The Role of Autophagy in Lamellar Body Formation and Surfactant Production in Type 2 Alveolar Epithelial Cells

The lamellar body (LB), a concentric structure loaded with surfactant proteins and phospholipids, is an organelle specific to type 2 alveolar epithelial cells (AT2). However, the origin of LBs has not been fully elucidated. We have previously reported that autophagy regulates Weibel-Palade bodies (WPBs) formation, and here we demonstrated that autophagy is involved in LB maturation, another lysosome-related organelle. We found that during development, LBs were transformed from autophagic vacuoles containing cytoplasmic contents such as glycogen. Fusion between LBs and autophagosomes was observed in wild-type neonate mice. Moreover, the markers of autophagic activity, microtubule-associated protein 1 light chain 3B (LC3B), largely co-localized on the limiting membrane of the LB. Both autophagy-related gene 7 (Atg7) global knockout and conditional Atg7 knockdown in AT2 cells in mice led to defects in LB maturation and surfactant protein B production. Additionally, changes in autophagic activity altered LB formation and surfactant protein B production. Taken together, these results suggest that autophagy plays a critical role in the regulation of LB formation during development and the maintenance of LB homeostasis during adulthood.     

Thirty years of resistance: Zig-zag through the plant immune system

Understanding the plant immune system is crucial for using genetics to protect crops from diseases. Plants resist pathogens via a two-tiered innate immune detection-and-response system. The first plant Resistance (R) gene was cloned in 1992 . Since then, many cell-surface pattern recognition receptors (PRRs) have been identified, and R genes that encode intracellular nucleotide-binding leucine-rich repeat receptors (NLRs) have been cloned. Here, we provide a list of characterized PRRs and NLRs. In addition to immune receptors, many components of immune signaling networks were discovered over the last 30 years. We review the signaling pathways, physiological responses, and molecular regulation of both PRR- and NLR-mediated immunity. Recent studies have reinforced the importance of interactions between the two immune systems. We provide an overview of interactions between PRR- and NLR-mediated immunity, highlighting challenges and perspectives for future research.

A review of major research advances in plant immunity during the last three decades and individual characterized immune receptors, their immune signaling pathways, and interactions between immune systems     

人巨细胞病毒的生物素DNA探针的制备和应用

本研究用克隆的HCMV AD169株DNA片段,制备了生物素标记的DNA探针,建立了检测临床脐带血、尿标本中HCMV DNA的核酸探针杂交方法。该探针可测出100pg同源DNA,不与人胚肺细胞、Hep-2细胞DNA以及其他疱疹病毒的DNA发生反应。用核酸杂交方法检测了30份脐带血标本,有11例阳性,阳性率为33％。10例孕妇尿标本中,3例阳性,阳性率为30％。检测结果表明：我们建立的生物素标记的HCMV DNA探针的点杂交法,具有高度的特异性、敏感性,比分离病毒法更迅速,可用于HCMV感染的临床标本的病毒核酸检测。     

Decreased PLTP mass but elevated PLTP activity linked to insulin resistance in HTG: effects of bezafibrate therapy

Hypertriglyceridemia (HTG) is associated with insulin resistance, increased cholesteryl ester transfer (CET), and low HDL cholesterol. Phospholipid transfer protein (PLTP) may be involved in these relationships. Associations between CET, lipids, insulin resistance, CETP and PLTP activities, and PLTP mass were investigated in 18 HTG patients and 20 controls. Effects of 6 weeks of bezafibrate treatment were studied in HTG patients. HTG patients had higher serum triglycerides, insulin resistance, free fatty acid (FFA), and CET, lower levels of HDL cholesterol (-44%) and PLTP mass (-54%), and higher CETP (+20%) and PLTP activity (+48%) than controls. Bezafibrate reduced triglycerides, CET (-37%), insulin resistance (-53%), FFA (-48%), CETP activity (-12%), PLTP activity (-8%), and increased HDL cholesterol (+27%), whereas PLTP mass remained unchanged. Regression analysis showed a positive contribution of PLTP mass (P = 0.001) but not of PLTP activity to HDL cholesterol, whereas insulin resistance positively contributed to PLTP activity (P < 0.01). Bezafibrate-induced change in CET and HDL cholesterol correlated with changes in CETP activity and FFAs, but not with change in PLTP activity. Bezafibrate-induced change in PLTP activity correlated with change in FFAs (r = 0.455, P = 0.058). We propose that elevated PLTP activity in HTG is related to insulin resistance and not to increased PLTP mass. Bezafibrate-induced diminished insulin resistance is associated with a reduction of CET and PLTP activity.     

From subgenome analysis to protein structure

Groups of related genes abound in large eukaryotic genomes. In such 'subgenomes', homology modeling carried out for a few genes will probably have relevance to the entire group. Subgenomes also afford unique ways of determining protein structural information. In addition to analyses based on the quantification of residue variability in paralogs, two-way comparisons, both within and among species, help to disclose functional amino acids. Comparative studies of gene families throughout the mammalian genome will also help elucidate the functional significance of single nucleotide polymorphisms in coding regions.     

Interaction between calcium-free calmodulin and IQ motif of neurogranin studied by nuclear magnetic resonance spectroscopy

The interaction of Ca(2+)-free calmodulin (apoCaM) with the IQ motif corresponding to the calmodulin-binding domain of neurogranin has been studied by nuclear magnetic resonance (NMR) methods. The NMR spectra of uncomplexed apoCaM and apoCaM in complex with the IQ motif recorded at 750 MHz were studied and the backbone assignments of the protein in both forms were obtained by triple-resonance multidimensional NMR experiments. Chemical shift perturbations were used to map the binding surfaces. Only a single set of resonances was observed throughout the titration, indicating that the binding interaction is under fast exchange. Analysis of chemical shift changes indicates that (a) the main interaction and conformational changes occur in the C-terminal domain of calmodulin and (b) linker-1 (residues 40-44) between EF-1 and EF-2, linker-3 (residues 112-117) between EF-3 and EF-4, and the end of the alpha-helix H (residues 145-148) may be involved in the binding process. The dissociation constant (K(d)), estimated by fitting the chemical shift changes against the IQ peptide concentration, ranged from about 1.2 x 10(-5) to 8.8 x 10(-5) M. This result demonstrates that the interaction falls into the weak binding regime.     

A novel GTP-binding protein hGBP3 interacts with NIK/HGK

A novel human guanylate-binding protein (GBP) hGBP3 was identified and characterized. Similar as the two human guanylate-binding proteins hGBP1 and hGBP2, hGBP3 has the first two motifs of the three classical guanylate-binding motifs, GXXXXGKS (T) and DXXG, but lacks the N (T) KXD motif. Escherichia coli-expressed hGBP3 protein specifically binds to guanosine triphosphate (GTP). Using a yeast two-hybrid system, it was revealed that the N-terminal region of hGBP3 binds to the C-terminal regulatory domain of NIK/HGK, a member of the group I GCK (germinal center kinase) family. This interaction was confirmed by in vitro glutathione-S-transferase (GST) pull-down and co-immunoprecipitation assays.     

人脯氨肽酶活性中心结构的计算机模拟

氨酰基脯氨酸二肽酶 (脯氨肽酶 )为广泛分布于生物界的细胞内二肽水解酶 .它特异性地水解以脯氨酸或羟脯氨酸为羧基端的二肽 (X Pro) ,而且只对反式肽键有催化活性 .此酶与脯氨酸代谢、胶原蛋白合成及细胞生长有密切关系 .文献报道 ,从Alteromonas细菌中提取的脯氨肽酶有水解梭曼的活性 ,其有机磷酸酐水解酶也有脯氨肽酶活性 .用重组基因表达的人肝脯氨肽酶也同时具有脯氨肽酶活性和水解梭曼的活性 .研究脯氨肽酶活性中心的结构具有重要理论意义和潜在实用价值 .但目前尚无人脯氨肽酶晶体结构的报道 .本文采用蛋白质结构模式识别 (threading)方法对脯氨肽酶的高级结构进行模拟 ,以大肠杆菌甲硫氨酸氨肽酶 (1MAT)为模板 ,模建了人脯氨肽酶C端结构域的空间结构 .通过对模建结构的 3D评估及电荷分布分析 ,对人脯氨肽酶活力中心结构进行了预测 .模建的人脯氨肽酶活性中心位于C端结构域 ,为 6条β折叠围成的一个疏水性口袋 ,外面被 5条α螺旋及一些loop包围 ,活力中心位于疏水结构中央 ,其中有 5个保守氨基酸 ,形成 1个较强的负电荷区 ,周围有 3个较弱的正电荷区域 .实验还发现 ,虽然Mn2 + 或Co2 + 对酶的活性极其重要 ,但对酶蛋白结构的贡献很小 .提示它们可能是在催化反应的电荷转移过程中发挥着重要作用     

Enhancement in the viability and biosensing activity of freeze-dried recombinant bioluminescent bacteria

The genetically-engineeredEscherichia coli strain, DPD2540, which contains afabA::luxCDABE fusion gene, gives a bioluminescent output when membrane fatty acid synthesis is needed. For more practical application of this strain in the field as biosensor, freeze-drying was adopted. A 12% sucrose solution with Luria-Bertani (LB) broth, as determined by the viability after freeze-drying, was found to be the most effective composition for lyophilization solution among various compositions tested. Rapid freezing with liquid nitrogen also gave the best viability after freeze-drying as compared to samples frozen at −70°C and −20°C. The biosensing activities of the cells showed a greater sensitivity when the cells from the exponential phase were freeze-dried. Finally, the optimum temperature for use of the freeze-dried cells in the biosensor field was determined.     

电针对大鼠足底疼痛病灶所属脊髓节段一氧化氮合酶阳性反应的影响

采用NADPH-d组织化学方法,观察电针对大鼠一侧足底注射5%福尔马林50μl后病灶所属脊髓节段（L4-L6）一氧化氮合酶（NOS）阳性反应的影响。结果表明,福尔马林组大鼠L4-L6节段脊髓胶状质NOS阳性反应较生理盐水组显著增强,电针治疗组大鼠L4-L6节段脊髓胶状质NOS阳性反应与生理盐水组无显著差异。但明显弱于福尔马林组,提示电针治疗能抑制大鼠足底疼痛病灶所属脊髓节段NO合成,减弱脊髓神经元的敏感化,从而发挥镇痛作用。