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1.
Nat Commun ; 12(1): 5855, 2021 10 06.
Artigo em Inglês | MEDLINE | ID: mdl-34615869

RESUMO

Karyotype alterations have emerged as on-target complications from CRISPR-Cas9 genome editing. However, the events that lead to these karyotypic changes in embryos after Cas9-treatment remain unknown. Here, using imaging and single-cell genome sequencing of 8-cell stage embryos, we track both spontaneous and Cas9-induced karyotype aberrations through the first three divisions of embryonic development. We observe the generation of abnormal structures of the nucleus that arise as a consequence of errors in mitosis, including micronuclei and chromosome bridges, and determine their contribution to common karyotype aberrations including whole chromosome loss that has been recently reported after editing in embryos. Together, these data demonstrate that Cas9-mediated germline genome editing can lead to unwanted on-target side effects, including major chromosome structural alterations that can be propagated over several divisions of embryonic development.


Assuntos
Sistemas CRISPR-Cas , Estruturas Cromossômicas , Edição de Genes/métodos , Instabilidade Genômica , Animais , Segregação de Cromossomos , Embrião de Mamíferos , Desenvolvimento Embrionário/genética , Cariótipo , Camundongos , Sequenciamento Completo do Genoma
2.
Nat Commun ; 12(1): 5221, 2021 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-34471117

RESUMO

Bacteria of the genus Streptomyces are prolific producers of specialized metabolites, including antibiotics. The linear chromosome includes a central region harboring core genes, as well as extremities enriched in specialized metabolite biosynthetic gene clusters. Here, we show that chromosome structure in Streptomyces ambofaciens correlates with genetic compartmentalization during exponential phase. Conserved, large and highly transcribed genes form boundaries that segment the central part of the chromosome into domains, whereas the terminal ends tend to be transcriptionally quiescent compartments with different structural features. The onset of metabolic differentiation is accompanied by a rearrangement of chromosome architecture, from a rather 'open' to a 'closed' conformation, in which highly expressed specialized metabolite biosynthetic genes form new boundaries. Thus, our results indicate that the linear chromosome of S. ambofaciens is partitioned into structurally distinct entities, suggesting a link between chromosome folding, gene expression and genome evolution.


Assuntos
Antibacterianos/metabolismo , Cromossomos Bacterianos , Streptomyces/genética , Streptomyces/metabolismo , Estruturas Cromossômicas , Regulação Bacteriana da Expressão Gênica , Genoma Bacteriano , Família Multigênica , Transcriptoma
3.
Elife ; 102021 07 19.
Artigo em Inglês | MEDLINE | ID: mdl-34279217

RESUMO

Supercoiling impacts DNA replication, transcription, protein binding to DNA, and the three-dimensional organization of chromosomes. However, there are currently no methods to directly interrogate or map positive supercoils, so their distribution in genomes remains unknown. Here, we describe a method, GapR-seq, based on the chromatin immunoprecipitation of GapR, a bacterial protein that preferentially recognizes overtwisted DNA, for generating high-resolution maps of positive supercoiling. Applying this method to Escherichia coli and Saccharomyces cerevisiae, we find that positive supercoiling is widespread, associated with transcription, and particularly enriched between convergently oriented genes, consistent with the 'twin-domain' model of supercoiling. In yeast, we also find positive supercoils associated with centromeres, cohesin-binding sites, autonomously replicating sites, and the borders of R-loops (DNA-RNA hybrids). Our results suggest that GapR-seq is a powerful approach, likely applicable in any organism, to investigate aspects of chromosome structure and organization not accessible by Hi-C or other existing methods.


Assuntos
Proteínas de Bactérias/genética , Imunoprecipitação da Cromatina , Estruturas Cromossômicas , Cromossomos/metabolismo , Proteínas de Bactérias/metabolismo , Sítios de Ligação , Proteínas de Ciclo Celular , Proteínas Cromossômicas não Histona , Cromossomos Bacterianos , DNA/metabolismo , Replicação do DNA , DNA Bacteriano , Escherichia coli/genética , Ligação Proteica , Saccharomyces cerevisiae/genética , Transcrição Genética
4.
Proc Natl Acad Sci U S A ; 118(11)2021 03 16.
Artigo em Inglês | MEDLINE | ID: mdl-33836591

RESUMO

White adipose tissue (WAT) is a key regulator of systemic energy metabolism, and impaired WAT plasticity characterized by enlargement of preexisting adipocytes associates with WAT dysfunction, obesity, and metabolic complications. However, the mechanisms that retain proper adipose tissue plasticity required for metabolic fitness are unclear. Here, we comprehensively showed that adipocyte-specific DNA methylation, manifested in enhancers and CTCF sites, directs distal enhancer-mediated transcriptomic features required to conserve metabolic functions of white adipocytes. Particularly, genetic ablation of adipocyte Dnmt1, the major methylation writer, led to increased adiposity characterized by increased adipocyte hypertrophy along with reduced expansion of adipocyte precursors (APs). These effects of Dnmt1 deficiency provoked systemic hyperlipidemia and impaired energy metabolism both in lean and obese mice. Mechanistically, Dnmt1 deficiency abrogated mitochondrial bioenergetics by inhibiting mitochondrial fission and promoted aberrant lipid metabolism in adipocytes, rendering adipocyte hypertrophy and WAT dysfunction. Dnmt1-dependent DNA methylation prevented aberrant CTCF binding and, in turn, sustained the proper chromosome architecture to permit interactions between enhancer and dynamin-1-like protein gene Dnm1l (Drp1) in adipocytes. Also, adipose DNMT1 expression inversely correlated with adiposity and markers of metabolic health but positively correlated with AP-specific markers in obese human subjects. Thus, these findings support strategies utilizing Dnmt1 action on mitochondrial bioenergetics in adipocytes to combat obesity and related metabolic pathology.


Assuntos
Adipócitos/metabolismo , DNA (Citosina-5-)-Metiltransferase 1/metabolismo , Epigênese Genética , Dinâmica Mitocondrial , Adipócitos/patologia , Tecido Adiposo/metabolismo , Tecido Adiposo/patologia , Adiposidade , Animais , Fator de Ligação a CCCTC/metabolismo , Estruturas Cromossômicas , DNA (Citosina-5-)-Metiltransferase 1/deficiência , DNA (Citosina-5-)-Metiltransferase 1/genética , Metilação de DNA , Dinaminas/genética , Dinaminas/metabolismo , Metabolismo Energético , Elementos Facilitadores Genéticos , Perfilação da Expressão Gênica , Metabolismo dos Lipídeos , Camundongos , Mitocôndrias/metabolismo , Obesidade/metabolismo , Obesidade/patologia , Regiões Promotoras Genéticas , Ligação Proteica
6.
Chromosome Res ; 29(1): 5-17, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33587223

RESUMO

This review describes image analyses for chromosome visible structures, focusing on the chromosome imaging system CHIAS (Chromosome Image Analyzing System). CHIAS is the first comprehensive imaging system for the analysis and characterization of plant chromosomes. A simulation method for human vision for capturing band positive regions was developed and used for the image analysis of large plant chromosomes with bands. Applying this method to C-banded Crepis chromosomes enabled recognition of band positive regions as seen by human vision. Furthermore, a new image parameter, condensation pattern was developed and successfully applied to identify small plant chromosomes such as rice and brassicas. Condensation profile (CP) derived from condensation pattern was also effective in developing quantitative chromosome maps. The result was quantitative chromosomal maps of several plants with small chromosomes, including Arabidopsis, diploid brassicas, rapeseed, rice, spinach, and sugarcane. In the final chapter, various applications of imaging techniques to the analysis of pachytene chromosomes, improved visibility of multicolor FISH images, 3D reconstruction of a human chromosome based on cross-section images obtained by a FIB/SEM, automatic extraction of chromosomal regions by machine learning, etc. are described.


Assuntos
Cromossomos de Plantas , Oryza , Estruturas Cromossômicas , Cromossomos de Plantas/genética , Humanos , Oryza/genética
7.
Mol Biol Evol ; 38(3): 904-910, 2021 03 09.
Artigo em Inglês | MEDLINE | ID: mdl-32986808

RESUMO

Microchromosomes are common yet poorly understood components of many vertebrate genomes. Recent studies have revealed that microchromosomes contain a high density of genes and possess other distinct characteristics compared with macrochromosomes. Whether distinctive characteristics of microchromosomes extend to features of genome structure and organization, however, remains an open question. Here, we analyze Hi-C sequencing data from multiple vertebrate lineages and show that microchromosomes exhibit consistently high degrees of interchromosomal interaction (particularly with other microchromosomes), appear to be colocalized to a common central nuclear territory, and are comprised of a higher proportion of open chromatin than macrochromosomes. These findings highlight an unappreciated level of diversity in vertebrate genome structure and function, and raise important questions regarding the evolutionary origins and ramifications of microchromosomes and the genes that they house.


Assuntos
Evolução Biológica , Estruturas Cromossômicas , Genoma , Vertebrados/genética , Animais
8.
Elife ; 92020 11 17.
Artigo em Inglês | MEDLINE | ID: mdl-33200984

RESUMO

Mutations of SMC5/6 components cause developmental defects, including primary microcephaly. To model neurodevelopmental defects, we engineered a mouse wherein Smc5 is conditionally knocked out (cKO) in the developing neocortex. Smc5 cKO mice exhibited neurodevelopmental defects due to neural progenitor cell (NPC) apoptosis, which led to reduction in cortical layer neurons. Smc5 cKO NPCs formed DNA bridges during mitosis and underwent chromosome missegregation. SMC5/6 depletion triggers a CHEK2-p53 DNA damage response, as concomitant deletion of the Trp53 tumor suppressor or Chek2 DNA damage checkpoint kinase rescued Smc5 cKO neurodevelopmental defects. Further assessment using Smc5 cKO and auxin-inducible degron systems demonstrated that absence of SMC5/6 leads to DNA replication stress at late-replicating regions such as pericentromeric heterochromatin. In summary, SMC5/6 is important for completion of DNA replication prior to entering mitosis, which ensures accurate chromosome segregation. Thus, SMC5/6 functions are critical in highly proliferative stem cells during organism development.


Assuntos
Proteínas de Ciclo Celular/metabolismo , Segregação de Cromossomos/fisiologia , Estruturas Cromossômicas/fisiologia , Neurogênese/fisiologia , Animais , Encéfalo/embriologia , Proteínas de Ciclo Celular/genética , Replicação do DNA , Embrião de Mamíferos , Desenvolvimento Embrionário , Regulação da Expressão Gênica no Desenvolvimento , Genótipo , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Mutação
9.
Nat Commun ; 11(1): 5495, 2020 10 30.
Artigo em Inglês | MEDLINE | ID: mdl-33127907

RESUMO

Protecting replication fork integrity during DNA replication is essential for maintaining genome stability. Here, we report that SDE2, a PCNA-associated protein, plays a key role in maintaining active replication and counteracting replication stress by regulating the replication fork protection complex (FPC). SDE2 directly interacts with the FPC component TIMELESS (TIM) and enhances its stability, thereby aiding TIM localization to replication forks and the coordination of replisome progression. Like TIM deficiency, knockdown of SDE2 leads to impaired fork progression and stalled fork recovery, along with a failure to activate CHK1 phosphorylation. Moreover, loss of SDE2 or TIM results in an excessive MRE11-dependent degradation of reversed forks. Together, our study uncovers an essential role for SDE2 in maintaining genomic integrity by stabilizing the FPC and describes a new role for TIM in protecting stalled replication forks. We propose that TIM-mediated fork protection may represent a way to cooperate with BRCA-dependent fork stabilization.


Assuntos
Proteínas de Ciclo Celular/metabolismo , Replicação do DNA/fisiologia , Proteínas de Ligação a DNA/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas de Ciclo Celular/genética , Quinase 1 do Ponto de Checagem/metabolismo , Estruturas Cromossômicas/metabolismo , Dano ao DNA , Reparo do DNA , Replicação do DNA/genética , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Regulação da Expressão Gênica , Técnicas de Silenciamento de Genes , Instabilidade Genômica/fisiologia , Células HEK293 , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteína Homóloga a MRE11/metabolismo , Proteínas Nucleares/metabolismo , Fosforilação , Domínios Proteicos
10.
Elife ; 92020 09 08.
Artigo em Inglês | MEDLINE | ID: mdl-32897188

RESUMO

DNA replication is needed to duplicate a cell's genome in S phase and segregate it during cell division. Previous work in Leishmania detected DNA replication initiation at just a single region in each chromosome, an organisation predicted to be insufficient for complete genome duplication within S phase. Here, we show that acetylated histone H3 (AcH3), base J and a kinetochore factor co-localise in each chromosome at only a single locus, which corresponds with previously mapped DNA replication initiation regions and is demarcated by localised G/T skew and G4 patterns. In addition, we describe previously undetected subtelomeric DNA replication in G2/M and G1-phase-enriched cells. Finally, we show that subtelomeric DNA replication, unlike chromosome-internal DNA replication, is sensitive to hydroxyurea and dependent on 9-1-1 activity. These findings indicate that Leishmania's genome duplication programme employs subtelomeric DNA replication initiation, possibly extending beyond S phase, to support predominantly chromosome-internal DNA replication initiation within S phase.


Assuntos
Estruturas Cromossômicas , Replicação do DNA/genética , Duplicação Gênica/genética , Genoma de Protozoário/genética , Leishmania major/genética , Estruturas Cromossômicas/química , Estruturas Cromossômicas/genética , Estruturas Cromossômicas/metabolismo , Cromossomos/química , Cromossomos/genética , Histonas/genética , Histonas/metabolismo , Fase S/genética
12.
Nat Commun ; 11(1): 4796, 2020 09 22.
Artigo em Inglês | MEDLINE | ID: mdl-32963231

RESUMO

Cortical thickness, surface area and volumes vary with age and cognitive function, and in neurological and psychiatric diseases. Here we report heritability, genetic correlations and genome-wide associations of these cortical measures across the whole cortex, and in 34 anatomically predefined regions. Our discovery sample comprises 22,824 individuals from 20 cohorts within the Cohorts for Heart and Aging Research in Genomic Epidemiology (CHARGE) consortium and the UK Biobank. We identify genetic heterogeneity between cortical measures and brain regions, and 160 genome-wide significant associations pointing to wnt/ß-catenin, TGF-ß and sonic hedgehog pathways. There is enrichment for genes involved in anthropometric traits, hindbrain development, vascular and neurodegenerative disease and psychiatric conditions. These data are a rich resource for studies of the biological mechanisms behind cortical development and aging.


Assuntos
Envelhecimento/genética , Encéfalo , Estudo de Associação Genômica Ampla , Transtornos Mentais/genética , Doenças Neurodegenerativas/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Estruturas Cromossômicas , Cognição , Feminino , Genômica , Humanos , Masculino , Pessoa de Meia-Idade , Fenótipo , Polimorfismo de Nucleotídeo Único
13.
Nucleic Acids Res ; 48(18): 10353-10367, 2020 10 09.
Artigo em Inglês | MEDLINE | ID: mdl-32926139

RESUMO

The vast majority of the genome is transcribed by RNA polymerases. G+C-rich regions of the chromosomes and negative superhelicity can promote the invasion of the DNA by RNA to form R-loops, which have been shown to block DNA replication and promote genome instability. However, it is unclear whether the R-loops themselves are sufficient to cause this instability or if additional factors are required. We have investigated replisome collisions with transcription complexes and R-loops using a reconstituted bacterial DNA replication system. RNA polymerase transcription complexes co-directionally oriented with the replication fork were transient blockages, whereas those oriented head-on were severe, stable blockages. On the other hand, replisomes easily bypassed R-loops on either template strand. Replication encounters with R-loops on the leading-strand template (co-directional) resulted in gaps in the nascent leading strand, whereas lagging-strand template R-loops (head-on) had little impact on replication fork progression. We conclude that whereas R-loops alone can act as transient replication blocks, most genome-destabilizing replication fork stalling likely occurs because of proteins bound to the R-loops.


Assuntos
Replicação do DNA/genética , Proteínas de Ligação a DNA/genética , Estruturas R-Loop/genética , Transcrição Genética , Composição de Bases/genética , Estruturas Cromossômicas/genética , Cromossomos/genética , DNA Helicases/genética , Reparo do DNA/genética , Escherichia coli/genética , Instabilidade Genômica/genética
14.
Nat Commun ; 11(1): 3888, 2020 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-32753666

RESUMO

First proposed as antimicrobial agents, histones were later recognized for their role in condensing chromosomes. Histone antimicrobial activity has been reported in innate immune responses. However, how histones kill bacteria has remained elusive. The co-localization of histones with antimicrobial peptides (AMPs) in immune cells suggests that histones may be part of a larger antimicrobial mechanism in vivo. Here we report that histone H2A enters E. coli and S. aureus through membrane pores formed by the AMPs LL-37 and magainin-2. H2A enhances AMP-induced pores, depolarizes the bacterial membrane potential, and impairs membrane recovery. Inside the cytoplasm, H2A reorganizes bacterial chromosomal DNA and inhibits global transcription. Whereas bacteria recover from the pore-forming effects of LL-37, the concomitant effects of H2A and LL-37 are irrecoverable. Their combination constitutes a positive feedback loop that exponentially amplifies their antimicrobial activities, causing antimicrobial synergy. More generally, treatment with H2A and the pore-forming antibiotic polymyxin B completely eradicates bacterial growth.


Assuntos
Anti-Infecciosos/farmacologia , Bactérias/efeitos dos fármacos , Bactérias/genética , Estruturas Cromossômicas/efeitos dos fármacos , Histonas/metabolismo , Prótons , Animais , Estruturas Cromossômicas/metabolismo , Cromossomos Bacterianos/metabolismo , DNA Bacteriano/metabolismo , Sinergismo Farmacológico , Escherichia coli/efeitos dos fármacos , Escherichia coli/metabolismo , Imunidade Inata , Mamíferos , Polimixina B/farmacologia , Análise de Sequência de RNA , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/metabolismo
15.
Life Sci Alliance ; 3(10)2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32820027

RESUMO

Most tumors lack the G1/S phase checkpoint and are insensitive to antigrowth signals. Loss of G1/S control can severely perturb DNA replication as revealed by slow replication fork progression and frequent replication fork stalling. Cancer cells may thus rely on specific pathways that mitigate the deleterious consequences of replication stress. To identify vulnerabilities of cells suffering from replication stress, we performed an shRNA-based genetic screen. We report that the RECQL helicase is specifically essential in replication stress conditions and protects stalled replication forks against MRE11-dependent double strand break (DSB) formation. In line with these findings, knockdown of RECQL in different cancer cells increased the level of DNA DSBs. Thus, RECQL plays a critical role in sustaining DNA synthesis under conditions of replication stress and as such may represent a target for cancer therapy.


Assuntos
Reparo do DNA/fisiologia , Replicação do DNA/fisiologia , RecQ Helicases/metabolismo , Animais , Linhagem Celular Tumoral , Estruturas Cromossômicas/metabolismo , DNA , Quebras de DNA de Cadeia Dupla , Reparo do DNA/genética , Proteínas de Ligação a DNA/genética , Instabilidade Genômica/genética , Humanos , Proteína Homóloga a MRE11/genética , Camundongos , RNA Interferente Pequeno/genética , Rad51 Recombinase/genética , RecQ Helicases/fisiologia
16.
Nat Commun ; 11(1): 4345, 2020 08 28.
Artigo em Inglês | MEDLINE | ID: mdl-32859945

RESUMO

Chromosome movements and programmed DNA double-strand breaks (DSBs) promote homologue pairing and initiate recombination at meiosis onset. Meiotic progression involves checkpoint-controlled termination of these events when all homologue pairs achieve synapsis and form crossover precursors. Exploiting the temporo-spatial organisation of the C. elegans germline and time-resolved methods of protein removal, we show that surveillance of the synaptonemal complex (SC) controls meiotic progression. In nuclei with fully synapsed homologues and crossover precursors, removing different meiosis-specific cohesin complexes, which are individually required for SC stability, or a SC central region component causes functional redeployment of the chromosome movement and DSB machinery, triggering whole-nucleus reorganisation. This apparent reversal of the meiotic programme requires CHK-2 kinase reactivation via signalling from chromosome axes containing HORMA proteins, but occurs in the absence of transcriptional changes. Our results uncover an unexpected plasticity of the meiotic programme and show how chromosome signalling orchestrates nuclear organisation and meiotic progression.


Assuntos
Caenorhabditis elegans/genética , Proteínas de Ciclo Celular/metabolismo , Proteínas Cromossômicas não Histona/metabolismo , Estruturas Cromossômicas/metabolismo , Meiose/fisiologia , Animais , Proteínas de Caenorhabditis elegans/metabolismo , Pontos de Checagem do Ciclo Celular , Quinase do Ponto de Checagem 2/metabolismo , Pareamento Cromossômico , Quebras de DNA de Cadeia Dupla , Complexo Sinaptonêmico/metabolismo
17.
Nucleic Acids Res ; 48(15): 8461-8473, 2020 09 04.
Artigo em Inglês | MEDLINE | ID: mdl-32633759

RESUMO

DNA polymerase ζ (Pol ζ) and Rev1 are essential for the repair of DNA interstrand crosslink (ICL) damage. We have used yeast DNA polymerases η, ζ and Rev1 to study translesion synthesis (TLS) past a nitrogen mustard-based interstrand crosslink (ICL) with an 8-atom linker between the crosslinked bases. The Rev1-Pol ζ complex was most efficient in complete bypass synthesis, by 2-3 fold, compared to Pol ζ alone or Pol η. Rev1 protein, but not its catalytic activity, was required for efficient TLS. A dCMP residue was faithfully inserted across the ICL-G by Pol η, Pol ζ, and Rev1-Pol ζ. Rev1-Pol ζ, and particularly Pol ζ alone showed a tendency to stall before the ICL, whereas Pol η stalled just after insertion across the ICL. The stalling of Pol η directly past the ICL is attributed to its autoinhibitory activity, caused by elongation of the short ICL-unhooked oligonucleotide (a six-mer in our study) by Pol η providing a barrier to further elongation of the correct primer. No stalling by Rev1-Pol ζ directly past the ICL was observed, suggesting that the proposed function of Pol ζ as an extender DNA polymerase is also required for ICL repair.


Assuntos
DNA Polimerase Dirigida por DNA/genética , DNA/genética , Nucleotidiltransferases/genética , Proteínas de Saccharomyces cerevisiae/genética , Estruturas Cromossômicas/efeitos dos fármacos , Estruturas Cromossômicas/genética , Dano ao DNA/efeitos dos fármacos , Dano ao DNA/genética , Reparo do DNA/efeitos dos fármacos , Reparo do DNA/genética , Replicação do DNA/genética , Complexos Multiproteicos/genética , Compostos de Mostarda Nitrogenada/farmacologia , Saccharomyces cerevisiae/genética
18.
Nat Commun ; 11(1): 3531, 2020 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-32669601

RESUMO

Homologous recombination (HR) factors were recently implicated in DNA replication fork remodeling and protection. While maintaining genome stability, HR-mediated fork remodeling promotes cancer chemoresistance, by as-yet elusive mechanisms. Five HR cofactors - the RAD51 paralogs RAD51B, RAD51C, RAD51D, XRCC2 and XRCC3 - recently emerged as crucial tumor suppressors. Albeit extensively characterized in DNA repair, their role in replication has not been addressed systematically. Here, we identify all RAD51 paralogs while screening for modulators of RAD51 recombinase upon replication stress. Single-molecule analysis of fork progression and architecture in isogenic cellular systems shows that the BCDX2 subcomplex restrains fork progression upon stress, promoting fork reversal. Accordingly, BCDX2 primes unscheduled degradation of reversed forks in BRCA2-defective cells, boosting genomic instability. Conversely, the CX3 subcomplex is dispensable for fork reversal, but mediates efficient restart of reversed forks. We propose that RAD51 paralogs sequentially orchestrate clinically relevant transactions at replication forks, cooperatively promoting fork remodeling and restart.


Assuntos
Replicação do DNA , Rad51 Recombinase/metabolismo , Proteína BRCA2/metabolismo , Linhagem Celular Tumoral , Estruturas Cromossômicas/metabolismo , Cromossomos/ultraestrutura , Dano ao DNA , Reparo do DNA , Proteínas de Ligação a DNA/metabolismo , Instabilidade Genômica , Recombinação Homóloga , Humanos , Microscopia , Mutagênicos , Mutação , Osteossarcoma/metabolismo , RNA Interferente Pequeno/metabolismo
19.
J Immunol ; 204(10): 2617-2626, 2020 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-32366683

RESUMO

The past decade has increased our understanding of how genome topology controls RAG endonuclease-mediated assembly of lymphocyte AgR genes. New technologies have illuminated how the large IgH, Igκ, TCRα/δ, and TCRß loci fold into compact structures that place their numerous V gene segments in similar three-dimensional proximity to their distal recombination center composed of RAG-bound (D)J gene segments. Many studies have shown that CTCF and cohesin protein-mediated chromosome looping have fundamental roles in lymphocyte lineage- and developmental stage-specific locus compaction as well as broad usage of V segments. CTCF/cohesin-dependent loops have also been shown to direct and restrict RAG activity within chromosome domains. We summarize recent work in elucidating molecular mechanisms that govern three-dimensional chromosome organization and in investigating how these dynamic mechanisms control V(D)J recombination. We also introduce remaining questions for how CTCF/cohesin-dependent and -independent genome architectural mechanisms might regulate compaction and recombination of AgR loci.


Assuntos
Linfócitos B/imunologia , Receptores de Antígenos/genética , Linfócitos T/imunologia , Recombinação V(D)J/genética , Animais , Fator de Ligação a CCCTC/metabolismo , Proteínas de Ciclo Celular/metabolismo , Montagem e Desmontagem da Cromatina , Proteínas Cromossômicas não Histona/metabolismo , Estruturas Cromossômicas , Loci Gênicos , Humanos , Conformação Molecular , Receptores de Antígenos/metabolismo
20.
Nucleus ; 11(9): 111-116, 2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-32412326

RESUMO

In S. cerevisiae prophase meiotic chromosomes move by forces generated in the cytoplasm and transduced to the telomere via a protein complex located in the nuclear membrane. We know that chromosome movements require actin cytoskeleton [13,31] and the proteins Ndj1, Mps3, and Csm4. Until recently, the identity of the protein connecting Ndj1-Mps3 with the cytoskeleton components was missing. It was also not known the identity of a cytoplasmic motor responsible for interacting with the actin cytoskeleton and a protein at the outer nuclear envelope. Our recent work [36] identified Mps2 as the protein connecting Ndj1-Mps3 with cytoskeleton components; Myo2 as the cytoplasmic motor that interacts with Mps2; and Cms4 as a regulator of Mps2 and Myo2 interaction and activities (Figure 1). Below we present a model for how Mps2, Csm4, and Myo2 promote chromosome movements by providing the primary connections joining telomeres to the actin cytoskeleton through the LINC complex.


Assuntos
Cromossomos Fúngicos , Meiose , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Telômero/metabolismo , Citoesqueleto de Actina/metabolismo , Estruturas Cromossômicas , Cromossomos Fúngicos/genética , Cromossomos Fúngicos/metabolismo , Meiose/genética , Modelos Moleculares , Saccharomyces cerevisiae/citologia , Proteínas de Saccharomyces cerevisiae/metabolismo , Telômero/genética
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