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1.
PLoS Genet ; 17(9): e1009779, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34591845

RESUMO

Meiosis in angiosperm plants is followed by mitotic divisions to form multicellular haploid gametophytes. Termination of meiosis and transition to gametophytic development is, in Arabidopsis, governed by a dedicated mechanism that involves SMG7 and TDM1 proteins. Mutants carrying the smg7-6 allele are semi-fertile due to reduced pollen production. We found that instead of forming tetrads, smg7-6 pollen mother cells undergo multiple rounds of chromosome condensation and spindle assembly at the end of meiosis, resembling aberrant attempts to undergo additional meiotic divisions. A suppressor screen uncovered a mutation in centromeric histone H3 (CENH3) that increased fertility and promoted meiotic exit in smg7-6 plants. The mutation led to inefficient splicing of the CENH3 mRNA and a substantial decrease of CENH3, resulting in smaller centromeres. The reduced level of CENH3 delayed formation of the mitotic spindle but did not have an apparent effect on plant growth and development. We suggest that impaired spindle re-assembly at the end of meiosis limits aberrant divisions in smg7-6 plants and promotes formation of tetrads and viable pollen. Furthermore, the mutant with reduced level of CENH3 was very inefficient haploid inducer indicating that differences in centromere size is not the key determinant of centromere-mediated genome elimination.


Assuntos
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Proteínas de Transporte/genética , Genes de Plantas , Meiose/genética , Mutação , Arabidopsis/fisiologia , Fertilidade/genética , RNA Mensageiro/genética , Fuso Acromático
2.
New Phytol ; 240(5): 2085-2101, 2023 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-37823324

RESUMO

Recent studies have shown that correlations between chromatin modifications and transcription vary among eukaryotes. This is the case for marked differences between the chromatin of the moss Physcomitrium patens and the liverwort Marchantia polymorpha. Mosses and liverworts diverged from hornworts, altogether forming the lineage of bryophytes that shared a common ancestor with land plants. We aimed to describe chromatin in hornworts to establish synapomorphies across bryophytes and approach a definition of the ancestral chromatin organization of land plants. We used genomic methods to define the 3D organization of chromatin and map the chromatin landscape of the model hornwort Anthoceros agrestis. We report that nearly half of the hornwort transposons were associated with facultative heterochromatin and euchromatin and formed the center of topologically associated domains delimited by protein coding genes. Transposons were scattered across autosomes, which contrasted with the dense compartments of constitutive heterochromatin surrounding the centromeres in flowering plants. Most of the features observed in hornworts are also present in liverworts or in mosses but are distinct from flowering plants. Hence, the ancestral genome of bryophytes was likely a patchwork of units of euchromatin interspersed within facultative and constitutive heterochromatin. We propose this genome organization was ancestral to land plants.


Assuntos
Anthocerotophyta , Briófitas , Bryopsida , Filogenia , Cromatina , Heterocromatina/genética , Eucromatina/genética , Briófitas/genética , Anthocerotophyta/genética , Bryopsida/genética
3.
Proc Natl Acad Sci U S A ; 113(43): 12226-12231, 2016 10 25.
Artigo em Inglês | MEDLINE | ID: mdl-27729523

RESUMO

In plants, gametogenesis occurs late in development, and somatic mutations can therefore be transmitted to the next generation. Longer periods of growth are believed to result in an increase in the number of cell divisions before gametogenesis, with a concomitant increase in mutations arising due to replication errors. However, there is little experimental evidence addressing how many cell divisions occur before gametogenesis. Here, we measured loss of telomeric DNA and accumulation of replication errors in Arabidopsis with short and long life spans to determine the number of replications in lineages leading to gametes. Surprisingly, the number of cell divisions within the gamete lineage is nearly independent of both life span and vegetative growth. One consequence of the relatively stable number of replications per generation is that older plants may not pass along more somatically acquired mutations to their offspring. We confirmed this hypothesis by genomic sequencing of progeny from young and old plants. This independence can be achieved by hierarchical arrangement of cell divisions in plant meristems where vegetative growth is primarily accomplished by expansion of cells in rapidly dividing meristematic zones, which are only rarely refreshed by occasional divisions of more quiescent cells. We support this model by 5-ethynyl-2'-deoxyuridine retention experiments in shoot and root apical meristems. These results suggest that stem-cell organization has independently evolved in plants and animals to minimize mutations by limiting DNA replication.


Assuntos
Arabidopsis/genética , Replicação do DNA/genética , Genoma de Planta/genética , Meristema/genética , Arabidopsis/crescimento & desenvolvimento , Diploide , Regulação da Expressão Gênica de Plantas , Células Germinativas/crescimento & desenvolvimento , Meristema/crescimento & desenvolvimento , Mutação/genética , Acúmulo de Mutações , Células Vegetais , Raízes de Plantas/genética , Raízes de Plantas/crescimento & desenvolvimento , Brotos de Planta/genética , Brotos de Planta/crescimento & desenvolvimento , Caules de Planta/genética , Caules de Planta/crescimento & desenvolvimento , Análise de Sequência de DNA
4.
PLoS Genet ; 9(5): e1003508, 2013 May.
Artigo em Inglês | MEDLINE | ID: mdl-23671425

RESUMO

Meiosis is a modified cell division in which a single S-phase is followed by two rounds of chromosome segregation resulting in the production of haploid gametes. The meiotic mode of chromosome segregation requires extensive remodeling of the basic cell cycle machinery and employment of unique regulatory mechanisms. Cyclin-dependent kinases (CDKs) and cyclins represent an ancient molecular module that drives and regulates cell cycle progression. The cyclin gene family has undergone a massive expansion in angiosperm plants, but only a few cyclins were thoroughly characterized. In this study we performed a systematic immunolocalization screen to identify Arabidopsis thaliana A- and B-type cyclins expressed in meiosis. Many of these cyclins exhibit cell-type-specific expression in vegetative tissues and distinct subcellular localization. We found six A-type cyclins and a single B-type cyclin (CYCB3;1) to be expressed in male meiosis. Mutant analysis revealed that these cyclins contribute to distinct meiosis-related processes. While A2 cyclins are important for chromosome segregation, CYCB3;1 prevents ectopic cell wall formation. We further show that cyclin SDS does not contain a D-box and is constitutively expressed throughout meiosis. Analysis of plants carrying cyclin SDS with an introduced D-box motif determined that, in addition to its function in recombination, SDS acts together with CYCB3;1 in suppressing unscheduled cell wall synthesis. Our phenotypic and expression data provide extensive evidence that multiplication of cyclins is in plants accompanied by functional diversification.


Assuntos
Proteínas de Arabidopsis/genética , Parede Celular , Ciclina A/genética , Ciclina B/genética , Ciclinas/genética , Meiose/genética , Arabidopsis/genética , Parede Celular/genética , Parede Celular/fisiologia , Segregação de Cromossomos/genética , Quinases Ciclina-Dependentes/genética , Quinases Ciclina-Dependentes/metabolismo , Regulação da Expressão Gênica de Plantas , Especificidade de Órgãos , Recombinação Genética/genética
5.
PLoS Genet ; 6(6): e1000986, 2010 Jun 10.
Artigo em Inglês | MEDLINE | ID: mdl-20548962

RESUMO

Chromosome termini form a specialized type of heterochromatin that is important for chromosome stability. The recent discovery of telomeric RNA transcripts in yeast and vertebrates raised the question of whether RNA-based mechanisms are involved in the formation of telomeric heterochromatin. In this study, we performed detailed analysis of chromatin structure and RNA transcription at chromosome termini in Arabidopsis. Arabidopsis telomeres display features of intermediate heterochromatin that does not extensively spread to subtelomeric regions which encode transcriptionally active genes. We also found telomeric repeat-containing transcripts arising from telomeres and centromeric loci, a portion of which are processed into small interfering RNAs. These telomeric siRNAs contribute to the maintenance of telomeric chromatin through promoting methylation of asymmetric cytosines in telomeric (CCCTAAA)(n) repeats. The formation of telomeric siRNAs and methylation of telomeres relies on the RNA-dependent DNA methylation pathway. The loss of telomeric DNA methylation in rdr2 mutants is accompanied by only a modest effect on histone heterochromatic marks, indicating that maintenance of telomeric heterochromatin in Arabidopsis is reinforced by several independent mechanisms. In conclusion, this study provides evidence for an siRNA-directed mechanism of chromatin maintenance at telomeres in Arabidopsis.


Assuntos
Arabidopsis/genética , Metilação de DNA , RNA Interferente Pequeno/genética , Telômero , Cromatina/genética , Cromossomos de Plantas , Regulação da Expressão Gênica de Plantas , Mutação , Transcrição Gênica
6.
Elife ; 122023 07 19.
Artigo em Inglês | MEDLINE | ID: mdl-37467143

RESUMO

How different intrinsic sequence variations and regulatory modifications of histones combine in nucleosomes remain unclear. To test the importance of histone variants in the organization of chromatin we investigated how histone variants and histone modifications assemble in the Arabidopsis thaliana genome. We showed that a limited number of chromatin states divide euchromatin and heterochromatin into several subdomains. We found that histone variants are as significant as histone modifications in determining the composition of chromatin states. Particularly strong associations were observed between H2A variants and specific combinations of histone modifications. To study the role of H2A variants in organizing chromatin states we determined the role of the chromatin remodeler DECREASED IN DNA METHYLATION (DDM1) in the organization of chromatin states. We showed that the loss of DDM1 prevented the exchange of the histone variant H2A.Z to H2A.W in constitutive heterochromatin, resulting in significant effects on the definition and distribution of chromatin states in and outside of constitutive heterochromatin. We thus propose that dynamic exchanges of histone variants control the organization of histone modifications into chromatin states, acting as molecular landmarks.


Assuntos
Proteínas de Arabidopsis , Arabidopsis , Cromatina/genética , Histonas/genética , Arabidopsis/genética , Arabidopsis/metabolismo , Heterocromatina/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Nucleossomos/genética
7.
Curr Biol ; 33(20): 4367-4380.e9, 2023 10 23.
Artigo em Inglês | MEDLINE | ID: mdl-37738971

RESUMO

The mobility of transposable elements (TEs) contributes to evolution of genomes. Their uncontrolled activity causes genomic instability; therefore, expression of TEs is silenced by host genomes. TEs are marked with DNA and H3K9 methylation, which are associated with silencing in flowering plants, animals, and fungi. However, in distantly related groups of eukaryotes, TEs are marked by H3K27me3 deposited by the Polycomb repressive complex 2 (PRC2), an epigenetic mark associated with gene silencing in flowering plants and animals. The direct silencing of TEs by PRC2 has so far only been shown in one species of ciliates. To test if PRC2 silences TEs in a broader range of eukaryotes, we generated mutants with reduced PRC2 activity and analyzed the role of PRC2 in extant species along the lineage of Archaeplastida and in the diatom P. tricornutum. In this diatom and the red alga C. merolae, a greater proportion of TEs than genes were repressed by PRC2, whereas a greater proportion of genes than TEs were repressed by PRC2 in bryophytes. In flowering plants, TEs contained potential cis-elements recognized by transcription factors and associated with neighbor genes as transcriptional units repressed by PRC2. Thus, silencing of TEs by PRC2 is observed not only in Archaeplastida but also in diatoms and ciliates, suggesting that PRC2 deposited H3K27me3 to silence TEs in the last common ancestor of eukaryotes. We hypothesize that during the evolution of Archaeplastida, TE fragments marked with H3K27me3 were selected to shape transcriptional regulation, controlling networks of genes regulated by PRC2.


Assuntos
Arabidopsis , Complexo Repressor Polycomb 2 , Animais , Complexo Repressor Polycomb 2/genética , Complexo Repressor Polycomb 2/metabolismo , Histonas/genética , Histonas/metabolismo , Elementos de DNA Transponíveis/genética , Eucariotos/genética , Arabidopsis/genética , Regulação da Expressão Gênica de Plantas
8.
Elife ; 112022 08 23.
Artigo em Inglês | MEDLINE | ID: mdl-35996955

RESUMO

Complex mechanisms regulate gene dosage throughout eukaryotic life cycles. Mechanisms controlling gene dosage have been extensively studied in animals, however it is unknown how generalizable these mechanisms are to diverse eukaryotes. Here, we use the haploid plant Marchantia polymorpha to assess gene dosage control in its short-lived diploid embryo. We show that throughout embryogenesis, paternal chromosomes are repressed resulting in functional haploidy. The paternal genome is targeted for genomic imprinting by the Polycomb mark H3K27me3 starting at fertilization, rendering the maternal genome in control of embryogenesis. Maintaining haploid gene dosage by this new form of imprinting is essential for embryonic development. Our findings illustrate how haploid-dominant species can regulate gene dosage through paternal chromosome inactivation and initiates the exploration of the link between life cycle history and gene dosage in a broader range of organisms.


The reproductive cells of organisms that reproduce sexually ­ the egg and the sperm ­ each contain one copy of the organism's genome. An embryo forms upon fertilization of an egg by a sperm cell. This embryo contains two copies of the genome, one from each parent. Under most circumstances, it does not matter which parent a gene copy came from: both gene copies are expressed. However, in some species genes coming from only one of the parents are switched on. This unusual mode of gene expression is called genomic imprinting. The best-known example of this occurs in female mammals, which repress the genes on the paternal X chromosome. Genomic imprinting also exists in flowering plants. Both mammals and flowering plants evolved tissues that channel nutrients from the mother to the embryo during development; the placenta and the endosperm, respectively. Genomic imprinting had, until now, only been described in these two types of organisms. It was unknown whether imprinting also happens in other organisms, and specifically those in which embryos develop inside the mother but without the help of a placenta or endosperm. Here Montgomery et al. addressed this question by studying the liverwort, Marchantia polymorpha, a moss-like plant. Initial experiments showed that cells in the liverwort embryo mostly expressed the genes coming from the egg, and not the sperm. All the genetic material coming from the sperm had a molecular marker or tag called H3K27me3. This mark, which also appears on the paternal X chromosome in female mammals, switches off the genes it tags. M. polymorpha embryos thus suppress gene expression from all of the genetic material from the father, relying only on maternal genetic material for development. When Montgomery et al. deleted the maternal genes necessary for making the H3K27me3 mark, the paternal genes switched on, and this led to the death of the embryos. The survival of M. polymorpha embryos therefore depended on keeping only one set of genes active. Taken together these experiments indicate that genomic imprinting evolved about 480 million years ago, about 320 million years earlier than previously thought, in organisms for which embryo development depended only on one parent. This means there are likely many more organisms that control gene expression in this way, opening up opportunities for further research. Understanding imprinting in more detail will also shed light on how sexual reproduction evolved.


Assuntos
Diploide , Marchantia , Animais , Cromossomos , Impressão Genômica , Haploidia
9.
EMBO J ; 26(24): 5061-70, 2007 Dec 12.
Artigo em Inglês | MEDLINE | ID: mdl-18007598

RESUMO

Obligatory homologous recombination (HR) is required for chiasma formation and chromosome segregation in meiosis I. Meiotic HR is initiated by DNA double-strand breaks (DSBs), generated by Spo11, a homologue of the archaebacterial topoisomerase subunit Top6A. In Saccharomyces cerevisiae, Rad50, Mre11 and Com1/Sae2 are essential to process an intermediate of the cleavage reaction consisting of Spo11 covalently linked to the 5' termini of DNA. While Rad50 and Mre11 also confer genome stability to vegetative cells and are well conserved in evolution, Com1/Sae2 was believed to be fungal-specific. Here, we identify COM1/SAE2 homologues in all eukaryotic kingdoms. Arabidopsis thaliana Com1/Sae2 mutants are sterile, accumulate AtSPO11-1 during meiotic prophase and fail to form AtRAd51 foci despite the presence of unrepaired DSBs. Furthermore, DNA fragmentation in AtCom1 is suppressed by eliminating AtSPO11-1. In addition, AtCOM1 is specifically required for mitomycin C resistance. Interestingly, we identified CtIP, an essential protein interacting with the DNA repair machinery, as the mammalian homologue of Com1/Sae2, with important implications for the molecular role of CtIP.


Assuntos
Proteínas de Arabidopsis/metabolismo , Proteínas de Transporte/metabolismo , Proteínas de Ciclo Celular/metabolismo , Genes de Plantas , Meiose/fisiologia , Proteínas Nucleares/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Sequência de Aminoácidos , Animais , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Transporte/genética , Proteínas de Ciclo Celular/genética , Dano ao DNA , Fragmentação do DNA , Reparo do DNA , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Endodesoxirribonucleases , Endonucleases , Humanos , Mitomicina/metabolismo , Dados de Sequência Molecular , Mutação , Proteínas Nucleares/genética , Inibidores da Síntese de Ácido Nucleico/metabolismo , Fenótipo , Recombinação Genética , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Alinhamento de Sequência
10.
Nat Cell Biol ; 23(4): 391-400, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33833428

RESUMO

Mobile transposable elements (TEs) not only participate in genome evolution but also threaten genome integrity. In healthy cells, TEs that encode all of the components that are necessary for their mobility are specifically silenced, yet the precise mechanism remains unknown. Here, we characterize the mechanism used by a conserved class of chromatin remodelers that prevent TE mobility. In the Arabidopsis chromatin remodeler DECREASE IN DNA METHYLATION 1 (DDM1), we identify two conserved binding domains for the histone variant H2A.W, which marks plant heterochromatin. DDM1 is necessary and sufficient for the deposition of H2A.W onto potentially mobile TEs, yet does not act on TE fragments or host protein-coding genes. DDM1-mediated H2A.W deposition changes the properties of chromatin, resulting in the silencing of TEs and, therefore, prevents their mobility. This distinct mechanism provides insights into the interplay between TEs and their host in the contexts of evolution and disease, and potentiates innovative strategies for targeted gene silencing.


Assuntos
Proteínas de Arabidopsis/genética , Elementos de DNA Transponíveis/genética , Proteínas de Ligação a DNA/genética , Evolução Molecular , Histonas/genética , Fatores de Transcrição/genética , Arabidopsis/genética , Cromatina/genética , Montagem e Desmontagem da Cromatina/genética , Metilação de DNA/genética , Regulação da Expressão Gênica de Plantas/genética , Inativação Gênica , Genoma de Planta/genética , Heterocromatina/genética
11.
Curr Biol ; 30(4): 573-588.e7, 2020 02 24.
Artigo em Inglês | MEDLINE | ID: mdl-32004456

RESUMO

Genome packaging by nucleosomes is a hallmark of eukaryotes. Histones and the pathways that deposit, remove, and read histone modifications are deeply conserved. Yet, we lack information regarding chromatin landscapes in extant representatives of ancestors of the main groups of eukaryotes, and our knowledge of the evolution of chromatin-related processes is limited. We used the bryophyte Marchantia polymorpha, which diverged from vascular plants circa 400 mya, to obtain a whole chromosome genome assembly and explore the chromatin landscape and three-dimensional genome organization in an early diverging land plant lineage. Based on genomic profiles of ten chromatin marks, we conclude that the relationship between active marks and gene expression is conserved across land plants. In contrast, we observed distinctive features of transposons and other repetitive sequences in Marchantia compared with flowering plants. Silenced transposons and repeats did not accumulate around centromeres. Although a large fraction of constitutive heterochromatin was marked by H3K9 methylation as in flowering plants, a significant proportion of transposons were marked by H3K27me3, which is otherwise dedicated to the transcriptional repression of protein-coding genes in flowering plants. Chromatin compartmentalization analyses of Hi-C data revealed that repressed B compartments were densely decorated with H3K27me3 but not H3K9 or DNA methylation as reported in flowering plants. We conclude that, in early plants, H3K27me3 played an essential role in heterochromatin function, suggesting an ancestral role of this mark in transposon silencing.


Assuntos
Cromatina/fisiologia , Elementos de DNA Transponíveis/fisiologia , Embriófitas/fisiologia , Evolução Molecular , Heterocromatina/fisiologia
12.
Methods Mol Biol ; 1370: 169-82, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26659962

RESUMO

The ability to analyze cell division in both spatial and temporal dimensions within an organism is a key requirement in developmental biology. Specialized cell types within individual organs, such as those within shoot and root apical meristems, have often been identified by differences in their rates of proliferation prior to the characterization of distinguishing molecular markers. Replication-dependent labeling of DNA is a widely used method for assaying cell proliferation. The earliest approaches used radioactive labeling with tritiated thymidine, which were later followed by immunodetection of bromodeoxyuridine (BrdU). A major advance in DNA labeling came with the use of 5-ethynyl-2'deoxyuridine (EdU) which has proven to have multiple advantages over BrdU. Here we describe the methodology for analyzing EdU labeling and retention in whole plants and histological sections of Arabidopsis.


Assuntos
Arabidopsis/citologia , Proliferação de Células , DNA de Plantas/análise , Desoxiuridina/análogos & derivados , Coloração e Rotulagem/métodos , Arabidopsis/ultraestrutura , Replicação do DNA , Desoxiuridina/análise , Meristema/ultraestrutura , Inclusão em Parafina/métodos , Células Vegetais/ultraestrutura , Raízes de Plantas/ultraestrutura , Plântula/ultraestrutura
13.
Cell Host Microbe ; 16(3): 376-90, 2014 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-25211079

RESUMO

Nonsense-mediated mRNA decay (NMD) is a conserved eukaryotic RNA surveillance mechanism that degrades aberrant mRNAs. NMD impairment in Arabidopsis is linked to constitutive immune response activation and enhanced antibacterial resistance, but the underlying mechanisms are unknown. Here we show that NMD contributes to innate immunity in Arabidopsis by controlling the turnover of numerous TIR domain-containing, nucleotide-binding, leucine-rich repeat (TNL) immune receptor-encoding mRNAs. Autoimmunity resulting from NMD impairment depends on TNL signaling pathway components and can be triggered through deregulation of a single TNL gene, RPS6. Bacterial infection of plants causes host-programmed inhibition of NMD, leading to stabilization of NMD-regulated TNL transcripts. Conversely, constitutive NMD activity prevents TNL stabilization and impairs plant defense, demonstrating that host-regulated NMD contributes to disease resistance. Thus, NMD shapes plant innate immunity by controlling the threshold for activation of TNL resistance pathways.


Assuntos
Arabidopsis/genética , Degradação do RNAm Mediada por Códon sem Sentido , Pseudomonas syringae/fisiologia , Receptores Imunológicos/genética , Receptores Imunológicos/imunologia , Arabidopsis/imunologia , Arabidopsis/microbiologia , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/imunologia , Proteínas de Transporte/genética , Proteínas de Transporte/imunologia , Códon sem Sentido , Interações Hospedeiro-Patógeno , Doenças das Plantas/genética , Doenças das Plantas/imunologia , Doenças das Plantas/microbiologia , Pseudomonas syringae/genética , RNA Helicases/genética , RNA Helicases/imunologia , RNA Mensageiro/genética , RNA Mensageiro/imunologia
14.
J Cell Sci ; 121(Pt 13): 2208-16, 2008 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-18544632

RESUMO

Meiosis consists of two nuclear divisions that are separated by a short interkinesis. Here we show that the SMG7 protein, which plays an evolutionarily conserved role in nonsense-mediated RNA decay (NMD) in animals and yeast, is essential for the progression from anaphase to telophase in the second meiotic division in Arabidopsis. Arabidopsis SMG7 is an essential gene, the disruption of which causes embryonic lethality. Plants carrying a hypomorphic smg7 mutation exhibit an elevated level of transcripts containing premature stop codons. This suggests that the role of SMG7 in NMD is conserved in plants. Furthermore, hypomorphic smg7 alleles render mutant plants sterile by causing an unusual cell-cycle arrest in anaphase II that is characterized by delayed chromosome decondensation and aberrant rearrangement of the meiotic spindle. The smg7 phenotype was mimicked by exposing meiocytes to the proteasome inhibitor MG115. Together, these data indicate that SMG7 counteracts cyclin-dependent kinase (CDK) activity at the end of meiosis, and reveal a novel link between SMG7 and regulation of the meiotic cell cycle.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Proteínas de Transporte/metabolismo , Meiose , Estabilidade de RNA , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Transporte/genética , Códon sem Sentido , Quinases Ciclina-Dependentes/metabolismo , Mutação
15.
Mol Cell ; 27(1): 163-9, 2007 Jul 06.
Artigo em Inglês | MEDLINE | ID: mdl-17612498

RESUMO

Telomeres in mammals and plants are protected by the terminal t loop structure, the formation of which parallels the first steps of intrachromatid homologous recombination (HR). Under some circumstances, cells can also utilize an HR-based mechanism (alternative lengthening of telomeres [ALT]) as a back-up pathway for telomere maintenance. We have found that the Ku70/80 heterodimer, a central nonhomologous end-joining DNA repair factor, inhibits engagement of ALT in Arabidopsis telomerase-negative cells. To further assess HR activities at telomeres, we have developed a sensitive assay for detecting extrachromosomal telomeric circles (t circles) that may arise from t loop resolution and aberrant HR. We show that Ku70/80 specifically inhibits circle formation at telomeres, but not at centromeric and rDNA repeats. Ku inactivation results in increased formation of t circles that represent approximately 4% of total telomeric DNA. However, telomeres in ku mutants are fully functional, indicating that telomerase efficiently heals ongoing terminal deletions arising from excision of the t circles.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , DNA Helicases/metabolismo , DNA Circular/metabolismo , Proteínas de Ligação a DNA/metabolismo , Telômero/metabolismo , Cromossomos de Plantas/genética , Eletroforese em Gel Bidimensional , Mutação/genética
16.
J Cell Sci ; 119(Pt 12): 2486-96, 2006 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-16763194

RESUMO

Mnd1 has recently been identified in yeast as a key player in meiotic recombination. Here we describe the identification and functional characterisation of the Arabidopsis homologue, AtMND1, which is essential for male and female meiosis and thus for plant fertility. Although axial elements are formed normally, sister chromatid cohesion is established and recombination initiation appears to be unaffected in mutant plants, chromosomes do not synapse. During meiotic progression, a mass of entangled chromosomes, interconnected by chromatin bridges, and severe chromosome fragmentation are observed. These defects depend on the presence of SPO11-1, a protein that initiates recombination by catalysing DNA double-strand break (DSB) formation. Furthermore, we demonstrate that the AtMND1 protein interacts with AHP2, the Arabidopsis protein closely related to budding yeast Hop2. These data demonstrate that AtMND1 plays a key role in homologous synapsis and in DSB repair during meiotic recombination.


Assuntos
Proteínas de Arabidopsis/fisiologia , Arabidopsis/genética , Pareamento Cromossômico/fisiologia , Meiose/fisiologia , Recombinação Genética , Arabidopsis/fisiologia , Proteínas de Arabidopsis/genética , Pareamento Cromossômico/genética , Dano ao DNA , Mutação
17.
Int Arch Allergy Immunol ; 134(1): 1-9, 2004 May.
Artigo em Inglês | MEDLINE | ID: mdl-15051934

RESUMO

BACKGROUND: The major birch pollen allergen Bet v 1 (or Bet v 1a) is one of the main causes of seasonal type I allergies. Various environmental factors such as light, temperature and air pollution may influence the activity of the Bet v 1a gene. The creation of a model system to evaluate the role of environmental factors affecting the Bet v 1a gene expression would be highly desirable. We suggest the use of transgenic tobacco plants carrying a Bet v 1a promoter-reporter gene fusion as such a system. METHODS: The promoter of the Bet v 1a gene was isolated with the use of the Universal Genome Walker kit (BD Biosciences Clontech, USA). Web Software was used to search for putative cis-regulatory elements within the promoter. Transgenic tobacco plants harboring the promoter-beta-glucuronidase (GUS) reporter gene fusion were obtained via Agrobacterium tumefaciens-mediated transformation. Promoter activity was examined with histochemical and quantitative assays. RESULTS: Structural analysis predicted elements responsible for pollen-specific, light-, stress- and hormone-mediated induction within the Bet v 1a promoter. The evaluation of GUS activity in transgenic tobacco plants showed that the Bet v 1a promoter is pollen-specific. Moreover, the Bet v 1a promoter is considered to be the strongest isolated pollen-specific promoter reported to date. It was shown that temperature and abscisic acid positively regulate the activity of the Bet v 1a promoter during pollen development, providing evidence for environment-dependent regulation of the Bet v 1a gene. CONCLUSIONS: A model system to study the effect of environmental factors on the expression of the Bet v 1a gene encoding the major birch allergen in pollen was generated. Additionally, we suggest that this system could be used to search for factors that inhibit the activity of the gene in pollen in order to reduce the potential allergenicity of birch trees.


Assuntos
Poluentes Atmosféricos/imunologia , Alérgenos/biossíntese , Alérgenos/genética , Betula/genética , Betula/imunologia , Código Genético/genética , Proteínas de Plantas/biossíntese , Proteínas de Plantas/genética , Pólen/genética , Ácido Abscísico/farmacologia , Alérgenos/isolamento & purificação , Antígenos de Plantas , Sequência de Bases , DNA Complementar/genética , Fluorometria , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Regulação da Expressão Gênica de Plantas/genética , Genes de Plantas/efeitos dos fármacos , Genes de Plantas/genética , Glucuronidase/efeitos dos fármacos , Glucuronidase/genética , Humanos , Dados de Sequência Molecular , Reguladores de Crescimento de Plantas/farmacologia , Proteínas de Plantas/isolamento & purificação , Pólen/imunologia , Regiões Promotoras Genéticas/genética , Biossíntese de Proteínas/genética , Proteínas Recombinantes de Fusão/efeitos dos fármacos , Proteínas Recombinantes de Fusão/genética , Temperatura , Transcrição Gênica/genética
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