Flexibility on the karyotype evolution in bitterlings (Pisces, Cyprinidae)

In bitterlings (Acheilognathinae) C- and Ag-banding karyotypes of 6 species-subspecies collected in China and South Korea were analyzed. The chromosomal constitution of 2n=46 (4SM+42ST) in Rhodeus atremius fangi was quite different from that of 2n=48 (8M+20SM+20ST) in other species-subspecies in Rhodeus. It was concluded from the analysis of banded chromosomes that the increase in number of ST during the karyotype change from 2n=48 to 2n=46 was achieved by a series of pericentric inversions from 24 M-SM to 24 ST, and the decrease in the diploid number was caused by an additional tandem fusion of 4 ST chromosomes, forming a new ST pair in the 2n=46 karyotype. The karyotype of Tanakia koreensis, T. signifer, and Acheilognathus macropterus is 2n=48 (8M+20SM+20ST), 2n=48 (8M+20SM+14–16ST+4–6 A), 2n=44 (14M+16SM+14ST), respectively. In R. ocellatus ocellatus, T. koreensis, T. signifer and A. macropterus, karyotype changes from 2n=48 to 2n=44 due to centric fusion and inversion have also been estimated. It was suggested that C-banding heterochromatin was greatly concerned with the karyotype evolution in bitterlings.     

Substitutions of the Conserved Thr42 Increased the Roles of the ε-Subunit of Maize CF1 as CF1 Inhibitor and Proton Gate

The conserved residue Thr42 of -subunit of the chloroplast ATP synthase of maize (Zea mays L.) was substituted with Cys, Arg, and Ile, respectively, through site-directed mutagenesis. The over-expressed and refolded -proteins were purified by chromatography on DEAE-cellulose and FPLC on mono-Q column, which were as biologically active (inhibiting Ca²⁺-ATPase activity and blocking proton gate) as the native subunit isolated from chloroplasts. The T42C and T42R showed higher inhibitory activities on the soluble CF₁(–) Ca²⁺-ATPase than the WT. The T42I inhibited the Ca²⁺-ATPase activity of soluble CF₁ and restored photophosphorylation activity of membrane-bound CF₁ deficient in the most efficiently. Far-ultraviolet CD spectra showed that the portions of -helix and -sheet structures of the three mutants were somewhat different from WT. Thus the conserved residue Thr42 may be important for maintaining the structure and function of the -subunit and the basic functions of the -subunit as far as an inhibitor of Ca²⁺-ATPase and the proton gate are related.     

Effect of aldosterone on incorporation of amino acids into renal medullary proteins

Summary Studies on the effects of pretreatment with aldosterone on the incorporation of³H leucine or³H methionine into proteins in renal slices were carried out in Joklik-modified minimal essential medium. Administration of aldosterone (2 g/100 g body wt) to adrenalectomized rats increased³H leucine incorporation into trichloroacetic acid insoluble fractions of crude homogenates of cortical slices by 15.5±0.4% and of medullary slices by 53.5±1.3%. No increase in isotope incorporation was observed in slices of renal papilla or spleen prepared from the same rats. Aldosterone had no effect on the³H-leucine content of the trichloroacetic acid-soluble fractions of all three renal zones and the spleen. The dose of aldosterone that elicited a half-maximal increase in³H-methionine incorporation into proteins of renal medullary slices (0.45 g of aldosterone/100 g body wt) was indistinguishable from that needed to elicit a halfmaximal increase in the urinary K⁺/Na⁺ ratio (0.35 g of aldosterone/100 g body wt). Dexamethasone, a potent glucocorticoid, at a dose of 0.8 g/100 g body wt did not augment³H-leucine incorporation into renal medullary proteins but was effective at 8 g/100 g body wt. Spirolactone (SC-26304), a potent anti-mineralocorticoid, abolished the effect of aldosterone on amino acid incorporation into medullary proteins when administered at a 100-fold higher dosage [i.e., 80 gvs. 0.8 g (per 100 g body wt)]. These results imply that the action of aldosterone on amino acid incorporation is mediated by the mineralocorticoid rather than the glucocorticoid pathway, presumably the mineralocorticoid receptors. Moreover, pretreatment of the rats with actinomycin D (70–80 g/100 g body wt) erased the effect of aldosterone (0.8 g/100 g body wt) on amino acid incorporation into medullary proteins.In paired experiments with³H and³⁵S methionine, aldosterone (0.8 g/100 g body wt) increased methionine incorporation into trichloroacetic acid precipitable proteins of subcellular fractions of the renal medulla. The effect of aldosterone on incorporation of methionine into medullary cytosol proteins was analyzed further by polyacrylamide gel electrophoresis at pH 8.3 in tris-glycine buffer. The gel profiles indicate that aldosterone significantly increased methionine incorporation into at least one protein (independent of the isotope) with a molecular weight of 31,000. This increase was inhibited by either pretreatment of the rat with actinomycin D (70–80 g/100 g body wt or SC-26304 (80 g/100 g body wt). Dexamethasone (0.8 g/100 g body wt) did not increase incorporation of methionine into the medullary cytosol proteins resolved by polyacrylamide gel electrophoresis.     

Genetic Linkage Maps of the Guppy (Poecilia reticulata): Assignment of RAPD Markers to Multipoint Linkage Groups

Genetic linkage maps of the guppy (Poecilia reticulata) were constructed from independent crosses between the Tuxedo strain and a feral line (Wildtype). Segregation patterns of random amplified polymorphic DNA (RAPD) markers and phenotypic markers were investigated in F₂ offspring of Tuxedo × Wildtype and Wildtype × Tuxedo crosses. Among the 300 and 276 RAPD markers scored for the respective crosses, linkages were identified for 230 and 212, respectively. The Tuxedo × Wildtype and Wildtype × Tuxedo maps spanned 2100 Kosambi centiMorgans (cM_K) and 1900 cM_K, respectively, in 28 linkage groups. Average marker resolution was 10 cM_K. Genome length was estimated at 4410 cM_K and 4060 cM_K for the respective crosses, with an average physical distance of 166 kbp/cM_K. Several RAPD markers were closely linked to or mapped onto the loci for the sex-determining region (SdR), and the sex-linked black caudal-peduncle (Bcp) and red tail (Rdt) genes. These primary linkage maps are the initial step toward the construction of a composite high-density map to facilitate map-based cloning and marker-assisted selection of quantitative trait loci that are essential for the development of comprehensive breeding programs for the guppy.     

Expression in Bacillus subtilis of the gene for human urogastrone using synthetic ribosome binding sites

Summary A chemically synthesised gene coding for human urogastrone which was earlier cloned in E. coli (Smith et al. 1982) has now been cloned into expression vectors for Bacillus subtilis Two types of constructs have been made, one giving production of methionylurogastrone and the other giving rise to a methionyl-urogastrone- galactosidase fusion polypeptide facilitating quantification of expression levels.The ribosome binding sites used in the expression plasmids are synthetically made oligonucleotides residing on short restriction fragments to allow easy replacement by other ribosome binding sites.Using shuttle vectors and constitutive promoters from Bacillus phages 105 and SPP1, we were able to detect levels of expression amounting to a few thousand molecules per cell during logarithmic growth in both E. coli and B. subtilis.     

The effect of monoterpenes on astaxanthin production by a mutant ofPhaffia rhodozyma

Summary The total pigment and astaxanthin content ofPhaffia rhodozyma increased with increasing concentrations -pinene up to 500 l -pinene/l. Above this concentration the total pigment and astaxanthin content as well as the biomass production decreased. The addition of 500 l -pinene/l increased the total pigment content from 1652 g/g to 2201 g/g and the astaxanthin content from 1554 g/g to 1883 g/g. A sharp decrease in maximum specific growth rate occurred above 150 l -pinene/l.     

Cell wall localization of dihydroflavonol-glucoside β-glucosidase in flowers of Petunia hybrida

Limbs of flower buds from Petunia hybrida were investigated for -glucosidase activity with dihydroflavonol-glucosides and 4-methyl-umbelliferyl--D-glucoside as substrates. Dihydroflavonol-glucoside -glucosidase is localized in the cell wall. This activity has an acid pH optimum and is also active toward 4-methyl-umbelliferyl--glucoside. Besides this activity a neutral -glucosidase is present. This activity is soluble and is not active toward dihydroflavonol-glucosides. Using starch gel electrophoresis it was shown that no difference in -glucosidase activity is present between mutants able to convert dihydroflavonols into anthocyanins and mutants accumulating dihydroflavonol-glucosides. It is concluded that -glucosidase activity is not involved in anthocyanin synthesis.Abbreviations 4MU--glc 4-methylumbelliferyl--D-glucopyranoside - dHQ-7-g dihydroquercetin-7-glucoside - dHQ-4-g dihydroquercetin-4-glucoside - dHM-4-g dihydromyricetin-4-glucoside Deceased     

First survey on the natural occurrence of Fusarium mycotoxins in Bulgarian wheat

Wheat for human consumption (140 samples) was collected after harvest from all regions of Bulgaria. The 1995 crop year was characterized by heavy rainfall in the spring and summer months. The internal mycoflora of wheat samples was dominated by Fusarium spp. and Alternaria spp., and storage fungi were rarely present. The samples were analysed for contamination with Fusarium mycotoxins deoxynivalenol (DON), 3-acetyldeoxynivalenol (3-AcDON), 15-acetyldeoxynivalenol (15-AcDON), T-2 Toxin (T-2), diacetoxyscirpenol (DAS), and zearalenone (ZEA), using enzyme immunoassay methods. DON and ZEA were the predominant toxins, with a contamination frequency of 67% and 69%, respectively. The average levels of these toxins in positive samples were 180 g/kg (DON) and 17 g/kg (ZEA), maximum concentrations were 1800 g kg^–1 and 120 g kg^–1, respectively. Acetyl derivatives of DON, namely 3-AcDON and 15-AcDON, were found in 2.1 % and 0.7% of the samples, at at maximum level of about 100 g kg^–1. Only one sample was positive for T-2 (55 g/kg), DAS was not detected. This is the first report about the natural occurrence of a range of Fusarium mycotoxins in wheat for human consumption in Bulgaria.Abbreviations 3-AcDON 3-acetyldeoxynivalenol - 15-AcDON 15-acetyldeoxynivalenol - DAS diacetoxyscirpenol - DON deoxynivalenol - EIA enzyme immunoassay - T-2 T-2 toxin - ZEA zearalenone     

β-1-4-and β-1-3-glucan synthases are associated with the plasma membrane of the fungus Saprolegnia

Cytoplasmic membranes from mycelium or protoplasts of Saprolegnia monoica (a cellulosic cell-wall fungus) were separated by continuous sucrose-density-gradient centrifugation. Glucan synthases assayed at low (micromolar uridine 5-diphosphate (UDP) glucose for -1-4-glucan synthase) and high (millimolar UDP glucose for -1-3-glucan synthase) substrate concentrations were associated with membranes exhibiting vanadate-sensitive, oligomycin-insensitive ATPase and equilibrating at density 1.16 g cm^-3. Synthase activities were also bound to membranes of lower density (1.10 and 1.145 g cm^-3). Plasma membranes were stabilized by coating protoplasts with concanavalin A. After lysis of the protoplasts, plasma membranes recovered by low centrifugal forces were isolated in continuous isopycinic gradients. Both synthase activities peaked with [³H]concanavalin A and Na-vanadate ATPase indicating that the synthetases are located at the plasma membrane. Treatments of intact protoplasts with cold glutaraldehyde or proteases before disruption lead to a diminution of glucan-synthase activities indicating that at least part of the enzymes of plasma membrane face the outside of the cell.Abbreviations ConA concanavalin A - ER endoplasmic reticulum - GSI -1,4-glucan synthase - GSH -1,3-glucan synthase - UDP uridine 5-diphosphate     

Polymorphism of the HLA DR1 haplotype in the Israeli population investigated at the serological,cellular, and genomic levels

In the present report, we used serological, cellular, and restriction fragment length polymorphism (RFLP) to investigate the DR1 haplotype in the Israeli population. We describe an Israeli homozygous typing cell (HTC), HLA-DwLVA, which defines a new lymphocyte-activating determinant associated with Bw65, DR1 and distinct from Dwl. The parents of this donor, non-Ashkenazi Algerian Jews, are first cousins and share HLA-Cw8, Bw65, BfS, DR1, DQw1, DPw4. No specificity could be assigned to HLA-DwLVA using the 91 Ninth Workshop HTCs. Two families and forty unrelated DR1 individuals were studied with DwLVA and a panel of DR1/Dw1 HTCs. HLA-DwLVA showed segregation as a single determinant within families. This new specificity was present in 24 out of 40 (60%) unrelated DR1 individuals, indicating that in the Israeli population DwLVA is the main lymphocyte-defined determinant associated with the serologically defined DRI specificity, in contrast to non-Jewish Caucasoids where DR1 is significantly associated with Dw1. The vast majority of DwLVA-positive carriers were also Bw65 carriers, indicating that Bw65, DR1, DwLVA may represent a typical allele combination in the Israeli population. The RFLP analysis established the correlation of certain RFLPs with Dw1 and DwLVA. In addition, we describe a cluster of RFLPs that may correspond to a new Dw subtype associated with DR1, for which no serological and cellular reagents have been described so far.     

Studies of α- and βL-Crystallin Complex Formation in Solution at 60°C

Studies of molecular mechanisms of chaperone-like activity of -crystallin became an active field of research over last years. However, fine interactions between -crystallin and the damaged protein and their complex organization remain largely uncovered. Complexation between - and _L-crystallins was studied during thermal denaturation of _L-crystallin at 60°C using small-angle X-ray scattering (SAXS), light scattering, gel-permeation chromatography, and electrophoresis. A mixed solution of - and _L-crystallins at concentrations about 10 mg/ml incubated at 60°C was found to contain their soluble complexes with a mean radius of gyration 14 nm, mean molecular mass 4 MDa and maximal size over 40 nm. In pure _L-crystallin solution, no complexes were observed at 60°C. In SAXS studies, transitions in the -crystallin quaternary structure at 60°C were shown to occur and result in doubling of the molecular weight. This suggests that during the temperature-induced denaturation of _L-crystallin it binds with modified -crystallin or, alternatively, _L-crystallin complexation and -crystallin modifications are concurrent. Estimates of the -_L-crystallin complex size and relative contents of - and -_L-crystallins in the complex suggest that several -crystallin molecules are involved in complex formation.     

The mitochondrially encoded "phantom ATPase" of S. cerevisiae. II. A heterogeneous mitochondrial ATPase population in situ.

Summary The mitochondrial ATPase from a PHO 1 mutant (OLI 2, PHO 1, OLI 4 region on mit DNA of S. cerevisiae) was further examined. A new purification method using Lysolecithin instead of Triton allowed us to solubilize and separate a heterogeneous ATPase population from PHO 1-mitochondria: the major abnormal fraction had extremely low oligomycin-sensitivity (but normal specific immunological reactivity), while a minor normal fraction (representing about 20% of the initial mitochondrial ATPase activity) had high sensitivity and affinity for oligomycin.Moderate urea treatment of PHO 1-mitochondria leads to partial loss of ATPase activity and a concomitant increase of oligomycin-sensitivity, suggesting that a heterogeneous ATPase population exists in situ in the mitochondrial membrane: part of the major abnormal ATPase fraction is selectively inactivated by urea, producing a concomitant enrichment in the initially minor normal ATPase fraction.If the minor normal ATPase fraction is the only one capable of in vivo ATP synthesis, the deficient but oligomycin-sensitive cell growth and oxidative phosphorylation in vitro are readily explained.Further structural studies are under way to ascertain whether the minor normal ATPase fraction is strictly identical to the wild type, in which case PHO 1 is a regulatory gene, or not, in which case PHO 1 is a structural gene.     

Evidence of downstream migration of Sakhalin taimen, <Emphasis Type="Italic">Hucho perryi</Emphasis>, as revealed by Sr : Ca ratios of otolith

The migratory history of Sakhalin taimen, Hucho perryi, was examined in terms of strontium (Sr) and calcium (Ca) uptake in the otolith by using wavelength dispersive X-ray spectrometry on an electron microprobe. Otolioth Sr:Ca ratios of freshwater-reared samples remained consistently at low levels throughout the otolith. The Sr:Ca ratios of samples from Lake Aynskoye of Sakhalin Island showed a low value from the core up to a point of 700–2140µm. Thereafter, the ratios increased sharply and remained at higher levels up to the outermost regions. The difference in Sr:Ca ratio might be the result of the presence of individuals that underwent seawater and freshwater life history phases, probably reflecting the ambient salinity or the seawater–freshwater gradient in Sr concentration. Otolith Sr:Ca ratio analysis revealed downstream migration history in H. perryi.     

Enhanced acetyl esterase production by Fusarium oxysporum

Production of acetyl esterase (EC 3.1.1.6) by Fusarium oxysporum strain F3 was enhanced by optimization of growth conditions. Under optimal conditions, activities as high as 0.89U/ml of culture medium were obtained. The culture filtrate was equally active on p-nitrophenyl acetate and acetylxylan. The enzyme produced 71% deacetylation of acetylxylan in 2h at 40C. Activity was optimized at pH6.5 and at 55^C. The respective K_m values for p-nitrophenyl acetate and acetylxylan were 0.25mM and 1.05% (w/v) and the V_m values were 0.65 and 0.43mol acetate/min/mg protein.     

An epitope common to HLA class I and class II antigens,Ig light chains,and β 2-microglobulin

The homology of class I major histocompatibility complex (MHC) antigens, class II MHC antigens, and immunoglobulin molecules has suggested their divergence from a common ancestral gene. We report here a monoclonal antibody (mAb), PAC. M1, which reacts with HLA class I heavy chains, HLA class II and chains, and the light chain of human immunoglobulin by Western blot analysis. PAC.M1 reacted with 44 kd, 33 kd, and 29 kd species when tested on membrane glycoproteins from TRa1, a B-lymphoblastoid cell line (B-LCL). Two-dimensional electrophoresis and Western blotting of TRa1 glycoproteins showed that these species had the appropriate electrophoretic mobilities for class I heavy chain and class II and subunits. The presence of the epitope was verified on class II and subunits by Western blotting of purified -invariant chain complexes, and on class I heavy chains by Western blotting of purified class I antigens. The PAC. M1 mAb also reacted with immunoglobulin light chains when Western blotting was performed with normal human serum and purified IgG and IgM as antigens. While reactivity of the mAb with beta-2 microglobulin ( ₂m) was difficult to detect by Western blotting, binding of PAC.M1 to purified ₂m was detectable in a solid-phase binding assay. Thus, PAC.Ml reacts with a determinant shared by a number of members of the immunoglobulin superfamily.     

Optomotorische Untersuchung des visuellen systems einiger Augenmutanten der Fruchtfliege Drosophila

Summary The optical properties of the compound eye of Drosophila have been analysed using the optomotor reactions of flies with normal and mutant eye pigmentation. The stimulus was provided by cylindrical patterns with different periodic intensity distributions rotating at different speeds. The response consists of a torque about the vertical axis and was recorded under conditions of fixed flight. (Maximum reaction is about 0.04 dyn · cm). The transfer characteristics of the optical system are determined by the inter-ommatidial angle , influencing the resolving power and by the width of the visual field of single ommatidia , influencing the response at high spatial frequencies. The values = 4.6° and = 3.5° are obtained from stimulus-response experiments with Drosophila. They are independent of the presence of screening pigments. Differences in the response of flies with strong (+, se), weak (w ^a), and missing (w) pigmentation can be explained by the increased amount of scattered light in the pigment-deficient eyes. The overall intensities in the equally illuminated receptors are expected to be in the ratio 11825, respectively. The perception of motion depends only on the temporal, not on the spatial phase relations between periodic intensity variations in neighbouring ommatidia. Therefore the inhomogeneous distribution of the inter-ommatidial angle changes the resolving power in different parts of the eye without changing the response to motion. Different simultaneous stimuli of equal strength in different parts of the eye are averaged in the perceptive system of Drosophila according to the number of ommatidia in these parts.     

Charles Darwin's theory of evolution: A review of our present understanding

The paper characterizes Darwin's theory, providing a synthesis of recent historical investigations in this area. Darwin's reading of Malthus led him to appreciate the importance of population pressures, and subsequently of natural selection, with the help of the wedge metaphor. But, in itself, natural selection did not furnish an adequate account of the origin of species, for which a principle of divergence was needed. Initially, Darwin attributed this to geographical isolation, but later, following his work on barnacles which underscored the significance of variation, and arising from his work on botanical arithmetic, he supposed that diversity allowed more places to be occupied in a given region. So isolation was not regarded as essential. Large regions with intense competition, and with ample variation spread by blending, would facilitate speciation. The notion of place was different from niche, and it is questioned whether Darwin's views on ecology were as modern as is commonly supposed. Two notions of struggle are found in Darwin's theory; and three notions of variation. Criticisms of his theory led him to emphasize the importance of variation over a range of forms. Hence the theory was populational rather than typological. The theory required a Lamarckian notion of inheritable changes initiated by the environment as a source of variation. Also, Darwin deployed a use/habit theory; and the notion of sexual selection. Selection normally acted at the level of the individual, though kin selection was possible. Group selection was hinted at for man. Darwin's thinking (and also the exposition of his theory) was generally guided by the domestic-organism analogy, which satisfied his methodological requirement of a vera causa principle.     

Nuclear behavior in multinucleate Schizophyllum commune germlings

Summary The ultrastructure of a fir morphological mutant containing multinucleate cells is described in Schizophyllum commune. The germlings of basidiospores which arose from mating fir with wild-type mycelium were studied in culture by phase contrast microscopy to elucidate behavior of multinucleate cells. Nuclear division appeared synchronous from two nuclei yielding four progeny through six nuclei producing twelve products, beyond which loss of synchrony was indicated. Compensatory nuclear migration into an anucleate cell was presumed during synchronous division of nuclear aggregates in the adjacent cell of an individual germling. The migrant nucleus eventually returned to the cell of origin. However, the return route was not via the central pore of the septum but rather occurred at the juncture of the cross-wall with the germling-periphery. Ultrastructure of a partial septum in fir which could accommodate nuclear passage of this sort is described.     

α-Thalassemia and the production of different α chain variants in heterozygotes

The production of five chain variants (Hb G-Georgia, Hb St. Luke's, Hb Lloyd, Hb Montgomery, and Hb G-Philadelphia) in heterozygotes was evaluated through hematological observations, hemoglobin quantification, and biosynthetic studies. All heterozygotes for Hb St. Luke's and Hb Lloyd and most heterozygotes with Hb G-Georgia and Hb Montgomery had normal hematology and average / values of about 1.1. They were assigned a normal genotype (^G/), although the proportions of Hb St. Luke's and Hb G-Georgia were low (10 to 13%) and those of Hb Lloyd and Hb Montgomery twice as high (20%). Data from short-term incubations confirmed this genotype for some of these heterozygotes. Isolated Hb St. Luke's and Hb G-Georgia gave low ^G/ values (0.2 and 0.3) indicating that these Hb variants were defective at the level of Hb assembly. Isolated Hb Montgomery and Hb G-Philadelphia, however, gave higher ^G/ values of 0.6 and 0.8, respectively. A second type of variability existed among Hb G-Georgia (20 vs. 13%), Hb Montgomery (28 vs. 20%), and Hb G-Philadelphia (47 vs. 34%) heterozygotes, in whom the levels of Hb G differed. The occurrence of higher levels of these three chain heterozygosities was associated with hematological or biosynthetic evidence of a mild or moderate chain deficiency due to an -thalassemia-2 heterozygosity (^G/⁰ or ^0G/) or a homozygosity (^0G/⁰), respectively.This study was supported in part by USPHS Research Grants HLB-05168 and HLB-15158.     

Effect of 17β-estradiol on calcium response to phytohaemagglutinin in human lymphocytes

Pre-treatment of human lymphocytes with 17-estradiol diminishes the increase in concentration of cytosolic free calcium after stimulation with phytohaemagglutinin. The effect is dependent on 17-estradiol concentration and on the preincubation time. The effect is not due to an interaction between 17-estradiol and phytohaemagglutinin, but appears to be a consequence of the binding of the hormone to the cell surface. The effect is specific for 17-estradiol, since the isomer and other steroid hormones (progesterone, testosterone, diethylstilbestrol and 5-androstan), have no effect. Since the effect of the 17-estradiol can be suppressed by treatment of lymphocytes with ouabain, it appears that the effect of estradiol on the rise of cytosolic calcium induced by phytohaemagglutinin is mediated by the (Na, K)-ATPase.