dsRNA binding properties of RDE-4 and TRBP reflect their distinct roles in RNAi

Double-stranded RNA (dsRNA)-binding proteins facilitate Dicer functions in RNA interference. Caenorhabditis elegans RDE-4 facilitates cleavage of long dsRNA to small interfering RNA (siRNA), while human trans-activation response RNA-binding protein (TRBP) functions downstream to pass siRNA to the RNA-induced silencing complex. We show that these distinct in vivo roles are reflected in in vitro binding properties. RDE-4 preferentially binds long dsRNA, while TRBP binds siRNA with an affinity that is independent of dsRNA length. These properties are mechanistically based on the fact that RDE-4 binds cooperatively, via contributions from multiple domains, while TRBP binds noncooperatively. Our studies offer a paradigm for how dsRNA-binding proteins, which are not sequence specific, discern dsRNA length. Additionally, analyses of the ability of RDE-4 deletion constructs and RDE-4/TRBP chimeras to reconstitute Dicer activity suggest RDE-4 promotes activity using its dsRNA-binding motif 2 to bind dsRNA, its linker region to interact with Dicer, and its C-terminus for Dicer activation.     

Intracellular signaling pathways involved in post-mitotic dopaminergic PC12 cell death induced by 6-hydroxydopamine

Oxidative stress has been shown to mediate neuron damage in Parkinson's disease (PD). In the present report, we intend to clarify the intracellular pathways mediating dopaminergic neuron death after oxidative stress production using post-mitotic PC12 cells treated with the neurotoxin 6-hydroxydopamine (6-OHDA). The use of post-mitotic cells is crucial, because one of the suggested intracellular pathways implicated in neuron death relates to the re-entry of neurons (post-mitotic cells) in the cell cycle. We find that 6-OHDA sequentially increases intracellular oxidants, functional cell damage and caspase-3 activation, leading to cell death after 12 h of incubation. Prevention of cell damage by different antioxidants supports the implication of oxidative stress in the observed neurotoxicity. Oxidative stress-dependent phosphorylation of the MAPK JNK and oxidative stress-independent PKB/Akt dephosphorylation are involved in 6-OHDA neurotoxicity. Decrease in p21(WAF1/CIP1) and cyclin-D1 expression, disappearance of the non-phosphorylated band of retinoblastoma protein (pRb), and expression of proliferating cell nuclear antigen, not present in PC12 post-mitotic cells, suggest a re-entry of differentiated cells into cell cycle. Our results indicate that such a re-entry is mediated by oxidative stress and is involved in 6-OHDA-induced cell death. We conclude that at least three intracellular pathways are involved in 6-OHDA-induced cell death in differentiated PC12 cells: JNK activation, cell cycle progression (both oxidative stress-dependent), and Akt dephosphorylation (not related to the increase of oxidants); the three pathways are necessary for the cells to die, since blocking one of them is sufficient to keep the cells alive.     

Senescence-induced ectopic expression of the A. tumefaciens ipt gene in wheat delays leaf senescence, increases cytokinin content, nitrate influx, and nitrate reductase activity, but does not affect grain yield

The manipulation of cytokinin levels by senescence-regulated expression of the Agrobacterium tumefaciens ipt gene through its control by the Arabidopsis SAG12 (senescence-associated gene 12) promoter is an efficient tool for the prolongation of leaf photosynthetic activity which potentially can affect plant productivity. In the present study, the efficiency of this approach was tested on wheat (Triticum aestivum L.)-a monocarpic plant characterized by a fast switch from vegetative to reproductive growth, and rapid translocation of metabolites from leaves to developing grains after anthesis. When compared with the wild-type (WT) control plants, the SAG12::ipt wheat plants exhibited delayed chlorophyll degradation only when grown under limited nitrogen (N) supply. Ten days after anthesis the content of chlorophyll and bioactive cytokinins of the first (flag) leaf of the transgenic plants was 32% and 65% higher, respectively, than that of the control. There was a progressive increase in nitrate influx and nitrate reductase activity. However, the SAG12::ipt and the WT plants did not show differences in yield-related parameters including number of grains and grain weight. These results suggest that the delay of leaf senescence in wheat also delays the translocation of metabolites from leaves to developing grains, as indicated by higher accumulation of ((15)N-labelled) N in spikes of control compared with transgenic plants prior to anthesis. This delay interferes with the wheat reproductive strategy that is based on a fast programmed translocation of metabolites from the senescing leaves to the reproductive sinks shortly after anthesis.     

Novel ecological niche of Cetobacterium somerae, an anaerobic bacterium in the intestinal tracts of freshwater fish

AIMS: This study was conducted to clarify the taxonomic status of Bacteroides type A strains with high vitamin B(12)-producing ability that is widely distributed in the intestinal tracts of freshwater fish. METHODS AND RESULTS: Seventeen strains of Bacteroides type A isolated from five fish species were all rod-shaped and gram-negative. The strains were positive for esculin hydrolysis, nitrate reduction, resistance to bile, acid phosphatase, and negative for the production of catalase and urease and the susceptibility to vancomycin. The G+C content of DNA from the 17 strains was 29 x 1-31 x 9 mol%, and 16S rDNA sequence analysis revealed a close phylogenetic relationship between Bacteroides type A strains and Cetobacterium somerae sharing 99 x 7-100% sequence similarity. In addition, strains were capable of producing vitamin B(12) at a rate of 1 x 82-13 x 98 ng ml(-1) in 48 h. CONCLUSION: Phenotypic and phylogenetic characteristics indicated that all isolates previously classified as Bacteroides type A strains belong to C. someare. SIGNIFICANCE AND IMPACT OF THE STUDY: This study provided the important finding of novel niche of vancomycin-resistant bacteria such as C. somerae in the intestinal tract of freshwater fish.     

四倍体刺槐的抗盐性

 以四倍体刺槐(Robinia pseudoacacia)为主要试验材料,以二倍体刺槐为对照材料,在两种盐胁迫下,对苗木的形态、生理生化、光合特性和解剖结构等指标的变化规律进行了研究。利用NaCl和Na₂SO₄盐溶液对两种刺槐进行盐处理,在30 d后每7 d处理1次,共处理4次,并在处理前和处理后每7 d进行各项指标的测定。结果发现：1)在盐胁迫下二倍体刺槐的植株生长受到强烈抑制,叶片含水量和叶绿素含量显著低于对照,有明显的盐害症状;对四倍体刺槐植株的生长影响较小,叶片含水量和叶绿素含量也与对照差异不显著,无盐害症状。2)四倍体刺槐经过盐处理后相对电导率和脯氨酸(Proline, Pro)含量虽然也稍有上升,但与对照相比未达到显著水平,而二倍体刺槐显著高于对照;同时作为保护酶系统的过氧化物酶(Peroxidase, POD)、超氧化物歧化酶(Superoxide dismutase, SOD)、过氧化氢酶(Catalase, CAT)在四倍体刺槐经盐胁迫后期也保持了较高的活性,从而提高了其抗盐性,而对盐敏感的二倍体刺槐3个保护酶活性均较低。 3)经盐胁迫后对四倍体刺槐光合特性影响不大,净光合速率（Net photosynthetic rate, P_n）和胞间CO₂浓度（Intercellular CO₂ concentration, C_i）均无显著变化,而二倍体刺槐P_n和C_i显著下降。4)经盐胁迫后四倍体刺槐在解剖结构上做出了积极的反应：叶肉栅栏组织拉长、排列更为紧密、 海绵组织变小、排列紧密。而二倍体刺槐出现了相反的现象。综合以上分析认为：四倍体刺槐具有较强的抗盐性。     

EFFECTS OF LIGHT AND TEMPERATURE ON SEED GERMINATION OF FICUS HISPIDA IN XISHUANGBANNA, SOUTHWEST CHINA

 该研究应用人工气候箱设置不同的光照和温度梯度, 探讨对对叶榕(Ficus hispida)种子萌发的影响。结果表明, 荧光灯条件下(红光和远红光比例(R/FR)为4.56, 光量子密度(PPFD)约为90 μmol&;#8226;m^–2&;#8226;s^–1), 温度对种子萌发速率有显著影响, 而对最终萌发率的影响不显著。对叶榕的种子为光敏性种子, 萌发严格需光, R/FR对种子最终萌发率的影响显著, 较高的R/FR促进种子萌发, 而较低的 R/FR 抑制种子萌发。光强影响种子的萌发速率, 较弱的光照可以延缓种子萌发, 但并不能完全抑制其萌发。在30 ℃条件下, 对叶榕的种子可以在较高的 R/FR (0.42) 水平下萌发, 但温度为23/20 ℃ 时, 种子对 R/FR 的要求增高, 0.42的R/FR不能导致种子萌发。说明较低的温度和 R/FR 都可以显著抑制对叶榕的种子萌发, 而较高的温度和R/FR皆有利于种子的萌发。     

双歧杆菌及其完整肽聚糖对脐血来源树突状细胞分泌IL-12的影响

目的探讨两歧双歧杆菌和不同剂量双歧杆菌的完整肽聚糖（WPG）对脐血来源树突状细胞（DC）分泌IL-12的影响。方法以双歧杆菌全菌（量）和不同剂量双歧杆菌WPG（1-8μg/ml）与脐血来源树突状细胞共培养,用ELISA的方法测定培养上清中IL-12的量。结果双歧杆菌和其WPG（1-6μg/ml）与树突状细胞共培养后,树突状细胞分泌的IL-12的量显著高于阴性对照组（P〈0.01）,当WPG量为1-5μg/ml时树突状细胞分泌的IL-12的量呈剂量依赖性,其中WPG量为5μg/ml时作用最为显著,WPG量为6μg/ml时分泌IL-12量减少,WPG量为8μg/ml时,分泌的IL-12量与阴性对照组差异无显著性（P〉0.05）。结论两歧双歧杆菌及其WPG能够刺激脐血来源的树突状细胞IL-12分泌;双歧杆菌WPG的免疫刺激作用呈一定的量效关系     

Stretch-induced activation of ERK in myocytes is p38 and calcineurin-dependent

Activation of specific mitogen-activated protein kinases (MAPKs) has been suggested to be involved in phenotype modulation of cells subjected to mechanical strain, which may be common to different mechano-sensitive cell types. We have submitted C2C12 myocytes to a static stretch and examined its effect upon the activation of ERK. Stretch induced a rapid but transient activation of ERK. This activation was however prevented when cells were pre-treated with inhibitors of p38 and calcineurin. The dependence of strain-induced ERK activation upon p38 suggests a cross-talk between these two pathways when mediating a response to a mechanical stimulus in this cell type. This suggests that cross relationships between these MAP kinases may be of crucial importance for myocyte phenotype modulation and differentiation in response to a mechanical stimulus.