世界生命科学前沿动态周报（五十四）

（8.22-8.28/2011）
美宝国际集团:陶国新  

    主要内容：广谱抗病毒方法；在体外单向应力诱导血管内皮细胞形成血管样结构；通过翻译后合成磷酸丝氨酸扩展大肠杆菌遗传密码；代谢综合症病人脂肪组织调节异常更易造成糖尿病和心血管疾病；突触小囊的分子构成有所不同；Hsp90-Cdc37伴侣复合物调节线粒体自吞噬。

焦点动态：广谱抗病毒方法。

1. 广谱抗病毒方法
【动态】
目前的抗病毒药物还不多，而且大部分现有药物是高度病原体特异性的，或者有其他缺陷。美国MIT的科学家研发了一种广谱抗病毒方法，称为双链RNA激活的切冬酶寡聚体(DRACO)，能够选择性诱导含有病毒双链RNA的细胞进行凋亡，快速消灭被感染的细胞而不影响未感染细胞。他们制造了DRACO并证明对11种哺乳动物细胞是无毒的，能够对抗15种不同病毒，包括登革热病毒和H1N1流感病毒等。他们也证明了DRACO能够挽救感染H1N1流感病毒的老鼠。因为双链RNA检测域的广谱的敏感性、凋亡诱导区域的高活性，以及病毒从未遭遇过的二者之间新的直接联系，DRACO有潜力成为多种临床主要病毒的治疗或预防手段。

【点评】
选择性的广谱的清除被病毒感染的细胞是不错的抗病毒策略。配合清除病毒的手段一起使用很有潜力成为临床治疗病毒感染的有效途径。

【参考论文】
PLoS ONE 2011, 6(7): e22572. doi:10.1371/journal.pone.0022572
Broad-Spectrum Antiviral Therapeutics
Rider TH, Zook CE, Boettcher TL, et al.
Currently there are relatively few antiviral therapeutics, and most which do exist are highly pathogen-specific or have other disadvantages. We have developed a new broad-spectrum antiviral approach, dubbed Double-stranded RNA (dsRNA) Activated Caspase Oligomerizer (DRACO) that selectively induces apoptosis in cells containing viral dsRNA, rapidly killing infected cells without harming uninfected cells. We have created DRACOs and shown that they are nontoxic in 11 mammalian cell types and effective against 15 different viruses, including dengue flavivirus, Amapari and Tacaribe arenaviruses, Guama bunyavirus, and H1N1 influenza. We have also demonstrated that DRACOs can rescue mice challenged with H1N1 influenza. DRACOs have the potential to be effective therapeutics or prophylactics for numerous clinical and priority viruses, due to the broad-spectrum sensitivity of the dsRNA detection domain, the potent activity of the apoptosis induction domain, and the novel direct linkage between the two which viruses have never encountered.

2. 在体外单向应力诱导血管内皮细胞形成血管样结构
【动态】
骨骼肌组织工程带给那些由于外伤等损失肌肉的患者以再生治疗的很大希望，但是推进到临床应用还面临不少挑战。荷兰的科学家报道了对于制造协调一致的预先供血的肌肉组织的研究。他们假设协调一致的有血供的三维结构肌肉组织可以只通过单向应力在体外诱导生成。他们的研究显示内皮细胞和肌肉细胞不仅可以独立地在水凝胶基质的三维培养体系中沿着单向应力的方向排列，而且更重要的是，在共培养的三维架构中内皮细胞组织成血管结构。引人注目的是，这些培养体系里不需要添加另外的生长因子就能够诱导内皮细胞形成血管。肌肉组织沿着一个方向收缩时形成的单向应力刺激肌肉细胞生产VEGF。这一伴随有VEGF生产的单向应力似乎在中起关键作用。

【点评】
这一研究表明机械力的影响和调节是骨骼肌发育的重要因素。

【参考论文】
Tissue Engineering Part A, 2011; 110811095013006 DOI: 10.1089/ten.tea.2011.0214 Mechanoregulation of Vascularization in Aligned Tissue-Engineered Muscle: A Role for Vascular Endothelial Growth Factor
Daisy W.J. van der Schaft, Ariane C.C. van Spreeuwel, Hans C. van Assen, et al.
Skeletal muscle tissue engineering has major promise for regenerative treatment of patients suffering from muscle loss due to, for example, traumatic injury, but faces considerable challenges to progress toward clinical application. In the present study the creation of an aligned prevascularized muscle tissue was addressed. We hypothesized that an aligned vascularized three-dimensional (3D) muscle tissue can be induced in vitro by merely using uniaxial stress. The present study showed that not only do endothelial cells and muscle cells independently align in the direction of uniaxial stress in a hydrogel-based 3D culture system, but also, more importantly, the endothelial cells in the co-cultured 3D constructs organized into vascular structures. Strikingly, in these cultures no additional growth factors were needed to induce vascular formation of the endothelial cells. Vascular endothelial growth factor (VEGF) production by the muscle cells was stimulated by the uniaxial stress that develops in the tissue when constrained in one direction. This stress accompanied by VEGF production appeared to play a key role in the organization of the endothelial cells into vessel-like structures.

3. 通过翻译后合成磷酸丝氨酸扩展大肠杆菌遗传密码
【动态】
O-磷酸丝氨酸是真核细胞磷酸化蛋白组中含量最多的磷酸化氨基酸（Sep），不在遗传密码中编码，而是在翻译后合成。美国科学家报道了一个工程体系，通过拥有tRNASep、SepRS和设计制造的EF-Tu （EF-Sep）的大肠杆菌变种将丝氨酸特异性的共翻译整合到蛋白的任意位置。依赖工程再造EF-Tu来放松其质量控制功能允许Sep-tRNASep的结合来扩展遗传密码。

【点评】
该生物工程体系提供了一种有效的特定位置的蛋白磷酸化手段。

【参考论文】
Science, 2011; 333 (6046): 1151-1154 DOI: 10.1126/science.1207203
Expanding the Genetic Code of Escherichia coli with Phosphoserine
Hee-Sung Park, Michael J. Hohn, Takuya Umehara, et al. 
O-Phosphoserine (Sep), the most abundant phosphoamino acid in the eukaryotic phosphoproteome, is not encoded in the genetic code, but synthesized posttranslationally. Here, we present an engineered system for specific cotranslational Sep incorporation (directed by UAG) into any desired position in a protein by an Escherichia coli strain that harbors a Sep-accepting transfer RNA (tRNASep), its cognate Sep–tRNA synthetase (SepRS), and an engineered EF-Tu (EF-Sep). Expanding the genetic code rested on reengineering EF-Tu to relax its quality-control function and permit Sep-tRNASep binding. To test our system, we synthesized the activated form of human mitogen-activated ERK activating kinase 1 (MEK1) with either one or two Sep residues cotranslationally inserted in their canonical positions (Sep218, Sep222). This system has general utility in protein engineering, molecular biology, and disease research.

4. 代谢综合症病人脂肪组织调节异常更易造成糖尿病和心血管疾病
【动态】
代谢综合症与更高的糖尿病及心血管疾病风险有联系。已有很多研究表明在代谢综合症中循环中的炎症生物标记物增多，但是很少数据关于造成这一低度炎症的细胞来源。美国科学家研究了SC脂肪组织生物学在尚未伴有糖尿病及心血管疾病的早期代谢综合症中的作用。经过知情同意后招募代谢综合症病人和对照组受试者，采集空腹血，活检采集SC脂肪组织。相比对照组，代谢综合症病人的炎症和胰岛素抗性血液生物标记物、hsCRP、IL-6, IL-1β, 瘦素（leptin）, 血清淀粉样蛋白A和维生素A结合蛋白4 (RBP-4) 的浓度明显更高。代谢综合症病人的SC脂肪组织中，leptin, RBP-4, CRP, 血清淀粉样蛋白A, 纤溶酶原激活物抑制剂-1，IL-1, IL-6, IL-8, 和单核细胞趋化蛋白-1水平明显高于对照组。校正腰围因素后，除RBP-4外其余差异依然存在。另外，相对于对照组，代谢综合症病人的SC脂肪组织巨噬细胞浸润明显增多，花冠样结构增多。hsCRP与体内平衡模型评价和SC脂肪组织单核细胞趋化蛋白-1正相关，与脂联素负相关。体内平衡模型评价与纤溶酶原激活物抑制剂-1、RBP-4和SC脂肪组织单核细胞趋化蛋白-1正相关。他们新发现了代谢综合症病人的SC脂肪组织具有更多的花冠样结构特征的巨噬细胞，并造成更多细胞炎症产生更多与胰岛素抗性和低度炎症相关的生物学标记物。这一偏差可能造成代谢综合症的进展和更高风险的糖尿病及心血管疾病。

【点评】
该研究证明患有代谢综合症的肥胖病人患糖尿病及心血管疾病的风险更高。

【参考论文】
The Journal of Clinical Endocrinology & Metabolism, 2011 DOI: 10.1210/jc.2011-1577
Adipose Tissue Dysregulation in Patients with Metabolic Syndrome
Andrew A. Bremer, Sridevi Devaraj, Alaa Afify, and Ishwarlal Jialal.
Context: The metabolic syndrome (MetS) is associated with increased risk of diabetes and cardiovascular disease (CVD). Numerous groups have shown increased circulating biomarkers of inflammation in MetS. However, there are scanty data on the cellular sources contributing to this low-grade inflammation.
Objective: The aim of this study was to determine the role of sc adipose tissue (SAT) biology in nascent MetS without concomitant diabetes or CVD.
Patients and Methods: Subjects with MetS and controls were recruited after informed consent. Fasting blood was collected, and SAT was obtained by biopsy.
Results: Circulating biomarkers of inflammation and insulin resistance, high-sensitivity C-reactive protein (hsCRP), IL-6, IL-1β, leptin, serum amyloid A, and retinol-binding protein-4 (RBP-4) concentrations were significantly higher in the MetS subjects than controls, whereas adiponectin concentrations were lower. In SAT, leptin, RBP-4, CRP, serum amyloid A, plasminogen activator inhibitor-1, IL-1, IL-6, IL-8, and monocyte chemotactic protein-1 (MCP-1) levels were significantly higher in MetS than controls. These differences except for RBP-4 persisted after adjusting for waist circumference. In addition, there were significantly increased numbers of macrophages infiltrating the SAT of MetS and increased numbers of crown-like structures compared with controls. hsCRP correlated positively with homeostasis model assessment and SAT MCP-1 and negatively with adiponectin. Homeostasis model assessment correlated positively with plasminogen activator inhibitor-1, RBP-4, and SAT MCP-1.
Conclusions: We make the novel observation that SAT of MetS has increased macrophage recruitment with cardinal crown-like structure features and contributes to the increased cellular inflammation that produces increased levels of biomarkers that are correlated with both insulin resistance and low-grade inflammation. These aberrations could contribute to the progression of MetS and the increased risk for diabetes and CVD.

5. 突触小囊的分子构成有所不同
【动态】
突触小囊属于两个不同的储蓄池，负责唤起的神经递质释放的循环池和对刺激无反应的休眠池。突触小囊统一的外观暗示其位置和与细胞骨架联系的不同解释它们功能上的差别。美国科学家最近发现v-SNARE破伤风毒素不敏感小囊相关的膜蛋白(VAMP7)与其他突触小囊蛋白在两个池中的分布不同，证明它们的分子组成不同。他们还发现循环池和休眠池会同时释放，当去除longin区域来活化时，VAMP7影响了释放性能。更进一步，随着唤起的自然释放的内吞作用机制有所不同，特定序列给予了针对不同小囊池的靶标。研究结果提示不同内吞机制产生的突触小囊具有不同蛋白赋予小囊不同特性。

【点评】
该研究对于进一步深入了解神经递质的释放和摄取有很大帮助。

【参考论文】
Neuron, 2011; 71 (3): 474 DOI:10.1016/j.neuron.2011.06.010
v-SNARE Composition Distinguishes Synaptic Vesicle Pools
Zhaolin Hua, Sergio Leal-Ortiz, Sarah M. Foss, et al. 
Synaptic vesicles belong to two distinct pools, a recycling pool responsible for the evoked release of neurotransmitter and a resting pool unresponsive to stimulation. The uniform appearance of synaptic vesicles has suggested that differences in location or cytoskeletal association account for these differences in function. We now find that the v-SNARE tetanus toxin-insensitive vesicle-associated membrane protein (VAMP7) differs from other synaptic vesicle proteins in its distribution to the two pools, providing evidence that they differ in molecular composition. We also find that both resting and recycling pools undergo spontaneous release, and when activated by deletion of the longin domain, VAMP7 influences the properties of release. Further, the endocytosis that follows evoked and spontaneous release differs in mechanism, and specific sequences confer targeting to the different vesicle pools. The results suggest that different endocytic mechanisms generate synaptic vesicles with different proteins that can endow the vesicles with distinct properties.

6. Hsp90-Cdc37伴侣复合物调节线粒体自吞噬
【动态】
自吞噬，细胞的主要回收利用再循环途径，对于在正常生长条件下和细胞应激时保证线粒体的质量起关键作用。Hsp90-Cdc37伴侣复合物协调调节所选激酶的活性来指挥应激反应的多个方面。尽管二者都维持线粒体完整性，Hsp90-Cdc37和自吞噬之间的关系一直没有搞清楚。Ulk1，酵母菌Atg1的哺乳类同系物之一，是自吞噬所需的丝氨酸- 苏氨酸激酶。美国科学家的最新研究显示Ulk1 和Hsp90-Cdc37之间的相互作用稳定和激活Ulk1 , 这又是磷酸化Atg13和从Ulk1释放Atg13，以及招募Atg13到受损伤的线粒体所必须。Hsp90-Cdc37, Ulk1, 以及Atg13磷酸化都是有效清除线粒体所必须的。 这些发现建立了一种直接的方法整合Ulk1- 和 Atg13-指导的自吞噬和Hsp90 与Cdc37协调的应激反应。

【点评】
该研究可以帮助我们认识线粒体的自吞噬机制。

【参考论文】
Molecular Cell, 2011; 43 (4): 572 DOI: 10.1016/j.molcel.2011.06.018
Hsp90-Cdc37 Chaperone Complex Regulates Ulk1- and Atg13-Mediated Mitophagy
Joung Hyuck Joo, Frank C. Dorsey, Aashish Joshi, et al.
Autophagy, the primary recycling pathway of cells, plays a critical role in mitochondrial quality control under normal growth conditions and in the response to cellular stress. The Hsp90-Cdc37 chaperone complex coordinately regulates the activity of select kinases to orchestrate many facets of the stress response. Although both maintain mitochondrial integrity, the relationship between Hsp90-Cdc37 and autophagy has not been well characterized. Ulk1, one of the mammalian homologs of yeast Atg1, is a serine-threonine kinase required for mitophagy. Here we show that the interaction between Ulk1 and Hsp90-Cdc37 stabilizes and activates Ulk1, which in turn is required for the phosphorylation and release of Atg13 from Ulk1, and for the recruitment of Atg13 to damaged mitochondria. Hsp90-Cdc37, Ulk1, and Atg13 phosphorylation are all required for efficient mitochondrial clearance. These findings establish a direct pathway that integrates Ulk1- and Atg13-directed mitophagy with the stress response coordinated by Hsp90 and Cdc37.