·Nature导读·

解析花蜜的进化之谜

植物生产和分泌花蜜的过程仍然未知。研究人员对三种不同的开花植物(拟南芥、芜菁和野生烟草)的研究发现，糖转运蛋白SWEET9在三种植物中负责介导花蜜分泌的过程，将糖转运到蜜腺的细胞外区域，是花蜜合成和分泌中的关键元件。SWEET9还可能对于开花植物的进化至关重要，出现在开花植物进化的早期。

论文链接:Lin I W，et al..Nectar secretion requires sucrose phosphate synthases and the sugar transporter SWEET9.

Angiosperms developed floral nectaries that reward pollinating insects.Although nectar function and composition have been characterized，the mechanism of nectar secretion has remained unclear.Here we identify SWEET9 as a nectary-specific sugar transporter in three eudicot species:Arabidopsis thaliana，Brassica rapa(extrastaminal nectaries)and Nicotiana attenuata(gynoecial nectaries).We show that SWEET9 is essential for nectar production and can function as an efflux transporter.We also show that sucrose phosphate synthase genes，encoding key enzymes for sucrose biosynthesis，are highly expressed in nectaries and that their expression is also essential for nectar secretion.Together these data are consistent with a model in which sucrose is synthesized in the nectary parenchyma and subsequently secreted into the extracellular space via SWEET9，where sucrose is hydrolysed by an apoplasmic invertase to produce a mixture of sucrose，glucose and fructose.The recruitment of SWEET9 for sucrose export may have been a key innovation，and could have coincided with the evolution of core eudicots and contributed to the evolution of nectar secretion to reward pollinators.

蝴蝶翅膀图案多样化的基因控制

蝴蝶翅膀的拟态是自然选择的一种典型范例。有证据表明，燕尾蝶基因组中存在一个控制拟态的区域，鉴于拟态的复杂性，通常认为蝴蝶翅膀的拟态图案应该是多个基因共同作用的结果，而该研究发现，控制性别分化的doublesex基因单独承担了这项工作。单个基因能够控制如此复杂的机能的原因可能在于控制doublesex表达时机和地点的非编码DNA可能起到了重要作用，并且，doublesex是一个可激活其他基因的转录因子，其差异性可能是翅膀图案多样化的原因。

论文链接:Kunte K，et al..doublesex is a mimicry supergene.

Nature，doi:10.1038/nature13112.Published online:05 March，2014.Abstract:One of the most striking examples of sexual dimorphism is sex-limited mimicry in butterflies，a phenomenon in which one sex—usually the female—mimics a toxic model species，whereas the other sex displays a different wing pattern1.Sex-limited mimicry is phylogenetically widespread in the swallowtail butterfly genus Papilio，in which it is often associated with female mimetic polymorphism.In multiple polymorphic species，the entire wing pattern phenotype is controlled by a single Mendelian‘supergene’.Although theoretical work has explored the evolutionary dynamics of supergene mimicry，there are almost no empirical data that address the critical issue of what a mimicry supergene actually is at a functional level.Using an integrative approach combining genetic and association mapping，transcriptome and genome sequencing，and gene expression analyses，we show that a single gene，doublesex，controls supergene mimicry in Papilio polytes.This is in contrast to the long-held view that supergenes are likely to be controlled by a tightly linked cluster of loci.Analysis of gene expression and DNA sequence variation indicates that isoform expression differences contribute to the functional differences between dsx mimicry alleles，and protein sequence evolution may also have a role.Our results combine elements from different hypotheses for the identity of supergenes，showing that a single gene can switch the entire wing pattern among mimicry phenotypes but may require multiple，tightly linked mutations to do so.

细菌IV型分泌系统与细菌抗生素耐药性

细菌IV型分泌系统能够在细胞间分配遗传物质，如，抗生素抗性基因，该机制直接导致了抗生素耐药性传播。它还在引发溃疡、百日咳、军团病一类严重肺炎的感染毒素分泌过程起至关重要的作用。研究人员重建了大肠杆菌的IV型分泌系统，从分子结构和分泌机制等方面揭示出IV型分泌系统与其他的细菌分泌系统的显著差异。

论文链接:Low H H，et al..Structure of a type IV secretion system.

Nature，doi:10.1038/nature13081.Published online:09 March，2014.

Abstract:Bacterial type IV secretion systems translocate virulence factors into eukaryotic cells，distribute genetic material between bacteria and have shown potential as a tool for the genetic modification of human cells.Given the complex choreography of the substrate through the secretion apparatus，the molecular mechanism of the type IV secretion system has proved difficult to dissect in the absence of structural data for the entire machinery.Here we use electron microscopy to reconstruct the type IV secretion system encoded by the Escherichia coli R388 conjugative plasmid.We show that eight proteins assemble in an intricate stoichiometric relationship to form an approximately 3 megadalton nanomachine that spans the entire cell envelope.The structure comprises an outer membrane-associated core complex connected by a central stalk to a substantial inner membrane complex that is dominated by a battery of 12 VirB4 ATPase subunits organized as side-by-side hexameric barrels.Our results show a secretion system with markedly different architecture，and consequently mechanism，to other known bacterial secretion systems.

RNAi通路的关键酶在细菌中起防止DNA入侵的作用

RNA干涉(RNAi)在真核细胞中可以通过一个过程来保护细胞不受外来“小单链RNA”(ssRNAs)的影响。在这个过程中，被主体编码的短RNA结合同源RNA目标，并介导它们的降解。Argonaute(Ago)是真核细胞中由RNA引导的RNAi通路的一个关键酶。研究发现，原核生物的Ago(来自细菌Thermus thermophilus)保护细胞不受DNA而不是RNA入侵。在本研究中，Ago被加载了miRNA(与来自质体DNA的miRNA相似)，后者结合并掀开互补的DNA链。

论文链接:Swarts D C，et al..DNA-guided DNA interference by a prokaryotic Argonaute.

Nature，2014，507(7491)，258 －261.doi:10.1038/nature12971.

Abstract:RNA interference is widely distributed in eukaryotes and has a variety of functions，including antiviral defence and gene regulation.All RNA interference pathways use small single-stranded RNA(ssRNA)molecules that guide proteins of the Argonaute(Ago)family to complementary ssRNA targets:RNA-guided RNA interference.The role of prokaryotic Ago variants has remained elusive，although bioinformatics analysis has suggested their involvement in host defence.Here we demonstrate that Ago of the bacterium Thermus thermophilus(TtAgo)acts as a barrier for the uptake and propagation of foreign DNA.In vivo，TtAgo is loaded with 5'-phosphorylated DNA guides，13～25 nucleotides in length，that are mostly plasmid derived and have a strong bias for a 5'-end deoxycytidine.These small interfering DNAs guide TtAgo to cleave complementary DNA strands.Hence，despite structural homology to its eukaryotic counterparts，TtAgo functions in host defence by DNA-guided DNA interference.

Cas9-RNA“审查”DNA的机制

基于CRISPR/Cas9的DNA靶向技术成为合成生物学和基因组工程领域的一个领先工具。该研究报告了利用单分子和宏观生化实验来揭示RNA引导的Cas9如何识别基因组DNA并查找特定的切割位点。结果显示，一个很短的三核苷酸protospacer adjacent motif(PAM)，在将Cas9-RNA复合物向潜在DNA靶点上招募中以及在激活核酸酶催化活性中起作用。竞争测定法实验证据表明，DNA链的分离和RNA-DNA异源双链体的形成开始于PAM，并朝向靶序列的远末端进行。

论文链接:Sternberg S H，et al..DNA interrogation by the CRISPR RNA-guided endonuclease Cas9.

Nature，2014，507(7490)，62 －67.doi:10.1038/nature13011.

Abstract:The clustered regularly interspaced short palindromic repeats(CRISPR)-associated enzyme Cas9 is an RNA-guided endonuclease that uses RNA-DNA base-pairing to target foreign DNA in bacteria.Cas9-guide RNA complexes are also effective genome engineering agents in animals and plants.Here we use single-molecule and bulk biochemical experiments to determine how Cas9-RNA interrogates DNA to find specific cleavage sites.We show that both binding and cleavage of DNA by Cas9-RNA require recognition of a short trinucleotide protospacer adjacent motif(PAM).Non-target DNA binding affinity scales with PAM density，and sequences fully complementary to the guide RNA but lacking a nearby PAM are ignored by Cas9-RNA.Competition assays provide evidence that DNA strand separation and RNA-DNA heteroduplex formation initiate at the PAM and proceed directionally towards the distal end of the target sequence.Furthermore，PAM interactions trigger Cas9 catalytic activity.These results reveal how Cas9 uses PAM recognition to quickly identify potential target sites while scanning large DNA molecules，and to regulate scission of double-stranded DNA.

肺换气相关细胞的“兼性”干细胞功能

肺换气发生在结构精妙的肺泡囊中，其内壁有两种上皮细胞类型:鳞状1-型肺泡细胞(AT1细胞)和骰状2-型肺泡细胞(AT2细胞)。前者介导换气，后者分泌防止肺泡在呼吸过程中崩溃的表面活性剂。采用肺泡标记物、遗传世系追踪和克隆分析来在小鼠整个生命周期中对肺泡祖细胞进行活体识别，发现AT1和AT2细胞是在发育过程中从一种“双潜力”祖细胞形成的。在出生后，成熟AT2细胞起“兼性”干细胞的作用，形成再生肺泡的慢慢增大的单克隆位点。有致癌作用的Kras(G12D)突变永久性地激发AT2自我更新，劫持这种“兼性”干细胞功能来启动肺癌。

论文链接:Desa T J，et al..Alveolar progenitor and stem cells in lung development，renewal and cancer.

Nature，2014，507(7491)，190 －194.doi:10.1038/nature12930.

Abstract:Alveoli are gas-exchange sacs lined by squamous alveolar type(AT)1 cells and cuboidal，surfactant-secreting AT2 cells.Classical studies suggested that AT1 arise from AT2 cells，but recent studies propose other sources.Here we use molecular markers，lineage tracing and clonal analysis to map alveolar progenitors throughout the mouse lifespan.We show that，during development，AT1 and AT2 cells arise directly from a bipotent progenitor，whereas after birth new AT1 cells derive from rare，self-renewing，long-lived，mature AT2 cells that produce slowly expanding clonal foci of alveolar renewal.This stem-cell function is broadly activated by AT1 injury，and AT2 self-renewal is selectively induced by EGFR(epidermal growth factor receptor)ligands in vitro and oncogenic Kras(G12D)in vivo，efficiently generating multifocal，clonal adenomas.Thus，there is a switch after birth，when AT2 cells function as stem cells that contribute to alveolar renewal，repair and cancer.We propose that local signals regulate AT2 stem-cell activity:a signal transduced by EGFR-KRAS controls self-renewal and is hijacked during oncogenesis，whereas another signal controls reprogrammi ng to AT1 fate.

RNA毒性积累是导致C9orf72引发ALS和FTD两种疾病的原因

C9orf72六核苷酸重复扩张已被发现是造成“肌萎缩侧索硬化”(ALS)和“额颞痴呆”(FTD)这两种疾病的原因。正常的C9orf72含有多达25个重复片段，而患病者的该基因则会含有数千个。研究人员在新研究中找出了六核苷酸重复扩增(HRE)结构多态性导致ALS/FTD的病理分子机制。转录的C9orf72六核苷酸重复片段被发现以一种依赖于构形的方式与特定的核糖核蛋白如核仁蛋白相结合。因此，核仁蛋白被错误局部化，并且在功能上受损，导致“核仁应激”。RNA毒性的增加在ALS/FTD两种疾病中是造成与C9orf72相关病理的原因。

论文链接:Haeusler A R，et al..C9orf72 nucleotide repeat structures initiate molecular cascades of disease.

Nature，2014，507(7491)，195 －200.doi:10.1038/nature13124.

Abstract:A hexanucleotide repeat expansion(HRE)，(GGGGCC)n，in C9orf72 is the most common genetic cause of the neurodegenerative diseases amyotrophic lateral sclerosis(ALS)and frontotemporal dementia(FTD).Here we identify a molecular mechanism by which structural polymorphism of the HRE leads to ALS/FTD pathology and defects.The HRE forms DNA and RNA G-quadruplexes with distinct structures and promotes RNA·DNA hybrids(R-loops).The structural polymorphism causes a repeat-length-dependent accumulation of transcripts aborted in the HRE region.These transcribed repeats bind to ribonucleoproteins in a conformation-dependent manner.Specifically，nucleolin，an essential nucleolar protein，preferentially binds the HRE G-quadruplex，and patient cells show evidence of nucleolar stress.Our results demonstrate that distinct C9orf72 HRE structural polymorphism at both DNA and RNA levels initiates molecular cascades leading to ALS/FTD pathologies，and provide the basis for a mechanistic model for repeat-associated neurodegenerative dise ases.

可应用于疫苗研发的蛋白设计方法

在一项采用计算蛋白设计方法来生成人“呼吸道合胞体病毒”(RSV)疫苗的新颖候选药物的研究中，准确模仿一个RSV抗原表位结构的人造蛋白支架，被发现能在恒河猴身上诱导产生中和RSV的有效抗体。此研究所用的蛋白设计方法，即，表位集中和支架为基础的疫苗的设计，进一步加以开发可用于多种其他目标的疫苗，包括抗原性高度可变的病原体，例如，人免疫缺陷病毒和流感。

论文链接:Correia B E，et al..Proof of principle for epitope-focused vaccine design.

Nature，507(7491)，201 －206.doi:10.1038/nature12966.

Abstract:Vaccines prevent infectious disease largely by inducing protective neutralizing antibodies against vulnerable epitopes.Several major pathogens have resisted traditional vaccine development，although vulnerable epitopes targeted by neutralizing antibodies have been identified for several such cases.Hence，new vaccine design methods to induce epitope-specific neutralizing antibodies are needed.Here we show，with a neutralization epitope from respiratory syncytial virus，that computational protein design can generate small，thermally and conformationally stable protein scaffolds that accurately mimic the viral epitope structure and induce potent neutralizing antibodies.These scaffolds represent promising leads for the research and development of a human respiratory syncytial virus vaccine needed to protect infants，young children and the elderly.More generally，the results provide proof of principle for epitope-focused and scaffold-based vaccine design，and encourage the evaluation and further development of these strategies for a variety of other vaccine targets，including antigenically highly variable pathogens such as human immunodeficiency virus and infl uenza.

首次全面揭示食管鳞癌基因组异常改变

食管癌是中国高发的恶性肿瘤。目前，食管癌发生和发展的机制尚不明了，临床治疗缺乏特异性的分子靶点和有效的治疗药物。研究人员通过高通量测序、比较基因组杂交芯片分析、生物学功能和临床验证研究，全面系统揭示了食管鳞癌的遗传突变背景，发现了与食管鳞癌发生发展进程和临床预后相关的基因。

论文链接:Song Y M，et al..Identification of genomic alterations in oesophageal squamous cell cancer.

Nature，doi:10.1038/nature13176.Published online:16 March，2014.

Abstract:Oesophageal cancer is one of the most aggressive cancers and is the sixth leading cause of cancer death worldwide.Approximately 70%of global oesophageal cancer cases occur in China，with oesophageal squamous cell carcinoma(ESCC)being the histopathological form in the vast majority of cases(＞90%).Currently，there are limited clinical approaches for the early diagnosis and treatment of ESCC，resulting in a 10%five-year survival rate for patients.However，the full repertoire of genomic events leading to the pathogenesis of ESCC remains unclear.Here we describe a comprehensive genomic analysis of 158 ESCC cases，as part of the International Cancer Genome Consortium research project.We conducted whole-genome sequencing in 17 ESCC cases and whole-exome sequencing in 71 cases，of which 53 cases，plus an additional 70 ESCC cases not used in the whole-genome and whole-exome sequencing，were subjected to array comparative genomic hybridization analysis.We identified eight significantly mutated genes，of which six are well known tumour-associated genes(TP53，RB1，CDKN2A，PIK3CA，NOTCH1，NFE2L2)，and two have not previously been described in ESCC(ADAM29 and FAM135B).Notably，FAM135B is identified as a novel cancer-implicated gene as assayed for its ability to promote malignancy of ESCC cells.Additionally，MIR548K，a microRNA encoded in the amplified 11q13.3 ～ 13.4 region，is characterized as a novel oncogene，and functional assays demonstrate that MIR548K enhances malignant phenotypes of ESCC cells.Moreover，we have found that several important histone regulator genes(MLL2(also called KMT2D)，ASH1L，MLL3(KMT2C)，SETD1B，CREBBP and EP300)are frequently altered in ESCC.Pathway assessment reveals that somatic aberrations are mainly involved in the Wnt，cell cycle and Notch pathways.Genomic analyses suggest that ESCC and head and neck squamous cell carcinoma share some common pathogenic mechanisms，and ESCC development is associated with alcohol drinking.This study has explored novel biological markers and tumorigenic pathways that would greatly improve therapeutic strategies for ESCC.