谷类作物β-葡聚糖合成酶基因家族的研究进展

Beta-葡聚糖是由β-(1,3)和β-(1,4)糖苷键连接的非纤维素多糖,主要分布在谷类作物籽粒胚乳及糊粉层中,在高尔基体合成,经由囊泡运输到质膜,最终在细胞壁上沉积。通过增加胆汁酸排泄,延迟葡萄糖吸收,β-葡聚糖可有效降低胆固醇及血糖水平。Beta-葡聚糖合成酶基因家族成员最早在水稻(Oryza sativa)中得到鉴定,后在其他作物中陆续被发现。该基因家族包括3个主要成员：CslF、CslH和CslJ亚基因家族,起源于不同分支,经过趋同演化,执行合成β-葡聚糖的功能。Beta-葡聚糖基因家族成员均受到负选择压力,演化过程中序列高度保守。CslF亚家族基因成员相对较多,常在染色体上形成基因簇,CslF6是介导β-葡聚糖合成的主效基因。CslF亚家族在叶基部等幼嫩组织中表达水平相对较高,且明显受到光照强度的影响;CslH和CslJ亚家族成员较少,其中CslH亚家族在叶尖等成熟组织中的表达水平高,而CslJ亚家族在籽粒中有较高的表达水平。该文综述了β-葡聚糖合成酶基因家族成员的系统发育关系、表达模式,β-葡聚糖合成酶的亚细胞定位,以及作物中的定向育种研究进展,提出β-葡聚糖合成酶基因家...     

Comparisons of biomass,water use efficiency and water use strategies across five genomic groups of Populus and its hybrids

Genetic improvement and hybridization in the Populus genus have led to the development of genotypes exhibiting fast growth, high rooting ability and disease resistance. However, while large biomass production is important for bioenergy crops, efficient use of resources including water is also important in sites lacking irrigation and for maintaining ecosystem water availability. In addition, comparison of water use strategies across a range of growth rates and genetic variability can elucidate whether certain strategies are shared among the fastest growing and/or most water use efficient genotypes. We estimated tree water use throughout the second growing season via sapflow sensors of 48 genotypes from five Populus taxa; P. deltoides W. Bartram ex Marshall × P. deltoides (D × D), P. deltoides × P. maximowiczii A. Henry (D × M), P. deltoides × P. nigra L. (D × N), P. deltoides × P. trichocarpa Torr. & Gray (D × T) and P. trichocarpa × P. deltoides (T × D) and calculated average canopy stomatal conductance (G_S). We regressed G_S and atmospheric vapor pressure deficit (VPD) wherein the slope of the relationship represents stomatal sensitivity to VPD. At the end of the second growing season, trees were harvested, and their dry woody biomass was used to calculate whole tree water use efficiency (WUE_T). We found that D × D and D × M genotypes exhibited differing water use strategies with D × D genotypes exhibiting high stomatal sensitivity while retaining leaves while D × M genotypes lost leaf area throughout the growing season but exhibited low stomatal sensitivity. Across measured taxa, biomass growth was positively correlated with WUE_T, and genotypes representing each measured taxa except D × N and T × D had high 2-year dry biomass of above 6 kg/tree. Overall, these data can be used to select Populus genotypes that combine high biomass growth with stomatal sensitivity and WUE_T to limit the negative impacts of bioenergy plantations on ecosystem water resources.     

Tomato exo-(1-->4)-beta-D-galactanase. Isolation, changes during ripening in normal and mutant tomato fruit, and characterization of a related cDNA clone.

An exo-(1-->4)-beta-D-galactanase was isolated from ripe tomato fruit (Lycopersicon esculentum Mill. cv Ailsa Craig and cv Better Boy) using anion-exchange, gel filtration, and cation-exchange chromatography. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the most active fraction revealed a predominant protein band at 75 kD and several minor bands. A 30-amino acid N-terminal sequence from this 75-kD protein showed a high degree of homology with other recently identified beta-galactosidase/ galactanase proteins from persimmon and apple fruits (I.-K. Kang, S.-G. Suh, K.C. Gross, J.-K. Byun [1994] Plant Physiol 105: 975-979; G.S. Ross, T. Wegrzyn, E.A. MacRae, R.J. Redgwell [1994] Plant Physiol 106: 521-528) and with the predicted polypeptide sequence encoded by the ethylene-regulated SR12 gene in carnation (K.G. Raghothama, K.A. Lawton, P.B. Goldsbrough, W.R. Woodson [1991] Plant Mol Biol 17: 61-71). The enzyme focused to a single band of beta-galactosidase activity on an isoelectrofocusing gel at pH 9.8. The enzyme was specific for (1-->4)-beta-D-galactan substrates with a pH optimum of 4.5. The only reaction product detected was monomeric galactose, indicating that the enzyme was an exo (1-->4)-beta-D-galactanase. beta-Galactanase activity increased at the onset of ripening in normal fruit, but no similar increase was detected in the nonripening mutants nor and rin. A tomato homolog (pTombetagal1) was isolated using the SR12 cDNA clone from carnation as a probe. This clone showed 73% identify at the amino acid level with beta-galactosidase-related sequences from apple and asparagus and 66% identity with SR12. pTombetagal1 is a member of a gene family. Northern analysis demonstrated that pTombetagal1 expression was ripening related in normal fruits, with lower levels apparent in the nonsoftening mutants.     

Energy-dispersive X-ray analysis of the extracellular cadmium sulfide crystallites of Klebsiella aerogenes

Klebsiella aerogenes forms electron-dense partieles on the cell surface in response to the presence of cadmium ions in the growth medium. These particles ranged from 20 to 200 nm in size, and quantitative energy dispersive X-ray analysis established that they comprise cadmium and sulfur in a 1:1 ratio. This observation leads to the conclusion that the particles are cadmium sulfide crystallites. A combination of atomic absorption spectroscopy, inductively coupled plasma mass spectrometry, and acid-labile sulfide analysis revealed that the total intracellular and bound extracellular cadmium:sulfur ratio is also 1:1, which suggests that the bulk of the cadmium is fixed as extracellular cadmium sulfide. The tolerance of K. acrogenes to cadmium ions and the formation of the cadmium sulfide crystallites were dependent on the buffer composition of the growth medium. The addition of cadmium ions to phosphate-buffered media resulted in cadmium phosphate precipitates that remove the potentially toxic cadmium ions from the growth medium. Electrondense particles formed on the surfaces of bacteria grown under these conditions were a combination of cadmium sulfide and cadmium phosphates. The specific bacterial growth rate in the exponential phase of batch cultures was not affected by up to 2mM cadmium in Tricine-buffered medium, but formation of cadmium sulfide crystallites was maximal during the stationary phase of batch culture. Cadmium tolerance was much lower (10 to 150 M) in growth media buffered with Tris, Bistris propane, Bes, Tes, or Hepes. These results illustrate the importance of considering medium composition when comparing levels of bacterial cadmium tolerance.Abbreviations EDXA Energy dispersive X-ray analysis - AAS Atomic absorption spectroscopy - TEM Transmission electron microscopy - SEM Scanning electron microscopy - ICP-MS Inductively coupled plasma mass spectrometry - ALSA Acid-labile sulfide analysis     

Peptide modification by introduction ofα-trifluoromethyl substituted amino acids

Summary Methodology for the synthesis and incorporation of-trifluoromethyl substituted amino acids into N- and C-terminal position of peptides is described. The incorporation of-trifluoromethyl substituted amino acids into strategical positions of peptides enhances proteolytic stability and lipophilicity. Furthermore, it improves transport rates in vivo and permeability through certain body barriers.     

Analytical subcellular fractionation of needle-biopsy specimens from human liver.

1. Fragments (2-20 mg wet wt.) of closed needle-biopsy specimens from human liver were disrupted in iso-osmotic sucrose and subjected to low-speed centrifugation. The supernatant was layered on a linear sucrose-density gradient in the Beaufay small-volume automatic zonal rotor. The following organelles, with equilibrium densities (g/ml) and principal marker enzyme shown in parentheses, were resolved: plasma membrane (1.12-1.14; 5'-nucleotidase); lysosomes (1.15-1.20; N-acetyl-beta-glucosaminidase); mitochondria (1.20; malate dehydrogenase); endoplasmic reticulum (1.17-1.21; neutral alpha-glucosidase); peroxisomes (1.22-1.24; catalase). 2. The distribution of particulate alkaline phosphatase and, to a lesser degree, leucine 2-naphthylamidase followed that of 5'-nucleotidase. gamma-Glutamyltransferase was associated with membranes of significantly higher equilibrium density than was 5'-nucleotidase. 3. The distribution of 12 acid hydrolases was determined in the density-gradient fractions. beta-Glucosidase had a predominantly cytosolic localization, but the other enzymes showed a broad distribution of activity throughout the gradient. Evidence was presented for two populations of lysosomes with equilibrium densities of 1.15 and 1.20 g/ml, but containing differing amounts of each enzyme. Further evidence of lysosomal heterogeneity was demonstrated by studying the distribution of isoenzymes of hexosaminidase and of acid phosphatase. 4. The resolving power of the centrifugation procedure can be further enhanced with membrane perturbants. Digitonin (0.12 mM) selectively disrupted lysosomes, markedly increased the equilibrium density of plasma-membrane components and lowered the density of the endoplasmic reticulum, but did not affect the mitochondria or peroxisomes. Pyrophosphate (15 mM) selectively lowered the equilibrium density of the endoplasmic reticulum.     

Adaptations and responses of Dasymutilla occidentalis (Hymenoptera: Mutillidae) to predators

The mutillid wasp Dasymutilla occidentalis possesses several adaptations and exhibits a number of responses wich appear to be of defensive value: a long mobile sting with powerful venom; a strong, rounded and slippery cuticle; an ability to run very rapidly and evasively; an aposematic warning coloration pattern; and the ability to respond to an attack by making stridulatory sounds and by releasing a chemical secretion or both. The effectiveness of these defenses is supported by tests utilizing various vertebrate and arthropod predators. The raison d'être of the multiple lines of defense possessed by D. occidentalis and the relative value of each line of defense are discussed. It is postulated that aposematic coloration, audible stridulation, and a volatile defensive exudate all function primarily as part of an early warning system enabling a predator to recognize this wasp-with its very algogenic venom-as unpalatable and potentially dangerous.

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Zusammenfassung Die Mutillide Wespe Dasymutilla occidentalis besitzt verschiedene Anpassungen und zeigt eine Anzahl von Reaktionen, die für die Verteidigung von Wert sind: ein langer, beweglicher Stachel mit starkem Gift, eine starke, runde und glatte Kutikula, die Möglichkeit, sehr schnell und ausweichend zu laufen sowie ein aposematisches Warn-Farben-Muster. Desweiteren ist sie fähig, einem Angriff mit einem knisternden Ton entgegenzutreten sowie ein chemisches Sekret abzugeben oder auch beides. Im Labor durchgeführte Zusammenstöße zwischen vertebraten und invertebraten Räubern und D. occidentalis beweisen klar den Wert der ganzen Anzahl von Verteidigungsmechanismen für die Wespe.Diese Zusammenstöße liefern auch einen Einblick in die raison d'être der vielseitigen Verteidigung. die stärkere Kutikula und ihre Glätte, Haupteigenschaften zum überleben gegen Räuber, funktionieren gleichzeitig mit dem Stich zum Schutz gegen Vertebraten und mit schneller Fluchtmöglichkeit zum Schutz gegen die meisten Invertebraten. Das akustische Geräusch scheint eine Hilfsverteidigung zu sein wenigstens gegen einige Spinnen und vermutlich auch gegen einige Vertebraten. Die chemische Sekretion, die hauptsächlich aus 4-methyl-3-heptanon besteht, scheint möglicherweise direkten Verteidigungswert gegen einige Eidechsen zu haben und funktioniert höchstwahrscheinlich hauptsächlich im Zusammenhang mit der roten und schwarzen aposematischen Färbung und dem knisternden Ton als vielseitiges Warnsystem, das fähig ist, durch Signalisieren allen potentiellen vertebraten Räubern mitzuteilen, daß dieses Insekt ungenießbar ist.

     

An in situ assay for induced diphtheria-toxin-resistant mutants of diploid human fibroblasts

The sensitivity of diploid human fibroblasts to the cytotoxic effects of diphtheria toxin (DT) depended on the cell growth status. Exponentially growing cells treated with 10^?3-1 lethal flocculating units (LF) of DT/ml for 4 days survived with a frequency of 4 × 10^?4. However, the DT-resistant phenotype of colonies isolated under these conditions was not stable. When the growth of the cells had been arrested by confluence or deprivation of serum growth factors prior to treatment with DT (4 days, 10^?3-0.6 LF/ml), the survival decreased to 2 × 10^?6 and the resistance of isolated colonies was stable. An in situ assay for induced DT-resistant mutants was developed in order to avoid problems associated with the possible reduced viability of the mutants relative to that of wild-type cells. A reproducible and linear dose response was obtained for the induction of DT-resistant mutants by ethylnitrosourea. The mutants were induced with high frequency by this compound (e.g., 10^?3 mutants/viable cell at a 37% survival dose); complete expression of the mutant phenotype occurred after 6 generations of growth under nonselective conditions. Isolated mutant colonies showed stable resistance to DT and were cross-resistant to Pseudomonas aeruginosa exotoxin A.