A molecular marker confirms that the rate of adult maturation is largely independent of the rate of pre-adult development in Drosophila melanogaster

The separation of adult from pre-adult life seen with animals such as Drosophila melanogaster, which are holometabolous and undergo complete metamorphosis, provides the opportunity to examine the contribution of pre-adult rate of development on the rate of maturation and aging of the adult. Recent work has shown that when ambient temperature is used to alter the rate of development there is little effect on adult life span. From this work it has been concluded that the rate of aging is largely independent of the rate of pre-adult development. However, the techniques used to examine life span did not allow for the examination of the earliest events of adult life. Our experimental design used a molecular marker linked to life span as a sensitive measure of determining physiological age. In this way, we were able to evaluate the effect of pre-adult rate of development on the earliest events of adult life. Using ambient temperature to alter both the rate of development in the pre-adult and the rate of aging in the adult independently, we were able to show that it is the ambient temperature at which the adults are living that is the principle determinant of the rate of maturation and aging of the adult. Little effect was seen on the rate of adult maturation in response to an acceleration or a slowing down of the rate of pre-adult development as measured by our molecular marker. These data support the conclusions drawn by others who examined the effect of the rate of development on adult life expectancy. The timing mechanisms at work during pre-adult and adult life appear to be largely regulated separately. If there is such a thing as a physiological clock, it appears to be reset upon eclosion. © 1996 Wiley-Liss, Inc.     

Structure-guided affinity maturation of a single-chain variable fragment antibody against the Fu-bc epitope of the dengue virus envelope protein

Dengue is one of the most dominant arthropod-borne viral diseases, infecting at least 390 million people every year throughout the world. Despite this, there is no effective treatment against dengue, and the only available vaccine has already been withdrawn owing to the significant adverse effects. Therefore, passive immunotherapy using monoclonal antibodies is now being sought as a therapeutic option. To date, many dengue monoclonal antibodies have been identified, most of which are serotype-specific, and only a few of which are cross-reactive. Furthermore, antibodies that cross-react within serotypes are weakly neutralizing and frequently induce antibody-dependent enhancement, which promotes viral entry and replication. Therefore, broadly neutralizing antibodies with no risk of antibody-dependent enhancement are required for the treatment of dengue. Here, we developed a single-chain variable fragment (scFv) antibody from an anti-fusion loop E53 antibody (PDB: 2IGF). We introduced previously predicted favorable complementarity-determining region (CDR) mutations into the gene encoding the scFv antibody for affinity maturation, and the resultant variants were tested in vitro against the highly conserved fusion and bc epitope of the dengue virus envelope protein. We show some of these scFv variants with two to three substitution mutations in three different CDRs possess affinity constants (K_D) ranging from 20 to 200 nM. The scFv-mutant15, containing D31L, Y105W, and S227W substitutions, showed the lowest affinity constant, (K_D = 24 ± 7 nM), approximately 100-fold lower than its parental construct. We propose that the scFv-derivative antibody may be a good candidate for the development of an effective and safe immunotherapy.     

The refined structure of a protein catenane: the HK97 bacteriophage capsid at 3.44 A resolution

The HK97 bacteriophage capsid is a unique example of macromolecular catenanes: interlocked rings of covalently attached protein subunits. The chain mail organization of the subunits stabilizes a particle in which the maximum thickness of the protein shell is 18A and the maximum diameter is 550A. The electron density has the appearance of a balloon illustrating the extraordinary strength conferred by the unique subunit organization. The refined structure shows novel qualities of the HK97 shell protein, gp5 that, together with the protease gp4, guides the assembly and maturation of the virion. Although gp5 forms hexamers and pentamers and the subunits exist in different structural environments, the tertiary structures of the seven protein molecules in the viral asymmetric unit are closely similar. The interactions of the subunits in the shell are exceptionally complex with each subunit interacting with nine other subunits. The interactions of the N-terminus released after gp5 cleavage appear important for organization of the loops that become crosslinked to the core of a neighboring subunit at the maturation. A comparison with a model of the Prohead II structure revealed that the surfaces of non-covalent contact between the monomers that build up hexamers/pentamers are completely redefined during maturation.     

Tillering in grain sorghum over a wide range of population densities: identification of a common hierarchy for tiller emergence,leaf area development and fertility

Most studies of tiller development have not related the physiological and morphological features of each culm to its subsequent fertility. This introduces problems when trying to account for the effects of tillering on yield in crop models. The objective of this study was to detect the most likely early determinants of tiller fertility in sorghum by identifying hierarchies for emergence, fertility and grain number of tillers over a wide range of assimilate availabilities. Emergence, phenology, leaf area development and dry weight partitioning were quantified weekly for individual tillers and main culms of tillering and uniculm plants grown at one of four densities, from two to 16 plants m(-2). For a given plant in any given density, the same tiller hierarchy applied for emergence of tillers, fertility of the emerged tillers and their subsequent grain number. These results were observed over a range of tiller fertility rates (from 7 to 91%), fertile tiller number per plant at maturity (from 0.2 to 4.7), and tiller contribution to grain yield (from 5 to 78 %). Tiller emergence was most probably related to assimilate supply and light quality. Development, fertility and contribution to yield of a specific tiller were highly dependent on growing conditions at the time of tiller emergence, particularly via early leaf area development of the tiller, which affected its subsequent leaf area accumulation. Assimilate availability in the main culm at the time of tiller emergence was the most likely early determinant of subsequent tiller fertility in this study.     

Fructan contents and dry matter deposition in different tissues of the wheat grain during development

The role of fructan metabolism in the assimilate relations of the grain of wheat (Triticum aestivum L.) was investigated by determination of the dry matter and fructan content of grain components at short intervals during grain filling. During the initial phase of rapid expansion, most of the assimilates entering the grain were partitioned to the outer pericarp. A large fraction of these assimilates were used for the synthesis of fructan. Dry matter deposition and fructan synthesis in the outer pericarp ceased at about 5d after anthesis. At the same time, the endosperm and the inner pericarp and testa started to accumulate dry matter at a fast rate. This was also associated with significant fructan synthesis in the latter tissues. The outer pericarp lost about 45% of its former maximum dry weight between 9 and 19 d after anthesis. This loss was due almost entirely to the near complete disappearance of water-soluble carbohydrates, most of which was fructan. The inner pericarp and testa accumulated dry matter until about mid-grain filling. The fructan contents of the inner pericarp and testa and the endosperm decreased slowly towards the end of grain filling. Most of the fructans in the inner pericarp and testa and the endosperm had a low molecular weight, whereas higher molecular weight fructans predominated in the outer pericarp. The embryo did not contain fructan. The presence of low molecular weight fructans in the endosperm cavity at mid-grain filling was confirmed. It is suggested that fructan synthesis is closely linked to growth-related water deposition in the different tissues of the wheat grain and serves to sequester the surplus of imported sucrose.     

Experimental Identification of “Songlan” (Isatis indigotica) and Woad (I.tinctoria) Introduced into China

“Song Lan” is a source of Chinese drugs such as “Daqingye”, “Banlangen” or “Qingdai”. We have discovered that the two species, “Song Lan” (Isatis indigotica Fort.) and woad (I. tinctoria L.), were mistakenly described in the literature due to their morphological polymorphism. In order to clarify the two species, cytology examination, pollen analysis, electrophoretic analysis of isoenzymes and soluble protein were performed. The results show that previous non-trichiferous type of woad is a “Song Lan”. As in woad, “Song Lan” is also morphologically of great variability. The base of canline leaves in this species may be sagittate or auriculate. We have not found the non-trichiferous type of woad in our country. It is reported for the first time that the chromosome number for “Song Lan” is 2n=14. The content of the indole glucoside in fresh leaves of “Song Lan” is about five timeshigher than in woad. For medicine cultivation of “Song Lan” is favorable.     

海甘蓝种子成熟过程中脂肪酸含量以及脱落酸和山梨醇的效应

海甘蓝种子在成熟过程中,棕榈酸、硬脂酸和亚麻酸的含量不断下降,而二十碳烯酸和芥酸的含量呈上升趋势。选用开花后２５～２７ｄ的海甘蓝幼胚分别在含不同浓度的ＡＢＡ或高渗透剂的培养基中培养１～３ｄ,发现其各种脂肪酸的变化趋势和种子自然成熟过程中脂肪酸的变化相似,说明ＡＢＡ或高渗透剂可能是种子成熟过程中各种脂肪酸合成和相互转化所需的条件。     

Involvement of abscisic acid and cytokinins in the senescence and remobilization of carbon reserves in wheat subjected to water stress during grain filling

This study investigated the possibility that abscisic acid (ABA) and cytokinins may mediate the effect of water deficit that enhances plant senescence and remobilization of pre‐stored carbon reserves. Two high lodging‐resistant wheat (Triticum aestivum L.) cultivars were field grown and treated with either a normal or high amount of nitrogen at heading. Well‐watered (WW) and water‐stressed (WS) treatments were imposed from 9 d post‐anthesis until maturity. Chlorophyll (Chl) and photosynthetic rate (Pr) of the flag leaves declined faster in WS plants than in WW plants, indicating that the water deficit enhanced senescence. Water stress facilitated the reduction of non‐structural carbohydrate in the stems and promoted the re‐allocation of prefixed ¹⁴C from the stems to grains, shortened the grain filling period and increased the grain filling rate. Water stress substantially increased ABA but reduced zeatin (Z) + zeatin riboside (ZR) concentrations in the stems and leaves. ABA correlated significantly and negatively, whereas Z + ZR correlated positively, with Pr and Chl of the flag leaves. ABA but not Z + ZR, was positively and significantly correlated with remobilization of pre‐stored carbon and grain filling rate. Exogenous ABA reduced Chl in the flag leaves, enhanced the remobilization, and increased grain filling rate. Spraying with kinetin had the opposite effect. The results suggest that both ABA and cytokinins are involved in controlling plant senescence, and an enhanced carbon remobilization and accelerated grain filling rate are attributed to an elevated ABA level in wheat plants when subjected to water stress.     

Bovine cumulus cell expansion does not depend on the presence of an oocyte secreted factor

Communication between the oocyte and its somatic cells has been shown to be important in oocyte development. Here we examined how the oocyte may be involved in bovine cumulus cell expansion. Intact bovine cumulus oocyte complexes (COC) were obtained by puncturing antral follicles. From the intact COC, oocytectomised complexes (OOX) were produced by micro surgical removal of the oocyte. Clumps of cumulus cells (CC) were obtained by micro-dissection. Intact or OOX complexes or CC were matured in the presence of fetal calf serum and hFSH (6 mlU/ml) for 24 hr and the degree of expansion measured. The presence of the oocyte is not essential to allow bovine cumulus expansion to occur as expansion occurred in all groups. Murine OOX complexes from eCG primed 35–40-day-old C57BL6/CBA F₁ hybrids (known to require the presence of an oocyte secreted factor for cumulus expansion) were cultured with or without denuded bovine oocytes (1 oocyte/μl). Murine OOX complexes expanded only in the presence of denuded bovine oocytes. Thus some factor produced by bovine oocytes enabled expansion of murine OOX complexes. To determine whether the factor is secreted by bovine oocytes, murine OOX were cultured with or without media conditioned by bovine oocytes (1 oocyte/μl for 4 hr). Significant expansion of murine OOX occurred in media conditioned by bovine oocytes. This shows that the cumulus expansion enabling effect of bovine oocytes is released into the surrounding media. Media conditioned by bovine oocytes and then frozen for up to 1 month showed that the activity by the factor can withstand freezing. © 1995 wiley-Liss, Inc.