RESUMO
Two alleles of a gene can be transcribed independently or coordinatedly, which can lead to temporal expression heterogeneity with potentially distinct impacts on cell fate. Here, we profiled genome-wide allelic transcriptional burst kinetics during the reprogramming of MEF to induced pluripotent stem cells. We show that the degree of coordination of allelic bursting differs among genes, and alleles of many reprogramming-related genes burst in a highly coordinated fashion. Notably, we show that the chromatin accessibility of the two alleles of highly coordinated genes is similar, unlike the semi-coordinated or independent genes, suggesting the degree of coordination of allelic bursting is linked to allelic chromatin accessibility. Consistently, we show that many transcription factors have differential binding affinity between alleles of semi-coordinated or independent genes. We show that highly coordinated genes are enriched with chromatin accessibility regulators such as H3K4me3, H3K4me1, H3K36me3, H3K27ac, histone variant H3.3, and BRD4. Finally, we demonstrate that enhancer elements are highly enriched in highly coordinated genes. Our study demonstrates that epigenomic states contribute to coordinated allelic bursting to fine-tune gene expression during induced pluripotent stem cell reprogramming.
Assuntos
Células-Tronco Pluripotentes Induzidas , Células-Tronco Pluripotentes Induzidas/metabolismo , Alelos , Epigenômica , Proteínas Nucleares/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Cromatina/genética , Cromatina/metabolismoRESUMO
In mammals, transcriptional inactivation of one X chromosome in female compensates for the dosage of X-linked gene expression between the sexes. Additionally, it is believed that the upregulation of active X chromosome in male and female balances the dosage of X-linked gene expression relative to autosomal genes, as proposed by Ohno. However, the existence of X chromosome upregulation (XCU) remains controversial. Here, we have profiled gene-wise dynamics of XCU in pre-gastrulation mouse embryos at single-cell level and found that XCU is dynamically linked with X chromosome inactivation (XCI); however, XCU is not global like XCI. Moreover, we show that upregulated genes are enriched with activating marks and have enhanced burst frequency. Finally, our In-silico model predicts that recruitment probabilities of activating factors and a surge of these factors upon X-inactivation trigger XCU. Altogether, our study provides significant insight into the gene-wise dynamics and mechanistic basis of XCU during early development and extends support for Ohno's hypothesis.
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XX female and XY male therian mammals equalize X-linked gene expression through the mitotically-stable transcriptional inactivation of one of the two X chromosomes in female somatic cells. Here, we describe an essential function of the X-linked homolog of an ancestral X-Y gene pair, Kdm5c-Kdm5d, in the expression of Xist lncRNA, which is required for stable X-inactivation. Ablation of Kdm5c function in females results in a significant reduction in Xist RNA expression. Kdm5c encodes a demethylase that enhances Xist expression by converting histone H3K4me2/3 modifications into H3K4me1. Ectopic expression of mouse and human KDM5C, but not the Y-linked homolog KDM5D, induces Xist in male mouse embryonic stem cells (mESCs). Similarly, marsupial (opossum) Kdm5c but not Kdm5d also upregulates Xist in male mESCs, despite marsupials lacking Xist, suggesting that the KDM5C function that activates Xist in eutherians is strongly conserved and predates the divergence of eutherian and metatherian mammals. In support, prototherian (platypus) Kdm5c also induces Xist in male mESCs. Together, our data suggest that eutherian mammals co-opted the ancestral demethylase KDM5C during sex chromosome evolution to upregulate Xist for the female-specific induction of X-inactivation.
Assuntos
Marsupiais , Ornitorrinco , RNA Longo não Codificante , Animais , Feminino , Genes Ligados ao Cromossomo X , Histona Desmetilases , Masculino , Mamíferos/genética , Marsupiais/genética , Camundongos , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Cromossomo X/genética , Cromossomo X/metabolismo , Inativação do Cromossomo X/genéticaRESUMO
Hsp90 gene of G. lamblia has a split nature comprising two ORFs separated by 777 kb on chromosome 5. The ORFs of the split gene on chromosome 5 undergo transcription to generate independent pre-mRNAs that join by a unique trans-splicing reaction that remains partially understood. The canonical cis-acting nucleotide elements such as 5'SS-GU, 3'SS-AG, polypyrimidine tract and branch point adenine are present in the independent pre-mRNAs and therefore trans-splicing of Hsp90 must be assisted by spliceosomes in vivo. Using an approach of RNA-protein pull down, we show that an RNA helicase selectively interacts with HspN pre-mRNA. Our experiments involving high resolution chromosome conformation capture technology as well as DNA FISH show that the trans-spliced genes of Giardia are in three-dimensional spatial proximity in the nucleus. Altogether our study provides a glimpse into the in vivo mechanisms involving protein factors as well as chromatin structure to facilitate the unique inter-molecular post-transcriptional stitching of split genes in G. lamblia.
Assuntos
Giardia lamblia/genética , Giardia lamblia/metabolismo , Proteínas de Choque Térmico HSP90/genética , Proteínas de Choque Térmico HSP90/metabolismo , Trans-Splicing , Humanos , Fases de Leitura Aberta , Precursores de RNA/genética , Precursores de RNA/metabolismo , Splicing de RNA , Spliceossomos/genética , Spliceossomos/metabolismoRESUMO
Recently, allele-specific single-cell RNA-seq analysis has demonstrated widespread dynamic random monoallelic expression of autosomal genes (aRME) in different cell types. However, the prevalence of dynamic aRME during pregastrulation remains unknown. Here, we show that dynamic aRME is widespread in different lineages of pregastrulation embryos. Additionally, the origin of dynamic aRME remains elusive. It is believed that independent transcriptional bursting from each allele leads to dynamic aRME. Here, we show that allelic burst is not perfectly independent; instead it happens in a semicoordinated fashion. Importantly, we show that semicoordinated allelic bursting of genes, particularly with low burst frequency, leads to frequent asynchronous allelic bursting, thereby contributing to dynamic aRME. Furthermore, we found that coordination of allelic bursting is lineage specific and genes regulating the development have a higher degree of coordination. Altogether, our study provides significant insights into the prevalence and origin of dynamic aRME and their developmental relevance during early development.
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Recently, a unique form of X chromosome dosage compensation has been demonstrated in human preimplantation embryos, which happens through the dampening of X-linked gene expression from both X chromosomes. Subsequently, X chromosome dampening has also been demonstrated in female human pluripotent stem cells (hPSCs) during the transition from primed to naive state. However, the existence of dampened X chromosomes in both embryos and hPSCs remains controversial. Specifically, in preimplantation embryos it has been shown that there is inactivation of X chromosome instead of dampening. Here, we performed allelic analysis of X-linked genes at the single-cell level in hPSCs and found that there is partial reactivation of the inactive X chromosome instead of chromosome-wide dampening upon conversion from primed to naive state. In addition, our analysis suggests that the reduced X-linked gene expression in naive hPSCs might be the consequence of erasure of active X chromosome upregulation.
Assuntos
Células-Tronco Embrionárias Humanas/metabolismo , Inativação do Cromossomo X , Alelos , Feminino , Humanos , RNA Longo não Codificante/genética , Análise de Célula ÚnicaRESUMO
Unliganded Estrogen receptor alpha (ERα) has been implicated in ligand-dependent gene regulation. Upon ligand exposure, ERα binds to several EREs relatively proximal to the pre-marked, unliganded ERα-bound sites and affects transient but robust gene expression. However, the underlying mechanisms are not fully understood. Here we demonstrate that upon ligand stimulation, persistent sites interact extensively, via chromatin looping, with the proximal transiently ERα-bound sites, forming Ligand Dependent ERα Enhancer Cluster in 3D (LDEC). The E2-target genes are regulated by these clustered enhancers but not by the H3K27Ac super-enhancers. Further, CRISPR-based deletion of TFF1 persistent site disrupts the formation of its LDEC resulting in the loss of E2-dependent expression of TFF1 and its neighboring genes within the same TAD. The LDEC overlap with nuclear ERα condensates that coalesce in a ligand and persistent site dependent manner. Furthermore, formation of clustered enhancers, as well as condensates, coincide with the active phase of signaling and their later disappearance results in the loss of gene expression even though persistent sites remain bound by ERα. Our results establish, at TFF1 and NRIP1 locus, a direct link between ERα condensates, ERα enhancer clusters, and transient, but robust, gene expression in a ligand-dependent fashion.
Assuntos
Montagem e Desmontagem da Cromatina , Elementos Facilitadores Genéticos , Receptor alfa de Estrogênio/metabolismo , Receptor alfa de Estrogênio/genética , Deleção de Genes , Histonas/metabolismo , Humanos , Ligantes , Células MCF-7 , Fator Trefoil-1/genéticaRESUMO
Around implantation, the epiblast (Epi) transits from naïve to primed pluripotency, before giving rise to the three germ layers. How chromatin is reconfigured during this developmental window remains poorly understood. We performed a genome-wide investigation of chromatin landscapes during this period. We find that enhancers in ectoderm are already pre-accessible in embryonic day 6.5 (E6.5) Epi when cells enter a primed pluripotent state. Unexpectedly, strong trimethylation of histone H3 at lysine 4 (H3K4me3) emerges at developmental gene promoters in E6.5 Epi and positively correlates with H3K27me3, thus establishing bivalency. These genes also show enhanced spatial interactions. Both the strong bivalency and spatial clustering are virtually absent in preimplantation embryos and are markedly reduced in fate-committed lineages. Finally, we show that KMT2B is essential for establishing bivalent H3K4me3 at E6.5 but becomes partially dispensable later. Its deficiency leads to impaired activation of developmental genes and subsequent embryonic lethality. Thus, our data characterize lineage-specific chromatin reconfiguration and a unique chromatin state for primed pluripotency.
Assuntos
Cromatina/genética , Embrião de Mamíferos/citologia , Células-Tronco Embrionárias/citologia , Epigenômica/métodos , Gastrulação , Regulação da Expressão Gênica no Desenvolvimento , Histona-Lisina N-Metiltransferase/fisiologia , Proteína de Leucina Linfoide-Mieloide/fisiologia , Animais , Metilação de DNA , Embrião de Mamíferos/metabolismo , Células-Tronco Embrionárias/metabolismo , Epigênese Genética , Feminino , Histonas/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos KnockoutRESUMO
The recent report of X-chromosome dampening in human preimplantation embryos remains controversial. Subsequently, Sahakyan et al. found evidence of X-chromosome dampening in human naïve pluripotent stem cells (hPSCs) as well. Here, we discuss whether X-dampening reported in hPSCs truly reflects the dampening of X-chromosomes or it is a consequence of the erasure of X-chromosome upregulation.
Assuntos
Cromossomos Humanos X , Mecanismo Genético de Compensação de Dose , Dosagem de Genes , Regulação da Expressão Gênica , Células-Tronco Pluripotentes/metabolismo , Animais , Desenvolvimento Embrionário/genética , Humanos , Células-Tronco Pluripotentes/citologia , Regulação para CimaRESUMO
Say-Meyer syndrome is a rare and clinically heterogeneous syndrome characterized by trigonocephaly, short stature, developmental delay and hypotelorism. Nine patients with this syndrome have been reported thus far although no causative gene has yet been identified. Here, we report two siblings with clinical phenotypes of Say-Meyer syndrome with moderate to severe intellectual disability and autism spectrum disorder. Cytogenetics and array-based comparative genomic hybridization did not reveal any chromosome abnormalities or copy number alterations. Exome sequencing of the patients revealed a novel X-linked recessive splice acceptor site variant c.145-2Aâ¯>â¯G in intron 5 of HUWE1 gene in both affected siblings. RT-PCR and sequencing revealed the use of an alternate cryptic splice acceptor site downstream, which led to deletion of six nucleotides resulting loss of two amino acids p.(Cys49-Glu50del) in HUWE1 protein. Deletion of these two amino acids, which are located in a highly conserved region, is predicted to be deleterious and quite likely to affect the function of HUWE1 protein. This is the first report of a potential candidate gene mutation for Say-Meyer syndrome, which was initially described four decades ago.
Assuntos
Anormalidades Múltiplas/genética , Transtorno do Espectro Autista/genética , Anormalidades Craniofaciais/genética , Transtornos do Crescimento/genética , Deficiência Intelectual/genética , Proteínas Supressoras de Tumor/genética , Ubiquitina-Proteína Ligases/genética , Anormalidades Múltiplas/patologia , Adolescente , Transtorno do Espectro Autista/patologia , Criança , Pré-Escolar , Hibridização Genômica Comparativa , Suturas Cranianas/diagnóstico por imagem , Suturas Cranianas/patologia , Anormalidades Craniofaciais/patologia , Exoma/genética , Feminino , Transtornos do Crescimento/patologia , Humanos , Deficiência Intelectual/patologia , Masculino , Isoformas de Proteínas/genética , Sítios de Splice de RNA/genética , Sequenciamento do ExomaRESUMO
Imprinted X-inactivation silences genes exclusively on the paternally-inherited X-chromosome and is a paradigm of transgenerational epigenetic inheritance in mammals. Here, we test the role of maternal vs. zygotic Polycomb repressive complex 2 (PRC2) protein EED in orchestrating imprinted X-inactivation in mouse embryos. In maternal-null (Eedm-/-) but not zygotic-null (Eed-/-) early embryos, the maternal X-chromosome ectopically induced Xist and underwent inactivation. Eedm-/- females subsequently stochastically silenced Xist from one of the two X-chromosomes and displayed random X-inactivation. This effect was exacerbated in embryos lacking both maternal and zygotic EED (Eedmz-/-), suggesting that zygotic EED can also contribute to the onset of imprinted X-inactivation. Xist expression dynamics in Eedm-/- embryos resemble that of early human embryos, which lack oocyte-derived maternal PRC2 and only undergo random X-inactivation. Thus, expression of PRC2 in the oocyte and transmission of the gene products to the embryo may dictate the occurrence of imprinted X-inactivation in mammals.
Assuntos
Camundongos/embriologia , Complexo Repressor Polycomb 2/metabolismo , Inativação do Cromossomo X , Animais , Camundongos Knockout , Complexo Repressor Polycomb 2/deficiência , RNA Longo não Codificante/metabolismoRESUMO
X-chromosome inactivation is a dosage compensation mechanism that equalizes X-linked gene expression between male and female mammals through the transcriptional silencing of most genes on one of the two X-chromosomes in females. With a few key exceptions, once the X-chromosome is inactivated replicated copies of that X-chromosome are maintained as inactive in all descendant cells. X-inactivation is therefore a paradigm of epigenetic inheritance. Imprinted X-inactivation is a specialized form of X-inactivation that results in the silencing of the paternally derived X-chromosome. Due to its parent-of-origin-specific pattern of inactivation, imprinted X-inactivation is a model of mitotic as well as meiotic, i.e., transgenerational, epigenetic inheritance. All cells of the early mouse embryo undergo imprinted X-inactivation, a pattern that is subsequently maintained in extraembryonic cell types in vivo and in vitro. Here, we describe both high- and low-throughput approaches to interrogate imprinted X-inactivation in the mouse embryo as well in cultured extraembryonic stem cells.
Assuntos
Alelos , Embrião de Mamíferos/metabolismo , Perfilação da Expressão Gênica/métodos , Hibridização in Situ Fluorescente/métodos , Inativação do Cromossomo X , Animais , Embrião de Mamíferos/fisiologia , Desenvolvimento Embrionário , Células-Tronco Embrionárias , Epigenômica/métodos , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Camundongos , Polimorfismo de Nucleotídeo Único , Análise de Sequência de RNA/métodosRESUMO
[This corrects the article DOI: 10.1371/journal.pone.0150763.].
RESUMO
BACKGROUND: Polycomb repressive complex 2 (PRC2) catalyzes histone H3K27me3, which marks many transcriptionally silent genes throughout the mammalian genome. Although H3K27me3 is associated with silenced gene expression broadly, it remains unclear why some but not other PRC2 target genes require PRC2 and H3K27me3 for silencing. RESULTS: Here we define the transcriptional and chromatin features that predict which PRC2 target genes require PRC2/H3K27me3 for silencing by interrogating imprinted mouse X-chromosome inactivation. H3K27me3 is enriched at promoters of silenced genes across the inactive X chromosome. To abrogate PRC2 function, we delete the core PRC2 protein EED in F1 hybrid trophoblast stem cells (TSCs), which undergo imprinted inactivation of the paternally inherited X chromosome. Eed -/- TSCs lack H3K27me3 and Xist lncRNA enrichment on the inactive X chromosome. Despite the absence of H3K27me3 and Xist RNA, only a subset of the inactivated X-linked genes is derepressed in Eed -/- TSCs. Unexpectedly, in wild-type (WT) TSCs these genes are transcribed and are enriched for active chromatin hallmarks on the inactive-X, including RNA PolII, H3K27ac, and H3K36me3, but not the bivalent mark H3K4me2. By contrast, PRC2 targets that remain repressed in Eed -/- TSCs are depleted for active chromatin characteristics in WT TSCs. CONCLUSIONS: A comparative analysis of transcriptional and chromatin features of inactive X-linked genes in WT and Eed -/- TSCs suggests that PRC2 acts as a brake to prevent induction of transcribed genes on the inactive X chromosome, a mode of PRC2 function that may apply broadly.
Assuntos
Impressão Genômica , Complexo Repressor Polycomb 2/metabolismo , Cromossomo X/genética , Animais , Cromatina/genética , Células-Tronco Embrionárias/metabolismo , Feminino , Inativação Gênica , Histonas/genética , Histonas/metabolismo , Masculino , Camundongos , Complexo Repressor Polycomb 2/genética , Trofoblastos/citologia , Inativação do Cromossomo XRESUMO
BACKGROUND: Family members of sucrose non-fermenting 1-related kinase 2 (SnRK2), being plant-specific serine/threonine protein kinases, constitute the central core of abscisic acid (ABA)-dependent and ABA-independent signaling pathways, and are key regulators of abiotic stress adaptation in plants. We report here the functional characterization of SAPK9 gene, one of the 10 SnRK2s of rice, through developing gain-of-function and loss-of-function phenotypes by transgenesis. RESULTS: The gene expression profiling revealed that the abundance of single gene-derived SAPK9 transcript was significantly higher in drought-tolerant rice genotypes than the drought-sensitive ones, and its expression was comparatively greater in reproductive stage than the vegetative stage. The highest expression of SAPK9 gene in drought-tolerant Oryza rufipogon prompted us to clone and characterise the CDS of this allele in details. The SAPK9 transcript expression was found to be highest in leaf and upregulated during drought stress and ABA treatment. In silico homology modelling of SAPK9 with Arabidopsis OST1 protein showed the bilobal kinase fold structure of SAPK9, which upon bacterial expression was able to phosphorylate itself, histone III and OsbZIP23 as substrates in vitro. Transgenic overexpression (OE) of SAPK9 CDS from O. rufipogon in a drought-sensitive indica rice genotype exhibited significantly improved drought tolerance in comparison to transgenic silencing (RNAi) lines and non-transgenic (NT) plants. In contrast to RNAi and NT plants, the enhanced drought tolerance of OE lines was concurrently supported by the upgraded physiological indices with respect to water retention capacity, soluble sugar and proline content, stomatal closure, membrane stability, and cellular detoxification. Upregulated transcript expressions of six ABA-dependent stress-responsive genes and increased sensitivity to exogenous ABA of OE lines indicate that the SAPK9 is a positive regulator of ABA-mediated stress signaling pathways in rice. The yield-related traits of OE lines were augmented significantly, which resulted from the highest percentage of fertile pollens in OE lines when compared with RNAi and NT plants. CONCLUSION: The present study establishes the functional role of SAPK9 as transactivating kinase and potential transcriptional activator in drought stress adaptation of rice plant. The SAPK9 gene has potential usefulness in transgenic breeding for improving drought tolerance and grain yield in crop plants.
Assuntos
Oryza/enzimologia , Proteínas de Plantas/metabolismo , Estômatos de Plantas/metabolismo , Proteínas Quinases/metabolismo , Secas , Regulação da Expressão Gênica de Plantas , Oryza/genética , Oryza/crescimento & desenvolvimento , Oryza/metabolismo , Osmose , Proteínas de Plantas/genética , Estômatos de Plantas/genética , Proteínas Quinases/genética , Sementes/enzimologia , Sementes/genética , Sementes/crescimento & desenvolvimento , Sementes/metabolismo , Sacarose/metabolismoRESUMO
The interconversion between naive and primed pluripotent states is accompanied by drastic epigenetic rearrangements. However, it is unclear whether intrinsic epigenetic events can drive reprogramming to naive pluripotency or if distinct chromatin states are instead simply a reflection of discrete pluripotent states. Here, we show that blocking histone H3K4 methyltransferase MLL1 activity with the small-molecule inhibitor MM-401 reprograms mouse epiblast stem cells (EpiSCs) to naive pluripotency. This reversion is highly efficient and synchronized, with more than 50% of treated EpiSCs exhibiting features of naive embryonic stem cells (ESCs) within 3 days. Reverted ESCs reactivate the silenced X chromosome and contribute to embryos following blastocyst injection, generating germline-competent chimeras. Importantly, blocking MLL1 leads to global redistribution of H3K4me1 at enhancers and represses lineage determinant factors and EpiSC markers, which indirectly regulate ESC transcription circuitry. These findings show that discrete perturbation of H3K4 methylation is sufficient to drive reprogramming to naive pluripotency.
Assuntos
Reprogramação Celular/efeitos dos fármacos , Histona-Lisina N-Metiltransferase/antagonistas & inibidores , Células-Tronco Embrionárias Murinas/efeitos dos fármacos , Proteína de Leucina Linfoide-Mieloide/antagonistas & inibidores , Oligopeptídeos/farmacologia , Células-Tronco Pluripotentes/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/farmacologia , Animais , Linhagem Celular , Camadas Germinativas/efeitos dos fármacos , Histona-Lisina N-Metiltransferase/deficiência , Histona-Lisina N-Metiltransferase/metabolismo , Camundongos , Células-Tronco Embrionárias Murinas/metabolismo , Proteína de Leucina Linfoide-Mieloide/deficiência , Proteína de Leucina Linfoide-Mieloide/metabolismo , Células-Tronco Pluripotentes/metabolismoRESUMO
Drought is one of the major limiting factors for productivity of crops including rice (Oryza sativa L.). Understanding the role of allelic variations of key regulatory genes involved in stress-tolerance is essential for developing an effective strategy to combat drought. The bZIP transcription factors play a crucial role in abiotic-stress adaptation in plants via abscisic acid (ABA) signaling pathway. The present study aimed to search for allelic polymorphism in the OsbZIP23 gene across selected drought-tolerant and drought-sensitive rice genotypes, and to characterize the new allele through overexpression (OE) and gene-silencing (RNAi). Analyses of the coding DNA sequence (CDS) of the cloned OsbZIP23 gene revealed single nucleotide polymorphism at four places and a 15-nucleotide deletion at one place. The single-copy OsbZIP23 gene is expressed at relatively higher level in leaf tissues of drought-tolerant genotypes, and its abundance is more in reproductive stage. Cloning and sequence analyses of the OsbZIP23-promoter from drought-tolerant O. rufipogon and drought-sensitive IR20 cultivar showed variation in the number of stress-responsive cis-elements and a 35-nucleotide deletion at 5'-UTR in IR20. Analysis of the GFP reporter gene function revealed that the promoter activity of O. rufipogon is comparatively higher than that of IR20. The overexpression of any of the two polymorphic forms (1083 bp and 1068 bp CDS) of OsbZIP23 improved drought tolerance and yield-related traits significantly by retaining higher content of cellular water, soluble sugar and proline; and exhibited decrease in membrane lipid peroxidation in comparison to RNAi lines and non-transgenic plants. The OE lines showed higher expression of target genes-OsRab16B, OsRab21 and OsLEA3-1 and increased ABA sensitivity; indicating that OsbZIP23 is a positive transcriptional-regulator of the ABA-signaling pathway. Taken together, the present study concludes that the enhanced gene expression rather than natural polymorphism in coding sequence of OsbZIP23 is accountable for improved drought tolerance and yield performance in rice genotypes.
Assuntos
Adaptação Fisiológica/genética , Secas , Regulação da Expressão Gênica de Plantas , Fases de Leitura Aberta/genética , Oryza/genética , Proteínas de Plantas/genética , Polimorfismo Genético , Ácido Abscísico/farmacologia , Alelos , Sequência de Bases , Clonagem Molecular , Dosagem de Genes , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Inativação Gênica/efeitos dos fármacos , Genes de Plantas , Genes Reporter , Genótipo , Germinação/efeitos dos fármacos , Proteínas de Fluorescência Verde/metabolismo , Dados de Sequência Molecular , Oryza/efeitos dos fármacos , Oryza/crescimento & desenvolvimento , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/genética , Folhas de Planta/efeitos dos fármacos , Folhas de Planta/fisiologia , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas , Regiões Promotoras Genéticas , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Estresse Fisiológico/efeitos dos fármacos , Estresse Fisiológico/genética , ÁguaRESUMO
To meet current challenges in agriculture, genome editing using sequence-specific nucleases (SSNs) is a powerful tool for basic and applied plant biology research. Here, we describe the principle and application of available genome editing tools, including zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs) and the clustered regularly interspaced short palindromic repeat associated CRISPR/Cas9 system. Among these SSNs, CRISPR/Cas9 is the most recently characterized and rapidly developing genome editing technology, and has been successfully utilized in a wide variety of organisms. This review specifically illustrates the power of CRISPR/Cas9 as a tool for plant genome engineering, and describes the strengths and weaknesses of the CRISPR/Cas9 technology compared to two well-established genome editing tools, ZFNs and TALENs.