlinked autosomal recessive genes, melanoid (m) and axanthic (ax). Purines were extracted from wild type (M-Ax-), melanoid (mm), axanthic (axax), and melanoid-axanthic (mm axax) somatopleur. This tissue is rich in iridophores in wild-type axolotl larvae. Purines in ethanol and in water extracts from each genotype were separated by paper chromatography. Patterns with respect to the kinds of purines present were distinct for each genotype. Two compounds present in wild type were missing in both melanoid and axanthic mutants, which lack iridophores. One compound was present in wild type and melanoid, which have xanthophores, but was lacking in the axanthic mutant, which does not. The double homozygote (mm axax) lacked a purine found in wild type and each of the mutants singly. Possibly in this case two biosynthetic pathways may produce the same intermediate product. Thus the actions of the melanoid and axanthic genes in the axolotl are demonstrable chemically as differences in the kinds of purines present in the various genotypes.     

Production of Riboflavin by Arthrobacter globiformis

S ummary . The yellow pigment excreted by a strain of Arthrobacter globiformis during exponential growth was identified as riboflav in. The amount produced was approximately 5 μ/mg of cells formed (dry weight) in all media tested, rich as well as poor. This value was also found in media containing 1% of Tween So, even though the concentration of internal flavins then was nearly one-half the normal value. Over production of riboflavin in most media was 40-50 fold. The specific growth rate of the riboflavin producer was sevral per cent less than that of a normal strain of the same species.     

Observations on the pigmentation of the pigeon iris

There are three genetically controlled iris types found in the pigeon, two of which contain stromal pigment cells, the third lacks pigment cells. The yellow (gravel) and white (pearl) iris types have pigment cells that contain birefringent pigment granules (crystals) and are ultrastructurally similar to iridophores of poikilothermic vertebrates. Both these iris types contain guanine as a major "pigment" and, in addition, the yellow iris contains at least two yellow fluorescing pigments that are tentatively identified as pteridines. The pigment cells of the yellow and white irises are structurally identical differing only in the presence or absence of these yellow pigments. The stromal pigment cells of the white iris correspond in structure and pigment chemistry to classical iridophores although they lack strong irridescence and are therefore perhaps best considered leucophores. The pigment cells of the yellow iris can be considered "reflecting xanthophores" having the combined properties of both classical xanthophores and iridophore/leucophores.     

Riboflavin, overproduced during sporulation of Ashbya gossypii, protects its hyaline spores against ultraviolet light

Riboflavin (vitamin B₂), essential in tiny amounts as a precursor for oxidoreductase coenzymes, is a yellow pigment. Although it causes cytotoxicity via photoinduced damage of macromolecules, several microorganisms are striking overproducers. A question, unanswered for decades, is whether riboflavin overproducers can benefit from this property. Here, we report an ultraviolet (UV) protective effect of riboflavin. The spores of Ashbya gossypii , a riboflavin-overproducing fungus, are more sensitive to UV than those of Aspergillus nidulans . The addition of riboflavin to suspensions improves the UV resistance of both spore types. Interestingly, we show that regulation of sporulation and riboflavin overproduction in A. gossypii are linked. In batch culture, both were elevated when growth ceased. At constant growth rates, obtained in a chemostat culture, neither was elevated. Supplementation of cultures by cAMP, a known stress signal, negatively affected sporulation as well as riboflavin overproduction, establishing a second, independent argument for the linkage.     

不产氧光合细菌光合色素的光氧调控机制

[目的]为不产氧光合细菌光合色素研究提供可行的较系统规范的研究方法和数据,揭示固氮红细菌(Rhodobacter azotoformans 134K20)光合色素光氧适应性机制.[方法]采用光谱法和色谱法对光和氧调控下的类胡萝卜素和细菌叶绿素合成代谢进行了研究.[结果]134K20菌株光照好氧时细胞得率最高.光照厌氧时主要合成3黄、1红、1紫、2绿、2蓝9种色素,黄色素大量表达.有氧时红色素大量表达,且启动2种新的红色素和1种新的紫色素表达,而黄色和蓝绿色素则受氧抑制.黑暗好氧主要合成2黄、3红、2紫、1绿、1蓝9种色素,但不同于光照厌氧.光照好氧时黄色素减少到1种,紫色素含量增加,其余同黑暗好氧.[结论]固氮红细菌(Rhodobacter azotoformans 134K20)是通过PpsR调节途径来调节光合基因表达的.黄色和红色素属于类胡萝卜素.黄色素1属于球形烯系列,其余两种黄色素是新的类胡萝卜素组分.红色素为新的球形烯酮组分,3种红色素极性、峰形和峰位差别显著,正己烷能显示其精细结构.紫色为极性较大的细菌脱镁叶绿素,绿色和蓝色为4种极性不同的细菌叶绿素a中间产物.乙醚甲醇法适合类胡萝卜素的提取,丙酮甲醇冰冻研磨法能快速有效完全提取光合色素.溶剂效应可有效鉴别细菌叶绿素a中间产物.     

Riboflavin Biosynthesis by Candida robusta

During the study on the oxidative sugar metabolism of yeasts, it was found that six strains of Candida robusta, isolated from fruits, produced large amounts of yellow pigment in shaking culture and this pigment was identified as riboflavin. Riboflavin production by C. robusta has never been reported.

Some notable characteristics of C. robusta in riboflavin production were found. As nitrogen sources, ammonium salts and urea were favorable, but nitrate and organic nitrogen sources such as glycine, asparagine and peptone were not utilized for riboflavin production. Riboflavin was not produced in still culture; a highly aerobic condition, as may be obtained by shaking culture, wa, essential. The addition of excess CaCO₃ was also necessary. Acetic acid, added as the Ca salt in its production as a sole carbon source, was more effective than sugars and optimum concentration of this acid was 7%. Riboflavin were obtained in yields as high as 32 to 34 mg % from the acetate medium after 8 days.     

Differential riboflavin deposition in white and variegated white mutants of Drosophila hydei

Riboflavin deposition in organs of Drosophila hydei was studied by means of a growth test using a riboflavin-deficient strain of the fungus Aspergillus nidulans. In wild-type animals, riboflavin is deposited in Malpighian tubules (MT) and testes but not in adult eyes. Certain white (w) mutants do not contain riboflavin, whereas intermediately colored w mutants contain minor amounts of the substance. Riboflavin-containing MT cells contain special globules that can be fixed and stained with the redox dye phenazine-methosulphate. The number and size of these granules is related to growth effect and point to a role of the w locus in the intracellular deposition of riboflavin in special organs. In white-mottled (wm) position-effect variegation mutants, a significant correlation was found between the extent of variegation (percentage of yellow cells) and riboflavin content (growth effect) of the MT. However, the individual variation of cell phenotype was extremely large and exaggerated types were observed indicating "overdominance" of the rearranged w+ gene. This contradicts an unsubstantiated dogma of position-effect variegation that assumes that the affected gene simply switches between the on and off state, as is discussed.     

A developmental stage of hyphal cells shows riboflavin overproduction instead of sporulation in Ashbya gossypii

The hemiascomycete Ashbya gossypii develops a mycelium. Nutritional stress leads to its differentiation into sporangia. These generate spores. In parallel, the yellow pigment riboflavin is produced. Intracellularly accumulated riboflavin, made visible as a bright green fluorescence, was observed in only 60 % of the hyphal cells. For the remaining 40 %, it was unclear whether these cells simply export riboflavin or its biosynthesis remains down-regulated in contrast to the accumulating cells. The approach followed in this work was to convert the hyphae into protoplasts by enzymatic degradation of the cell wall. Afterwards, the protoplasts were sorted by fluorescence-activated cell sorting on the basis of riboflavin accumulation. When a reporter strain expressing lacZ under the control of the most important riboflavin biosynthesis promoter, RIB3, was used, green protoplasts were found to have more than tenfold greater reporter activity than hyaline protoplasts. This was true on the basis of total protein as well as on the basis of hexokinase specific activity, a marker for constitutive expression. These results allow the conclusion that hyphal cells of A. gossypii differ in phenotype regarding riboflavin overproduction and accumulation.     

A light-sensitive yellow pigment from the housefly

Extraction of house-fly heads with neutral phosphate buffer yielded a dark brown solution from which a number of pigments were separated, either wholly or partially, by chromatography on a column of calcium phosphate mixed with celite. One of the pigments was light-sensitive, and had a yellow color, with a spectral absorption maximum at 437 mmicro in phosphate buffer at pH 6.5. Several consecutively eluted fractions from each chromatogram of the house-fly head extract showed the characteristic absorption curve of this pigment with no trace, spectroscopically, of the other pigments of the extract. The products of bleaching the pigment at pH 6.5 had an absorption curve showing plateaus at 440 to 460 mmicro and 350 to 360 mmicro and an inflection at about 250 mmicro. Above pH 8.0 the pigment bleached in the dark giving an absorption maximum at about 380 mmicro, and inflections at 290 mmicro and at about 250 mmicro. With 2.5 to 5 N HCl or H(2)SO(4) an absorption maximum at 470 to 475 mmicro was obtained with either the unbleached or the bleached pigment. With sulfosalicylic acid, ethanol, or heating at 100 degrees C., a part of the pigment was precipitated, leaving a light-stable yellow supernatant. This light-sensitive house-fly pigment cannot as yet be identified with any of the previously known insect pigments or with the photosensitive pigments of other animals, though these latter compounds exhibit some similarity in their spectroscopic properties.     

Morphological and Biochemical Characterization of the Cream Markings in the Integument of the Female Isopod,Armadillidium vulgare

Cream markings aligned along the dorsal region of the female isopod, A. vulgare, were investigated with light and a fluorescence microscope and an electron microscope. Biochemical studies were also carried out. The cream markings were observed in the dorsal integument as a group of cream-colored chromatophores that emit a yellow fluorescence. These chromatophores, which are distinguishable from ommochrome chromatophores, contained numerous granules in the cytoplasm, and these granules (0.6–3.0 μm in length by 0.4–1.5 μm in width) were electron-lucent and spheroidal in shape with a concentric arrangement of membranes. Based on various biochemical analyses, the principal component of the yellow pigment isolated from the cream markings was identified as sepiapterin. These facts revealed that the cream markings are the chromatophores that contain pteridine granules. The males have no cream markings like those of the females, since the cream-colored chromatophores are externally hidden by the ommochrome chromatophore layer. The content of sepiapterin in the males was about two times greater than that in the females. This quantitative difference in sepiapterin content between males and females suggests that the pteridine formation in this pigment cell may be regulated by hormones associated with sex determination.     

DISTRIBUTION OF MATURATION-PROMOTING FACTOR IN STARFISH OOCYTE STRATIFIED BY CENTRIFUGATION

In starfish, cytoplasm taken from maturing oocytes is capable of inducing breakdown of the germinal vesicle and subsequent maturation when injected into immature oocytes. The cytoplasmic factor has been designated as "maturation-promoting factor" (MPF). Ooplasm was stratified by centrifugation of maturing oocytes in density-graded Ficoll-seawater, without disrupting the cell. Three strata were distinguished beginning with the centripetal side: oil cap, hyaline layer and yellow layer. MPF activity was shown to be localized in the hyaline layer. Electron microscopic observation revealed that the hyaline layer contains Golgi complexes, many lucent vesicles and multi-vesicular bodies as distinct organelles, but seldom contains such inclusions as the lipid droplets forming the oil cap, mitochondria, yolk and pigment granules contained in the yellow layer. Based on these observations, a possible cytoplasmic component with MPF activity is discussed.     

Photoconversion of riboflavin to lumichrome in plant tissues

Free flavins have been extracted from shoots of etiolated corn (Zea mays L., var. Burpee Snowcross) and from yeast cells and separated from other substances by absorption on resorcinol-formaldehyde resin and talc columns and by thin layer chromatography. Riboflavin was the only free flavin present. Extracts of etiolated shoots of oats (Avena sativa L., var. Multiline E-69 and Clinford) yielded riboflavin plus a second free flavin previously demonstrated in oats. The areas of the chromatograms expected to contain lumichrome were completely clear. After illumination of any of the three organisms with artificial light (1100 ft-c) or sunlight for 6 hours, lumichrome (7,8-dimethylalloxazine) was found. In corn shoots after irradiation by sunlight, the amount of lumichrome present was equivalent to 2.5% of the total free flavin. Lumichrome was identified by thin layer chromatography in six solvent systems (including two two-dimensional systems), by its characteristic fluorescence in acetic acid, by its absorption spectrum, and by formation of a characteristic hydrate in ammonia-containing solutions. A comparison was made with in vitro photolysis of riboflavin and the possible role of photolysis of riboflavin (either free or bound) and of lumichrome formation in photo-responses of plants is discussed. Placing the shoots in the dark for 4 hours after irradiation in sunlight for 6 hours led to no detectable loss of the lumichrome which had been formed.     

Purification and molecular and biological characterisation of the 1-hydroxyphenazine,produced by an environmental strain of Pseudomonas aeruginosa

Pseudomonas aeruginosa—an opportunistic pathogen, perhaps best known for chronic lung infections, produces wide range of pigments that possess specific activities which either assist the organism’s survival or bring about changes within host. A similar blue-green diffusible pigment producing P. aeruginosa was isolated from dug-well water, so as to extract 1-hydroxyphenazine from its crude pigment. The compound was purified from the crude pigment using column chromatography followed by a preparative thin layer chromatography that showed a single yellow spot. Further molecular characterisation of the purified component was carried out using UV–Vis spectrophotometer, Fourier transform infrared spectroscopy, nuclear magnetic resonance spectroscopy and mass spectroscopy which showed respective peaks corresponding to 1-hydroxyphenazine. Biological characterisation using in vitro assays revealed that 1-hydroxyphenazine showed anti-bacterial activity only against Bacillus sp. and a concentration of 30 µg/ml induced noticeable morphological alteration in A549 human lung adenocarcinoma cells followed by cell death after 48 h. Thus, such active components within bacterial pigments can be characterized and used as possible anti-bacterial or anti-cancer agents.     

Association of red coloration with senescence of sugar maple leaves in autumn

We evaluated the association of red coloration with senescence in sugar maple (Acer saccharum Marsh.) leaves by assessing differences in leaf retention strength and the progression of the abscission layer through the vascular bundle of green, yellow, and red leaves of 14 mature open-grown trees in October 2002. Computer image analysis confirmed visual categorization of leaves as predominantly green, yellow or red, and chemical quantification of leaf pigment concentrations verified that leaf color reflected underlying differences in leaf biochemistry. Significantly lower chlorophyll concentrations within red and yellow leaves indicated that senescence was more advanced in leaves from these color categories relative to green leaves. Among leaf types, only red leaves contained high concentrations of anthocyanins. There were significant differences in leaf retention capacity among color categories, with the petioles of green leaves being the most firmly attached to twigs, followed by red and then yellow leaves. Microscopic analysis indicated that yellow leaves had the most advanced extension of the abscission layer through the vasculature, with green and red leaves having significantly less abscission layer progression than yellow. A more limited progression of the abscission layer through vascular bundles may be evidence of delayed leaf senescence that could extend resorption of mobile leaf constituents. Together, results from this study suggest an association between leaf anthocyanin content and functional delays in senescence.     

一株产黄色素红曲霉Monascus HB-5的生物学特性及发酵条件研究

研究了一株产黄色素的红曲霉菌株Monascus HB-5的生物学特性,并对其接种量、pH、碳源、氮源等主要发酵因素进行优化。结果表明该菌产出高浓度单一黄色素,色调为棕黄色,仅在370nm具有吸收峰。产黄色素最适发酵条件为接种量10%,pH4.6～5.7,温度30～32℃,适合的碳源为玉米粉,氮源为硝酸铵,摇瓶黄色素色价达200 U/ml。并对色素的稳定性及安全性进行了初步研究,橘霉素含量低于0.1mg/l。     

Flavin binding to the high affinity riboflavin transporter RibU

The first biochemical and spectroscopic characterization of a purified membrane transporter for riboflavin (vitamin B(2)) is presented. The riboflavin transporter RibU from the bacterium Lactococcus lactis was overexpressed, solubilized, and purified. The purified transporter was bright yellow when the cells had been cultured in rich medium. We used a detergent-compatible matrix-assisted laser desorption ionization time-of-flight mass spectrometry method (Cadene, M., and Chait, B. T. (2000) Anal. Chem. 72, 5655-5658) to show that the source of the yellow color was riboflavin that had been co-purified with the transporter. The method appears generally applicable for substrate identification of purified membrane proteins. Substrate-free RibU was produced by expressing the protein in cells cultured in chemically defined medium. Riboflavin, FMN, and roseoflavin bound to RibU with high affinity and 1:1 stoichiometry (K(d) for riboflavin is 0.6 nM), but FAD did not bind to the transporter. The absorption spectrum of riboflavin changed dramatically when the substrate bound to RibU. Well resolved bands appeared at 441, 464, and 486 nm, indicating a hydrophobic binding pocket. The fluorescence of riboflavin was almost completely quenched upon binding to RibU, and also the tryptophan fluorescence of the transporter was quenched when flavins bound. The results indicate that riboflavin is stacked with one or more tryptophan residues in the binding pocket of RibU. Mutagenesis experiments showed that Trp-68 was involved directly in the riboflavin binding. The structural properties of the binding site and mechanistic consequences of the exceptionally high affinity of RibU for its substrate are discussed in relation to soluble riboflavin-binding proteins of known structure.