DNA interrogation by the CRISPR RNA-guided endonuclease Cas9.

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.

journal_name

Nature

journal_title

Nature

authors

Sternberg SH,Redding S,Jinek M,Greene EC,Doudna JA

doi

10.1038/nature13011

subject

Has Abstract

pub_date

2014-03-06 00:00:00

pages

62-7

issue

7490

eissn

0028-0836

issn

1476-4687

pii

nature13011

journal_volume

507

pub_type

杂志文章

相关文献

NATURE文献大全
  • Structure of the Sec23/24-Sar1 pre-budding complex of the COPII vesicle coat.

    abstract::COPII-coated vesicles form on the endoplasmic reticulum by the stepwise recruitment of three cytosolic components: Sar1-GTP to initiate coat formation, Sec23/24 heterodimer to select SNARE and cargo molecules, and Sec13/31 to induce coat polymerization and membrane deformation. Crystallographic analysis of the Sacchar...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/nature01040

    authors: Bi X,Corpina RA,Goldberg J

    更新日期:2002-09-19 00:00:00

  • Synapse elimination and learning rules co-regulated by MHC class I H2-Db.

    abstract::The formation of precise connections between retina and lateral geniculate nucleus (LGN) involves the activity-dependent elimination of some synapses, with strengthening and retention of others. Here we show that the major histocompatibility complex (MHC) class I molecule H2-D(b) is necessary and sufficient for synaps...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/nature13154

    authors: Lee H,Brott BK,Kirkby LA,Adelson JD,Cheng S,Feller MB,Datwani A,Shatz CJ

    更新日期:2014-05-08 00:00:00

  • Australopithecus ramidus, a new species of early hominid from Aramis, Ethiopia.

    abstract::Seventeen hominoid fossils recovered from Pliocene strata at Aramis, Middle Awash, Ethiopia make up a series comprising dental, cranial and postcranial specimens dated to around 4.4 million years ago. When compared with Australopithecus afarensis and with modern and fossil apes the Aramis fossil hominids are recognize...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/371306a0

    authors: White TD,Suwa G,Asfaw B

    更新日期:1994-09-22 00:00:00

  • Flight of an aeroplane with solid-state propulsion.

    abstract::Since the first aeroplane flight more than 100 years ago, aeroplanes have been propelled using moving surfaces such as propellers and turbines. Most have been powered by fossil-fuel combustion. Electroaerodynamics, in which electrical forces accelerate ions in a fluid1,2, has been proposed as an alternative method of ...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/s41586-018-0707-9

    authors: Xu H,He Y,Strobel KL,Gilmore CK,Kelley SP,Hennick CC,Sebastian T,Woolston MR,Perreault DJ,Barrett SRH

    更新日期:2018-11-01 00:00:00

  • Role of melanocortinergic neurons in feeding and the agouti obesity syndrome.

    abstract::Dominant alleles at the agouti locus (A) cause an obesity syndrome in the mouse, as a consequence of ectopic expression of the agouti peptide. This peptide, normally only found in the skin, is a high-affinity antagonist of the melanocyte-stimulating hormone receptor (MC1-R), thus explaining the inhibitory effect of ag...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/385165a0

    authors: Fan W,Boston BA,Kesterson RA,Hruby VJ,Cone RD

    更新日期:1997-01-09 00:00:00

  • Control of MAP kinase activation by the mitogen-induced threonine/tyrosine phosphatase PAC1.

    abstract::Intracellular signalling following mitogenic stimulation of quiescent cells involves the initiation of a phosphorylation cascade that leads to the rapid and reversible activation of the mitogen-activated protein (MAP) kinases ERK1 and ERK2. MAP kinase activation is mediated by dual phosphorylation within the motif Thr...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/367651a0

    authors: Ward Y,Gupta S,Jensen P,Wartmann M,Davis RJ,Kelly K

    更新日期:1994-02-17 00:00:00

  • Somatic hypermutation of an immunoglobulin transgene in kappa transgenic mice.

    abstract::Initial studies of somatically acquired mutations in immunoglobulin V regions from hybridomas and myelomas that are not derived from joining aberrations, suggested a controlled and specific hypermutation process, because spontaneous mutation rates observed for other genes are extremely low. Some evidence for the idea ...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/326405a0

    authors: O'Brien RL,Brinster RL,Storb U

    更新日期:1987-03-26 00:00:00

  • An outburst from a massive star 40 days before a supernova explosion.

    abstract::Some observations suggest that very massive stars experience extreme mass-loss episodes shortly before they explode as supernovae, as do several models. Establishing a causal connection between these mass-loss episodes and the final explosion would provide a novel way to study pre-supernova massive-star evolution. Her...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/nature11877

    authors: Ofek EO,Sullivan M,Cenko SB,Kasliwal MM,Gal-Yam A,Kulkarni SR,Arcavi I,Bildsten L,Bloom JS,Horesh A,Howell DA,Filippenko AV,Laher R,Murray D,Nakar E,Nugent PE,Silverman JM,Shaviv NJ,Surace J,Yaron O

    更新日期:2013-02-07 00:00:00

  • Forensics: age written in teeth by nuclear tests.

    abstract::Establishing the age at death of individuals is an important step in their identification and can be done with high precision up to adolescence by analysis of dentition, but it is more difficult in adults. Here we show that the amount of radiocarbon present in tooth enamel as a result of nuclear bomb testing during 19...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/437333a

    authors: Spalding KL,Buchholz BA,Bergman LE,Druid H,Frisén J

    更新日期:2005-09-15 00:00:00

  • The genetics of monarch butterfly migration and warning colouration.

    abstract::The monarch butterfly, Danaus plexippus, is famous for its spectacular annual migration across North America, recent worldwide dispersal, and orange warning colouration. Despite decades of study and broad public interest, we know little about the genetic basis of these hallmark traits. Here we uncover the history of t...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/nature13812

    authors: Zhan S,Zhang W,Niitepõld K,Hsu J,Haeger JF,Zalucki MP,Altizer S,de Roode JC,Reppert SM,Kronforst MR

    更新日期:2014-10-16 00:00:00

  • Clonal interaction in tumours.

    abstract::The development of cancer is contingent on the emergence of at least one clone of transformed cells. One method used to investigate whether human tumours are monoclonal depends on the mosaicism in the normal tissues of women heterozygous for the two forms of the enzyme glucose-6-phosphate dehydrogenase (G-6-PD). This ...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/299822a0

    authors: Woodruff MF,Ansell JD,Forbes GM,Gordon JC,Burton DI,Micklem HS

    更新日期:1982-10-28 00:00:00

  • Role of the polycomb protein EED in the propagation of repressive histone marks.

    abstract::Polycomb group proteins have an essential role in the epigenetic maintenance of repressive chromatin states. The gene-silencing activity of the Polycomb repressive complex 2 (PRC2) depends on its ability to trimethylate lysine 27 of histone H3 (H3K27) by the catalytic SET domain of the EZH2 subunit, and at least two o...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/nature08398

    authors: Margueron R,Justin N,Ohno K,Sharpe ML,Son J,Drury WJ 3rd,Voigt P,Martin SR,Taylor WR,De Marco V,Pirrotta V,Reinberg D,Gamblin SJ

    更新日期:2009-10-08 00:00:00

  • Development of a preventive vaccine for Ebola virus infection in primates.

    abstract::Outbreaks of haemorrhagic fever caused by the Ebola virus are associated with high mortality rates that are a distinguishing feature of this human pathogen. The highest lethality is associated with the Zaire subtype, one of four strains identified to date. Its rapid progression allows little opportunity to develop nat...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/35046108

    authors: Sullivan NJ,Sanchez A,Rollin PE,Yang ZY,Nabel GJ

    更新日期:2000-11-30 00:00:00

  • Glaciations in response to climate variations preconditioned by evolving topography.

    abstract::Landscapes modified by glacial erosion show a distinct distribution of surface area with elevation (hypsometry). In particular, the height of these regions is influenced by climatic gradients controlling the altitude where glacial and periglacial processes are the most active, and as a result, surface area is focused ...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/nature11786

    authors: Pedersen VK,Egholm DL

    更新日期:2013-01-10 00:00:00

  • eIF4F is a nexus of resistance to anti-BRAF and anti-MEK cancer therapies.

    abstract::In BRAF(V600)-mutant tumours, most mechanisms of resistance to drugs that target the BRAF and/or MEK kinases rely on reactivation of the RAS-RAF-MEK-ERK mitogen-activated protein kinase (MAPK) signal transduction pathway, on activation of the alternative, PI(3)K-AKT-mTOR, pathway (which is ERK independent) or on modul...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/nature13572

    authors: Boussemart L,Malka-Mahieu H,Girault I,Allard D,Hemmingsson O,Tomasic G,Thomas M,Basmadjian C,Ribeiro N,Thuaud F,Mateus C,Routier E,Kamsu-Kom N,Agoussi S,Eggermont AM,Désaubry L,Robert C,Vagner S

    更新日期:2014-09-04 00:00:00

  • Differential usage of iso-accepting tRNASer species in silk glands of Bombyx mori.

    abstract::The rapid development of the silk glands of Bombyx mori during the last larval instar shows two phases. During the first 4 days, in both the middle and posterior parts of the silk glands, the ribosomal machinery is assembled and the synthesis of housekeeping proteins starts. During the second phase (the last 4 days), ...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/290267a0

    authors: Hentzen D,Chevallier A,Garel JP

    更新日期:1981-03-19 00:00:00

  • cMpl ligand is a humoral regulator of megakaryocytopoiesis.

    abstract::Megakaryocytopoiesis is the cellular developmental process that leads to platelet production. At least two humoral growth factors may be necessary for megakaryocyte proliferation and maturation. One is a megakaryocyte-colony stimulating factor (MK-CSF) which induces the proliferation and differentiation of megakaryocy...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/369571a0

    authors: Wendling F,Maraskovsky E,Debili N,Florindo C,Teepe M,Titeux M,Methia N,Breton-Gorius J,Cosman D,Vainchenker W

    更新日期:1994-06-16 00:00:00

  • On analyzing scientific fraud.

    abstract::Braunwald, a professor at Harvard Medical School, responds to a paper by Walter W. Stewart and Ned Feder ("The integrity of the scientific literature," Nature 1987 Jan 15; 325(6101): 207-214) concerning their findings of errors, discrepancies, and other lapses in publications coauthored by John Darsee, a biomedical re...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/325215a0

    authors: Braunwald E

    更新日期:1987-01-15 00:00:00

  • Clarifying the glass-transition behaviour of water by comparison with hyperquenched inorganic glasses.

    abstract::The formation of glasses is normal for substances that remain liquid over a wide temperature range (the 'good glassformers') and can be induced for most liquids if cooling is fast enough to bypass crystallization. During reheating but still below the melting point, good glassformers exhibit glass transitions as they a...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/nature02295

    authors: Yue Y,Angell CA

    更新日期:2004-02-19 00:00:00

  • Linguistic perception: neural processing of a whistled language.

    abstract::Silbo Gomero is a whistled language that is a rare and endangered surrogate of Spanish, used by shepherds on the island of La Gomera in the Canary Islands for communication over long distances on difficult terrain. Here we show that areas of the brain normally associated with spoken-language function are also activate...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/433031a

    authors: Carreiras M,Lopez J,Rivero F,Corina D

    更新日期:2005-01-06 00:00:00

  • Sequencing the nuclear genome of the extinct woolly mammoth.

    abstract::In 1994, two independent groups extracted DNA from several Pleistocene epoch mammoths and noted differences among individual specimens. Subsequently, DNA sequences have been published for a number of extinct species. However, such ancient DNA is often fragmented and damaged, and studies to date have typically focused ...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/nature07446

    authors: Miller W,Drautz DI,Ratan A,Pusey B,Qi J,Lesk AM,Tomsho LP,Packard MD,Zhao F,Sher A,Tikhonov A,Raney B,Patterson N,Lindblad-Toh K,Lander ES,Knight JR,Irzyk GP,Fredrikson KM,Harkins TT,Sheridan S,Pringle T,Schuste

    更新日期:2008-11-20 00:00:00

  • The Ndc80 kinetochore complex forms oligomeric arrays along microtubules.

    abstract::The Ndc80 complex is a key site of regulated kinetochore-microtubule attachment (a process required for cell division), but the molecular mechanism underlying its function remains unknown. Here we present a subnanometre-resolution cryo-electron microscopy reconstruction of the human Ndc80 complex bound to microtubules...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/nature09423

    authors: Alushin GM,Ramey VH,Pasqualato S,Ball DA,Grigorieff N,Musacchio A,Nogales E

    更新日期:2010-10-14 00:00:00

  • Do female hyaenas choose mates based on tenure?

    abstract::In their investigation into whether female mate-choice drives male dispersal, Höner et al. argue that female spotted hyaenas (Crocuta crocuta) prefer mates whose tenure in the social group is less than the females' age, to avoid paternal incest, and suggest that male dispersal reflects this preference. However, we are...

    journal_title:Nature

    pub_type: 评论,杂志文章

    doi:10.1038/nature07122

    authors: Van Horn RC,Watts HE,Holekamp KE

    更新日期:2008-07-10 00:00:00

  • Origin of luteinizing hormone-releasing hormone neurons.

    abstract::Neurons expressing luteinizing hormone-releasing hormone (LHRH), found in the septal-preoptic nuclei and hypothalamus, control the release of gonadotropic hormones from the anterior pituitary gland and facilitate reproductive behaviour. LHRH-expressing neurons are also found in the nervus terminalis, a cranial nerve t...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/338161a0

    authors: Schwanzel-Fukuda M,Pfaff DW

    更新日期:1989-03-09 00:00:00

  • Fast vesicle reloading and a large pool sustain high bandwidth transmission at a central synapse.

    abstract::What limits the rate at which sensory information can be transmitted across synaptic connections in the brain? High-frequency signalling is restricted to brief bursts at many central excitatory synapses, whereas graded ribbon-type synapses can sustain release and transmit information at high rates. Here we investigate...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/nature04509

    authors: Saviane C,Silver RA

    更新日期:2006-02-23 00:00:00

  • A salt-water reservoir as the source of a compositionally stratified plume on Enceladus.

    abstract::The discovery of a plume of water vapour and ice particles emerging from warm fractures ('tiger stripes') in Saturn's small, icy moon Enceladus raised the question of whether the plume emerges from a subsurface liquid source or from the decomposition of ice. Previous compositional analyses of particles injected by the...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/nature10175

    authors: Postberg F,Schmidt J,Hillier J,Kempf S,Srama R

    更新日期:2011-06-22 00:00:00

  • Gene for chronic proximal spinal muscular atrophies maps to chromosome 5q.

    abstract::Proximal spinal muscular atrophies represent the second most common fatal, autosomal recessive disorder after cystic fibrosis. The childhood form is classically subdivided into three groups: acute Werdnig-Hoffmann (type I), intermediate Werdnig-Hoffmann disease (type II) and Kugelberg-Welander disease (type III). Thes...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/344767a0

    authors: Melki J,Abdelhak S,Sheth P,Bachelot MF,Burlet P,Marcadet A,Aicardi J,Barois A,Carriere JP,Fardeau M

    更新日期:1990-04-19 00:00:00

  • Ad26/MVA therapeutic vaccination with TLR7 stimulation in SIV-infected rhesus monkeys.

    abstract::The development of immunologic interventions that can target the viral reservoir in HIV-1-infected individuals is a major goal of HIV-1 research. However, little evidence exists that the viral reservoir can be sufficiently targeted to improve virologic control following discontinuation of antiretroviral therapy. Here ...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/nature20583

    authors: Borducchi EN,Cabral C,Stephenson KE,Liu J,Abbink P,Ng'ang'a D,Nkolola JP,Brinkman AL,Peter L,Lee BC,Jimenez J,Jetton D,Mondesir J,Mojta S,Chandrashekar A,Molloy K,Alter G,Gerold JM,Hill AL,Lewis MG,Pau MG,Schuit

    更新日期:2016-12-08 00:00:00

  • Identity and dynamics of mammary stem cells during branching morphogenesis.

    abstract::During puberty, the mouse mammary gland develops into a highly branched epithelial network. Owing to the absence of exclusive stem cell markers, the location, multiplicity, dynamics and fate of mammary stem cells (MaSCs), which drive branching morphogenesis, are unknown. Here we show that morphogenesis is driven by pr...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/nature21046

    authors: Scheele CL,Hannezo E,Muraro MJ,Zomer A,Langedijk NS,van Oudenaarden A,Simons BD,van Rheenen J

    更新日期:2017-02-16 00:00:00

  • Genome-wide atlas of gene expression in the adult mouse brain.

    abstract::Molecular approaches to understanding the functional circuitry of the nervous system promise new insights into the relationship between genes, brain and behaviour. The cellular diversity of the brain necessitates a cellular resolution approach towards understanding the functional genomics of the nervous system. We des...

    journal_title:Nature

    pub_type: 杂志文章

    doi:10.1038/nature05453

    authors: Lein ES,Hawrylycz MJ,Ao N,Ayres M,Bensinger A,Bernard A,Boe AF,Boguski MS,Brockway KS,Byrnes EJ,Chen L,Chen L,Chen TM,Chin MC,Chong J,Crook BE,Czaplinska A,Dang CN,Datta S,Dee NR,Desaki AL,Desta T,Diep E,Dol

    更新日期:2007-01-11 00:00:00