Structural and functional analysis of the single-strand origin of replication from the lactococcal plasmid pWV01

The single-strand origin (SSO) of the rolling-circle (RC), broad-host-range lactococcal plasmid pWVO1 was functionally characterized. The activity of this SSO in the conversion of single-stranded DNA to double-stranded DNA was tested both in vivo and in vitro. In addition, the effect of this SSO on plasmid maintenance was determined. The functional pWVO1 SSO comprises a 250 by region, containing two inverted repeats (IRs). The activity of each IR was tested, separately and in combination, in a plasmid derivative that was otherwise completely devoid of structures that might function as SSO. One of the IRs (IR 1) showed some homology with other previously described SSOs of the SSO_A type, as well as with the conversion signal of the Escherichia coli phage X174. This IR was shown to have a partial, RNA polymerise-independent activity in complementary strand synthesis, both in vivo and in vitro. The second IR, which had no activity of its own, was required for full SSO activity, both in vivo and in vitro. The conversion of single-stranded DNA to the double-stranded form by the complete SSO was only partly sensitive to inhibition by rifampicin, indicating the existence of an RNA polymerase-independent pathway for this event. The results suggest that the pWVO1 SSO can be activated by two different routes: an RNA polymerise-dependent one (requiring the entire SSO), and an RNA polymerase-independent one (requiring only IR I).     

Analysis of randomly isolated cDNAs from developing endosperm of rice (Oryza sativa L.): evaluation of expressed sequence tags,and expression levels of mRNAs

Using a cDNA library prepared from poly(A)⁺ RNA from 10-day-old rice endosperm, partial nucleotide sequences of randomly isolated clones were analyzed. A total of 153 (30.6%) out of 500 cDNA clones showed high amino acid identity to previously identified genes. There was significant redundancy in cDNAs encoding prolamine and glutelin. About 21.0% of the cDNA clones were found to code for seed storage protein genes. Consequently, 37 independent genes were identified. Using cDNA clones encoding glutelin, prolamine, seed allergen, -1,4-glucan branching enzyme, glycine-rich RNA binding protein, metallothionein, non-specific lipid-transfer protein and ubiquitin conjugating enzyme the accumulation of mRNA during rice seed development was compared. Genes associated with seed storage protein and starch biosynthesis were expressed according to expected developmental stages. Glycinerich RNA binding protein genes as well as metallothionein-like protein genes were highly expressed in developing seeds, but low in leaves of whole plants.     

Plant regeneration from protoplasts isolated from cotyledons of Sesbania bispinosa

Protoplasts were isolated from cotyledons of Sesbania bispinosa (Jacq.) W.F. Wight. In a liquid-over-agar culture system with Murashige and Skoog (MS) medium supplemented with 1 mg l^-1 2,4-dichlorophenoxyacetic acid (2,4-d, 2 mg l^-1 benzyladenine (BA), 1 mg l^-1 glutamine and 0.5 and formed callus. The first division occurred after 3–4 days. Callus formed from the protoplasts differentiated shoots by organogenesis on MS medium with 1 mg l^-1 indolebutyric acid (IBA) and 1 mg l^-1 BA. These shoots developed into complete plantlets when excised and cultured on MS medium with 0.5 mg l^-1 IBA.     

Sequence requirements for the termination of rolling-circle replication of plasmid pT181

Most small multicopy plasmids of Gram-positive bacteria and many in Gram-negative bacteria replicate by a rolling-circle (RC) mechanism. The replication initiator proteins encoded by the RC plasmids and single-stranded bacteriophages of Escherichia coli have origin-specific nicking-closing activities that are required for the initiation and termination of RC replication. We have investigated the sequence requirements for termination of RC replication of plasmid pT181. The initiator nick site is located in the loop of a hairpin region (IRII) within the pT181 origin of replication. By mutational analysis, we have found that several nucleotides within the stem of IRII which are critical for the initiation activity are dispensable for termination of replication. We also demonstrate that nucleotides in the right arm of IRII, but not the left arm, are absolutely required for termination of RC replication. We have also identified specific nucleotides in IRII that are critical for its termination activity. The sequence of the right arm of the hairpin must be located downstream of the initiator nick site for termination, suggesting that termination requires a specific orientation of the initiator protein at the origin.     

Does PBAN play an alternative role of controlling pheromone emission in the cabbage looper moth,Trichoplusia ni (Hübner) (Lepidoptera: Noctuidae)?

There is an active process by which sex pheromone reserves of female cabbage looper moths, Trichoplusia ni, are transported to the gland's surface during the nocturnal period of calling. We hypothesized that this mobilization was controlled by a head factor, possibly related to the pheromone biosynthesis activating neuropeptides (PBAN) that in other species stimulate pheromone synthesis. We evaluated the impact of head extracts of T. ni on pheromone emission and glandular content of pheromone. During the photophase injected head extracts stimulated an increased pheromone emission rate in females, but glandular content of pheromone was not affected. Head extracts of H. virescens, a species with known PBAN activity, and synthetic PBAN stimulated an increased pheromone emission rate in T. ni. There was some specificity of the response of female T. ni to PBAN, in that several other unrelated polypeptides did not stimulate this type of response. Previously it had been determined that brain factors do not play a role in stimulating pheromone biosynthesis in T. ni. Our results indicate that there may be additional avenues by which PBAN or related neuropeptides control pheromone emission, including transport of pheromone reserves to the surface of the sex pheromone gland.     

δ-Glutamate Receptors Are Differentially Distributed at Parallel and Climbing Fiber Synapses on Purkinje Cells

Abstract: Neurons containing multiple excitatory inputs may sort and target glutamate receptor subtypes to subsets of synapses. A good model for testing this hypothesis is the Purkinje cell, which expresses significant levels of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionate, kainate, N -methyl- d -aspartate, δ-, and metabotropic glutamate receptors. Purkinje cells receive two excitatory inputs, the parallel and climbing fibers; the combined effect of stimulation of these two inputs is to produce long-term depression of parallel fiber/Purkinje cell neurotransmission. Distribution of glutamate receptors in these two synapse populations in rat cerebella was studied using preembedding immunocytochemistry with antibodies to GluR1, GluR2/3, GluR5-7, NR1, δ1/2, and mGluR1α. Moderate/dense postsynaptic staining was most frequent in postsynaptic densities and spines of both parallel and climbing fiber synapses with mGluR1α antibody, was intermediate in frequency with GluR2/3 and GluR5-7 antibodies, and was least frequent with GluR1 and NR1 antibodies. The most striking finding was the absence of significant postsynaptic staining with δ1/2 antibody in climbing fiber synapses in adult animals, even though postsynaptic staining was prevalent in parallel fiber synapses with this antibody. In contrast to adults, moderate/dense postsynaptic immunolabeling of climbing fiber synapses with δ1/2 antibody was common in rats at 10 days postnatal. This study provides direct morphological evidence that δ-glutamate receptors are differentially targeted to synapse populations. Our results support previous suggestions that δ2 is involved in development of parallel and climbing fiber synapses and in long-term depression of parallel fiber/Purkinje synaptic responses in adults.     

Physiologic regulation of mixtalol in rape senescence and its yield effects

Physiologic and yield effects of mixtalol at various concentrations sprayed on rape at the anthesis stage were examined. Foliar sprays of 4 and 2 ppm mixtalol significantly increased the chlorophyll content of rape leaves and pods, reduced the accumulation of malondialdehyde and ethylene production, and delayed the degradation of superoxide dismutase and catalase activities of the rape plant. Mixtalol also increased root oxidizability. Meanwhile, the number of branches and pods per plant was increased, and a 10.7% and 8.2% increase of seed yield over the controls was observed with treatments of 4 and 2 ppm mixtalol, respectively. No significant effects from mixtalol were observed on the maturation of plants or on the seed oil content or the erucic acid and glucosinolate content. Total rape oil production increased with 4 and 2 ppm mixtalol significantly by 12.4% and 10.5%, respectively, over the controls.Abbreviations MTL mixtalol - MDA malondialdehyde - TBA thiobarbituric acid - SOD Superoxide dismutase - CAT catalase - TTC tetrazolium     

Synthesis and antitumor activity of camptothecin derivatives bearing five-membered heterocycle containing 10-substituents

A series of new camptothecin derivatives bearing five-membered ring heterocycle containing substituents in the 10-position were synthesized and evaluated for in vitro cytotoxic activity. Camptothecin derivatives bearing a pyrrole or a thiophene ring were significantly more potent than camptothecin, however those bearing furan were less potent than camptothecin.     

科尔沁沙质草甸草场不同牧压条件植物群落分异数量分析

本文用ＴＷＩＮＳＰＡＮ分类和ＰＣＡ及ＤＣＡ排序技术分析了沙质草甸草场放牧试验引起地植物群落分异的特征,主要结论为：放牧强度是引起群落分异的主要原因,群落的分异主要表现在高度和现存生物量上。放牧引起群落变化的动力主要是放牧强度。而无牧条件下群落变化的主要动力是植物间的相互作用。当地的放牧强度处于中牧与重牧之间,属于过度放牧。这是引起草场退化及沙漠化的主要原因之一。     

显微红外研究巨噬细胞膜电融合后膜蛋白二级结构的变化

首次尝试了将显微傅里叶变换红外光谱术应用于研究电融合后细胞膜蛋白质二级结构的变化,发现脉冲电场作用于细胞具有穿透效应,施加电脉冲后,整个细胞的蛋白质体系能量增加,表明电泳冲对蛋白的二结构影响很大;同时,还发现用唾液酸苷酶和蛋白酶Ｐｒｏｎａｓｅ分析处理巨噬细胞膜表面后,膜上蛋白质二级结构无序化程度增加,用酶适度处理的细胞将更易发生电融合。