世界生命科学前沿动态周报（七十）

（12.12-12.18/2011）
美宝国际集团:陶国新 

主要内容：特别的GATA因子是内胚层上皮-间质转化的保守诱导物；影响疤痕形成的物理因素；蛋白亚磺酰化在细胞信号通路中的重要调节作用；HIV病毒进入细胞核的“钥匙”；确定造血干细胞的起源；葡聚糖水凝胶对三度烧伤的治疗效果。

焦点动态：影响疤痕形成的物理因素。

1. 特别的GATA因子是内胚层上皮-间质转化的保守诱导物
【动态】
上皮-间质转化（EMT）使静止的上皮细胞转变为有移动能力的间充质细胞状态，西班牙科学家最近证明果蝇内胚层的EMT依赖GATA因子Srp。当Srp异位激活后作用类似于高效的EMT触发剂。Srp通过下调但不阻断联接蛋白dE钙粘素（dE-Cad）而影响内胚层的EMT。而且，Srp通过直接抑制crumbs(crb)基因重定位dE-Cad。他们的研究还显示Srp的直接同源物人GATA-6在哺乳动物细胞中诱导类似的转变。类似于Srp，人GATA-6通过下调但不阻断E-Cad起作用，并诱导抑制Crumbs 的直接同源物crb2。总的看来，他们的研究发现一套在发育学和病理学中GATA因子是选择性的保守的触发物抑制上皮细胞的上皮特性而赋予其迁移能力。

【点评】
该研究发现上皮-间质转化依赖GATA因子，对于深入了解生物发育以及肿瘤发展有推动作用。

【参考论文】
Developmental Cell, 2011; 21 (6): 1051 DOI: 10.1016/j.devcel.2011.10.005
Specific GATA Factors Act as Conserved Inducers of an Endodermal-EMT
The epithelial-to-mesenchymal transition (EMT) converts cells from static epithelial to migratory mesenchymal states (Hay, 1995 ). Here, we demonstrate that EMT in the Drosophila endoderm is dependent on the GATA-factor Serpent (Srp), and that Srp acts as a potent trigger for this transition when activated ectopically. We show that Srp affects endodermal-EMT through a downregulation of junctional dE-Cadherin (dE-Cad) protein, without a block in its transcription. Moreover, the relocalization of dE-Cad is achieved through the direct repression of crumbs (crb) by Srp. Finally, we show that hGATA-6, an ortholog of Srp, induces a similar transition in mammalian cells. Similar to Srp, hGATA-6 acts through the downregulation of junctional E-Cad, without blocking its transcription, and induces the repression of a Crumbs ortholog, crb2. Together, these results identify a set of GATA factors as a conserved alternative trigger to repress epithelial characteristics and confer migratory capabilities on epithelial cells in development and pathogenesis.

2. 影响疤痕形成的物理因素
【动态】
纤维增生旺盛是受伤后常见的并发症，原因尚不明了。一个常被忽视的创伤修复的关键因素是机械力，机械力通过细胞内包括焦点粘连激酶（FAK）在内的焦点粘连成分调节细胞-基质相互作用。美国斯坦福大学的科学家最近报道了皮肤损伤后FAK被激活，这一过程被机械力负载所加强。在疤痕增生动物模型中，敲除成纤维细胞特异性的FAK的老鼠明显比对照老鼠炎症和纤维化都少。他们发现FAK通过细胞外相关激酶（ERK）物理性的触发单核细胞化学引诱物-1（MCP-1，也叫CCL2）的分泌，这是一种与人体纤维病变有关的高效趋化因子。类似地，敲除MCP-1的老鼠形成的疤痕最小，意味着炎症趋化因子途径是FAK通过力传导诱导纤维化的主要机理。用小分子抑制FAK能够在人体细胞中阻断这些作用，并且通过调节MCP-1信号和炎症细胞的招募而减少实验动物的疤痕形成。这些发现合在一起表明机械力通过炎症FAK–ERK–MCP-1途径调节纤维化以及针对FAK的分子策略能够有效的解除机械力与病理疤痕形成的关联。

【点评】
该研究阐明了机械力可以通过增强炎症反应诱导组织纤维化增生。解除机械力的影响能够改善纤维化状况。

【参考论文】
Nature Medicine, 2011; DOI: 10.1038/nm.2574
Focal adhesion kinase links mechanical force to skin fibrosis via inflammatory signaling
Victor W Wong, Kristine C Rustad, Satoshi Akaishi, et al.
Exuberant fibroproliferation is a common complication after injury for reasons that are not well understood. One key component of wound repair that is often overlooked is mechanical force, which regulates cell-matrix interactions through intracellular focal adhesion components, including focal adhesion kinase (FAK). Here we report that FAK is activated after cutaneous injury and that this process is potentiated by mechanical loading. Fibroblast-specific FAK knockout mice have substantially less inflammation and fibrosis than control mice in a model of hypertrophic scar formation. We show that FAK acts through extracellular-related kinase (ERK) to mechanically trigger the secretion of monocyte chemoattractant protein-1 (MCP-1, also known as CCL2), a potent chemokine that is linked to human fibrotic disorders. Similarly, MCP-1 knockout mice form minimal scars, indicating that inflammatory chemokine pathways are a major mechanism by which FAK mechanotransduction induces fibrosis. Small-molecule inhibition of FAK blocks these effects in human cells and reduces scar formation in vivo through attenuated MCP-1 signaling and inflammatory cell recruitment. These findings collectively indicate that physical force regulates fibrosis through inflammatory FAK–ERK–MCP-1 pathways and that molecular strategies targeting FAK can effectively uncouple mechanical force from pathologic scar formation.

3. 蛋白亚磺酰化在细胞信号通路中的重要调节作用
【动态】
蛋白亚磺酰化是一种翻译后修饰，在高等真核生物中逐渐显示出其重要性。但是，研究其多样性的作用还是很有挑战性的，尤其是在天然的细胞环境内。美国科学家最近开发利用DYn-2，一种新的化学选择性探针检测人体细胞内的亚磺酰化的蛋白。他们的研究表明表皮生长因子受体介导的信号导致过氧化氢的产生和下游蛋白的氧化。另外，他们还证明DYn-2能够细胞内亚磺酰化率的差异，这种差异与靶蛋白的定位差异有关。他们还发现过氧化氢在表皮生长因子受体的关键活性位点半胱氨酸（Cys797）进行直接修饰能够增强其酪氨酸激酶活性。总起来看，他们的发现显示亚磺酰化是类似于磷酸化的全局性信号机制，牵涉到其他受体酪氨酸激酶和针对蛋白中氧化敏感的半胱氨酸的不可逆抑制剂。
【点评】
蛋白亚磺酰化在细胞信号调节中作用的提高，会促进以调节蛋白亚磺酰化为目标的药物研究和开发。但是更重要的意义在于，这表明蛋白翻译后修饰有多种方式都很重要，生命自身的调节远比我们已知的更为复杂巧妙。

【参考论文】
Nature Chemical Biology, 11 December 2011 DOI: 10.1038/nchembio.736
Peroxide-dependent sulfenylation of the EGFR catalytic site enhances kinase activity
Candice E Paulsen, Thu H Truong, Francisco J Garcia, et al.
Protein sulfenylation is a post-translational modification of emerging importance in higher eukaryotes. However, investigation of its diverse roles remains challenging, particularly within a native cellular environment. Herein we report the development and application of DYn-2, a new chemoselective probe for detecting sulfenylated proteins in human cells. These studies show that epidermal growth factor receptor–mediated signaling results in H2O2 production and oxidation of downstream proteins. In addition, we demonstrate that DYn-2 has the ability to detect differences in sulfenylation rates within the cell, which are associated with differences in target protein localization. We also show that the direct modification of epidermal growth factor receptor by H2O2 at a critical active site cysteine (Cys797) enhances its tyrosine kinase activity. Collectively, our findings reveal sulfenylation as a global signaling mechanism that is akin to phosphorylation and has regulatory implications for other receptor tyrosine kinases and irreversible inhibitors that target oxidant-sensitive cysteines in proteins.

4. HIV病毒进入细胞核的“钥匙”
【动态】
慢病毒像HIV-1横穿核膜孔复合物（NPC）并感染终末分化的不分裂细胞，它们如何做到的还不清楚。以前的研究已发现胞浆NPC蛋白Nup358/RaBP2是HIV-1辅因子。最近英国和美国的科学家报道HIV-1病毒壳（CA）直接结合到Nup358/RaBP2的亲环素（cyclophilin, Cyp）结构域。该Cyp与三重TRIM5的融合产生了一种新的HIV-1复制抑制剂，与体内的一种相互作用一致。与CypA结合到HIV-1 CA相反，Nup358的结合对环孢霉素的抑制不敏感，借此可以区分CypA和Nup358的影响。抑制CypA减少了对Nup358和核篮蛋白Nup153的依赖，表明CypA调节病毒参与的核内转运机制的选择。相比野生型病毒，HIV-1 病毒壳的Cyp结合突变G89V和P90A在高密度转录单位的基因区域有更多整合和相关特征。野生型病毒在环孢霉素存在时的整合倾向性与高密度转录区域有类似的改变。相反，HIV-1 CA在使得病毒对Nup358 或 TRN-SR2 删除（CA N74D, N57A）更不敏感的壳表面另一区域的改变导致整合到少有转录单位的基因区域。两组CA突变在HeLa细胞和人单核细胞源巨噬细胞的复制中受到损害。他们的发现将HIV-1衔接亲环素与整合目标和复制效率联系起来，并对病毒亲环素招募的保守性提供了新见解。

【点评】
该研究阐明了艾滋病毒HIV-1与亲环素的相互作用决定了往核内转运的途径、整合的目标以及复制的效率。对于开发新的抗艾滋病药物会有推动作用。

【参考论文】
PLoS Pathogens, 2011; 7 (12): e1002439 DOI: 10.1371/journal.ppat.1002439 
HIV-1 Capsid-Cyclophilin Interactions Determine Nuclear Import Pathway, Integration Targeting and Replication Efficiency
Torsten Schaller, Karen E. Ocwieja, Jane Rasaiyaah, et al.
Lentiviruses such as HIV-1 traverse nuclear pore complexes (NPC) and infect terminally differentiated non-dividing cells, but how they do this is unclear. The cytoplasmic NPC protein Nup358/RanBP2 was identified as an HIV-1 co-factor in previous studies. Here we report that HIV-1 capsid (CA) binds directly to the cyclophilin domain of Nup358/RanBP2. Fusion of the Nup358/RanBP2 cyclophilin (Cyp) domain to the tripartite motif of TRIM5 created a novel inhibitor of HIV-1 replication, consistent with an interaction in vivo. In contrast to CypA binding to HIV-1 CA, Nup358 binding is insensitive to inhibition with cyclosporine, allowing contributions from CypA and Nup358 to be distinguished. Inhibition of CypA reduced dependence on Nup358 and the nuclear basket protein Nup153, suggesting that CypA regulates the choice of the nuclear import machinery that is engaged by the virus. HIV-1 cyclophilin-binding mutants CA G89V and P90A favored integration in genomic regions with a higher density of transcription units and associated features than wild type virus. Integration preference of wild type virus in the presence of cyclosporine was similarly altered to regions of higher transcription density. In contrast, HIV-1 CA alterations in another patch on the capsid surface that render the virus less sensitive to Nup358 or TRN-SR2 depletion (CA N74D, N57A) resulted in integration in genomic regions sparse in transcription units. Both groups of CA mutants are impaired in replication in HeLa cells and human monocyte derived macrophages. Our findings link HIV-1 engagement of cyclophilins with both integration targeting and replication efficiency and provide insight into the conservation of viral cyclophilin recruitment.

5. 确定造血干细胞的起源
【动态】
造血干细胞（HSCs）和更早一波的限定性的网织红细胞/骨髓祖细胞（EMPs）在孕体生血内皮细胞时区分开来。从胚胎干细胞或诱导多能干细胞能够体外生产EMPs，但生产HSCs的努力大多失败了。EMPS和HSCs的形成都需要转录因子Runx1及其非DNA-结合伴侣核结合因子β (CBFβ)。美国科学家最近的研究显示孕体中EMP和HSC形成中对CBFβ的需要在时间和空间上是不同的。在表达Tek的细胞中CBFβ的泛内皮表达对形成EMP已经足够，但对HSC形成却还不够。另一方面，在表达Ly6a的细胞中，CBFβ的表达对形成HSC足够但对形成EMP还不够。其数据表明EMPs和HSCs是从生血内皮细胞的不同群分化来的，而Ly6a特异性的标记产生HSC的生血内皮。
【点评】
该研究比较精确地确定了造血干细胞的前体细胞并发现了其特异性标志Ly6a。

【参考论文】
Cell Stem Cell, 2011; 9 (6): 541 DOI: 10.1016/j.stem.2011.10.003 
Erythroid/Myeloid Progenitors and Hematopoietic Stem Cells Originate from Distinct Populations of Endothelial Cells
Michael J. Chen, Yan Li, Maria Elena De Obaldia, et al.
Hematopoietic stem cells (HSCs) and an earlier wave of definitive erythroid/myeloid progenitors (EMPs) differentiate from hemogenic endothelial cells in the conceptus. EMPs can be generated in vitro from embryonic or induced pluripotent stem cells, but efforts to produce HSCs have largely failed. The formation of both EMPs and HSCs requires the transcription factor Runx1 and its non-DNA binding partner core binding factor β (CBFβ). Here we show that the requirements for CBFβ in EMP and HSC formation in the conceptus are temporally and spatially distinct. Panendothelial expression of CBFβ in Tek-expressing cells was sufficient for EMP formation, but was not adequate for HSC formation. Expression of CBFβ in Ly6a-expressing cells, on the other hand, was sufficient for HSC, but not EMP, formation. The data indicate that EMPs and HSCs differentiate from distinct populations of hemogenic endothelial cells, with Ly6a expression specifically marking the HSC-generating hemogenic endothelium.

6. 葡聚糖水凝胶对三度烧伤的治疗效果
【动态】
对深度烧伤的创伤愈合效果而言血管新生是个关键的决定因素。美国约翰霍普金斯医学院的科学家认为基于葡聚糖的水凝胶能够作为引导性平台促进3度烧伤的血管新生和皮肤再生。葡聚糖水凝胶软而韧，有机会提高烧伤治疗效果。他们首先制定了一套用葡聚糖水凝胶治疗老鼠烧伤的程序，其中遵循临床规范削除全厚层烧伤皮肤，然后用葡聚糖水凝胶和敷料层覆盖伤处。该程序能保证整个愈合期间水凝胶能够完好无损固定在伤处以简化烧伤治疗的处理。一项3周的对照研究表明葡聚糖水凝胶促进了带有完整附件的真皮再生。水凝胶平台促进了早期炎症细胞浸润导致其快速降解，促进了生血管细胞向愈合中伤处的浸润。内皮细胞进入水凝胶平台在第7日开始能够新生血管，使得血流比治疗和不治疗的对照组显著增加。到第21天，用水凝胶治疗的烧伤处产生成熟的上皮结构有毛囊和皮脂腺。治疗5周后，水凝胶促进了头发的新生，并且表皮形态和厚度与正常老鼠皮肤相似。总的看来，他们的证据显示定制的单独使用葡聚糖水凝胶，不加任何生长因子、细胞因子或细胞，能够促进明显的血管新生和皮肤再生，并可能引出新的皮肤创伤治疗方法。

【点评】
该研究发现在烧伤老鼠削痂后单独使用葡聚糖水凝胶能够促进血管新生和正常皮肤的再生。

【参考论文】
PNAS December 27, 2011 vol. 108 no. 52 20976-20981
Dextran hydrogel scaffolds enhance angiogenic responses and promote complete skin regeneration during burn wound healing
Guoming Suna, Xianjie Zhangb, Yu-I Shena, et al.

Neovascularization is a critical determinant of wound-healing outcomes for deep burn injuries. We hypothesize that dextran-based hydrogels can serve as instructive scaffolds to promote neovascularization and skin regeneration in third-degree burn wounds. Dextran hydrogels are soft and pliable, offering opportunities to improve the management of burn wound treatment. We first developed a procedure to treat burn wounds on mice with dextran hydrogels. In this procedure, we followed clinical practice of wound excision to remove full-thickness burned skin, and then covered the wound with the dextran hydrogel and a dressing layer. Our procedure allows the hydrogel to remain intact and securely in place during the entire healing period, thus offering opportunities to simplify the management of burn wound treatment. A 3-week comparative study indicated that dextran hydrogel promoted dermal regeneration with complete skin appendages. The hydrogel scaffold facilitated early inflammatory cell infiltration that led to its rapid degradation, promoting the infiltration of angiogenic cells into the healing wounds. Endothelial cells homed into the hydrogel scaffolds to enable neovascularization by day 7, resulting in an increased blood flow significantly greater than treated and untreated controls. By day 21, burn wounds treated with hydrogel developed a mature epithelial structure with hair follicles and sebaceous glands. After 5 weeks of treatment, the hydrogel scaffolds promoted new hair growth and epidermal morphology and thickness similar to normal mouse skin. Collectively, our evidence shows that customized dextran-based hydrogel alone, with no additional growth factors, cytokines, or cells, promoted remarkable neovascularization and skin regeneration and may lead to novel treatments for dermal wounds.