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1.
Cell Chem Biol ; 31(1): 139-149.e14, 2024 01 18.
Artigo em Inglês | MEDLINE | ID: mdl-37967558

RESUMO

A novel class of benzoxaboroles was reported to induce cancer cell death but the mechanism was unknown. Using a forward genetics platform, we discovered mutations in cleavage and polyadenylation specific factor 3 (CPSF3) that reduce benzoxaborole binding and confer resistance. CPSF3 is the endonuclease responsible for pre-mRNA 3'-end processing, which is also important for RNA polymerase II transcription termination. Benzoxaboroles inhibit this endonuclease activity of CPSF3 in vitro and also curb transcriptional termination in cells, which results in the downregulation of numerous constitutively expressed genes. Furthermore, we used X-ray crystallography to demonstrate that benzoxaboroles bind to the active site of CPSF3 in a manner distinct from the other known inhibitors of CPSF3. The benzoxaborole compound impeded the growth of cancer cell lines derived from different lineages. Our results suggest benzoxaboroles may represent a promising lead as CPSF3 inhibitors for clinical development.


Assuntos
Antineoplásicos , Compostos de Boro , Fator de Especificidade de Clivagem e Poliadenilação , Endonucleases , Precursores de RNA , Processamento Pós-Transcricional do RNA , Fator de Especificidade de Clivagem e Poliadenilação/antagonistas & inibidores , Fator de Especificidade de Clivagem e Poliadenilação/química , Endonucleases/antagonistas & inibidores , Precursores de RNA/genética , Precursores de RNA/metabolismo , Compostos de Boro/química , Compostos de Boro/farmacologia , Antineoplásicos/química , Antineoplásicos/farmacologia , Processamento Pós-Transcricional do RNA/efeitos dos fármacos , Humanos , Linhagem Celular Tumoral
2.
Cell Rep ; 27(3): 776-792.e7, 2019 04 16.
Artigo em Inglês | MEDLINE | ID: mdl-30995476

RESUMO

Many viruses shut off host gene expression to inhibit antiviral responses. Viral proteins and host proteins required for viral replication are typically spared in this process, but the mechanisms of target selectivity during host shutoff remain poorly understood. Using transcriptome-wide and targeted reporter experiments, we demonstrate that the influenza A virus endoribonuclease PA-X usurps RNA splicing to selectively target host RNAs for destruction. Proximity-labeling proteomics reveals that PA-X interacts with cellular RNA processing proteins, some of which are partially required for host shutoff. Thus, PA-X taps into host nuclear pre-mRNA processing mechanisms to destroy nascent mRNAs shortly after their synthesis. This mechanism sets PA-X apart from other viral host shutoff proteins that target actively translating mRNAs in the cytoplasm. Our study reveals a unique mechanism of host shutoff that helps us understand how influenza viruses suppress host gene expression.


Assuntos
Vírus da Influenza A/fisiologia , Splicing de RNA , RNA Mensageiro/metabolismo , Proteínas Repressoras/metabolismo , Proteínas não Estruturais Virais/metabolismo , Células A549 , Fator de Especificidade de Clivagem e Poliadenilação/antagonistas & inibidores , Fator de Especificidade de Clivagem e Poliadenilação/genética , Fator de Especificidade de Clivagem e Poliadenilação/metabolismo , Regulação para Baixo , Endorribonucleases/metabolismo , Células HEK293 , Interações Hospedeiro-Patógeno , Humanos , Interferons/genética , Interferons/metabolismo , Mutagênese Sítio-Dirigida , Interferência de RNA , Precursores de RNA/metabolismo , Sítios de Splice de RNA , RNA Interferente Pequeno/metabolismo , Proteínas Repressoras/genética , Regulação para Cima , Proteínas não Estruturais Virais/genética , Fatores de Poliadenilação e Clivagem de mRNA/antagonistas & inibidores , Fatores de Poliadenilação e Clivagem de mRNA/genética , Fatores de Poliadenilação e Clivagem de mRNA/metabolismo
3.
Proc Natl Acad Sci U S A ; 115(38): 9616-9621, 2018 09 18.
Artigo em Inglês | MEDLINE | ID: mdl-30185555

RESUMO

African trypanosomes cause lethal and neglected tropical diseases, known as sleeping sickness in humans and nagana in animals. Current therapies are limited, but fortunately, promising therapies are in advanced clinical and veterinary development, including acoziborole (AN5568 or SCYX-7158) and AN11736, respectively. These benzoxaboroles will likely be key to the World Health Organization's target of disease control by 2030. Their mode of action was previously unknown. We have developed a high-coverage overexpression library and use it here to explore drug mode of action in Trypanosoma brucei Initially, an inhibitor with a known target was used to select for drug resistance and to test massive parallel library screening and genome-wide mapping; this effectively identified the known target and validated the approach. Subsequently, the overexpression screening approach was used to identify the target of the benzoxaboroles, Cleavage and Polyadenylation Specificity Factor 3 (CPSF3, Tb927.4.1340). We validated the CPSF3 endonuclease as the target, using independent overexpression strains. Knockdown provided genetic validation of CPSF3 as essential, and GFP tagging confirmed the expected nuclear localization. Molecular docking and CRISPR-Cas9-based editing demonstrated how acoziborole can specifically block the active site and mRNA processing by parasite, but not host CPSF3. Thus, our findings provide both genetic and chemical validation for CPSF3 as an important drug target in trypanosomes and reveal inhibition of mRNA maturation as the mode of action of the trypanocidal benzoxaboroles. Understanding the mechanism of action of benzoxaborole-based therapies can assist development of improved therapies, as well as the prediction and monitoring of resistance, if or when it arises.


Assuntos
Fator de Especificidade de Clivagem e Poliadenilação/antagonistas & inibidores , Proteínas de Protozoários/antagonistas & inibidores , Tripanossomicidas/farmacologia , Trypanosoma brucei brucei/fisiologia , Tripanossomíase Africana/prevenção & controle , Animais , Benzamidas/farmacologia , Benzamidas/uso terapêutico , Compostos de Boro/farmacologia , Compostos de Boro/uso terapêutico , Sistemas CRISPR-Cas , Núcleo Celular/genética , Núcleo Celular/metabolismo , Fator de Especificidade de Clivagem e Poliadenilação/genética , Fator de Especificidade de Clivagem e Poliadenilação/metabolismo , Resistência a Medicamentos/efeitos dos fármacos , Resistência a Medicamentos/genética , Técnicas de Silenciamento de Genes , Biblioteca Gênica , Ensaios de Triagem em Larga Escala/métodos , Humanos , Simulação de Acoplamento Molecular , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , Processamento Pós-Transcricional do RNA/efeitos dos fármacos , RNA Mensageiro/metabolismo , RNA de Protozoário/metabolismo , Tripanossomicidas/uso terapêutico , Trypanosoma brucei brucei/efeitos dos fármacos , Tripanossomíase Africana/transmissão , Tripanossomíase Africana/veterinária , Valina/análogos & derivados , Valina/farmacologia , Valina/uso terapêutico
4.
Genes Dev ; 32(2): 127-139, 2018 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-29432121

RESUMO

Termination is a ubiquitous phase in every transcription cycle but is incompletely understood and a subject of debate. We used gene editing as a new approach to address its mechanism through engineered conditional depletion of the 5' → 3' exonuclease Xrn2 or the polyadenylation signal (PAS) endonuclease CPSF73 (cleavage and polyadenylation specificity factor 73). The ability to rapidly control Xrn2 reveals a clear and general role for it in cotranscriptional degradation of 3' flanking region RNA and transcriptional termination. This defect is characterized genome-wide at high resolution using mammalian native elongating transcript sequencing (mNET-seq). An Xrn2 effect on termination requires prior RNA cleavage, and we provide evidence for this by showing that catalytically inactive CPSF73 cannot restore termination to cells lacking functional CPSF73. Notably, Xrn2 plays no significant role in either Histone or small nuclear RNA (snRNA) gene termination even though both RNA classes undergo 3' end cleavage. In sum, efficient termination on most protein-coding genes involves CPSF73-mediated RNA cleavage and cotranscriptional degradation of polymerase-associated RNA by Xrn2. However, as CPSF73 loss caused more extensive readthrough transcription than Xrn2 elimination, it likely plays a more underpinning role in termination.


Assuntos
Fator de Especificidade de Clivagem e Poliadenilação/metabolismo , Exorribonucleases/fisiologia , RNA Polimerase II/metabolismo , Terminação da Transcrição Genética , Regiões 3' não Traduzidas , Linhagem Celular , Fator de Especificidade de Clivagem e Poliadenilação/antagonistas & inibidores , Fator de Especificidade de Clivagem e Poliadenilação/genética , Exorribonucleases/antagonistas & inibidores , Exorribonucleases/metabolismo , Sequenciamento de Nucleotídeos em Larga Escala , Histonas , Humanos , Ácidos Indolacéticos/farmacologia , Mutação , RNA Nuclear Pequeno/genética , Análise de Sequência de RNA
5.
Nat Commun ; 8: 14574, 2017 03 06.
Artigo em Inglês | MEDLINE | ID: mdl-28262680

RESUMO

Benzoxaboroles are effective against bacterial, fungal and protozoan pathogens. We report potent activity of the benzoxaborole AN3661 against Plasmodium falciparum laboratory-adapted strains (mean IC50 32 nM), Ugandan field isolates (mean ex vivo IC50 64 nM), and murine P. berghei and P. falciparum infections (day 4 ED90 0.34 and 0.57 mg kg-1, respectively). Multiple P. falciparum lines selected in vitro for resistance to AN3661 harboured point mutations in pfcpsf3, which encodes a homologue of mammalian cleavage and polyadenylation specificity factor subunit 3 (CPSF-73 or CPSF3). CRISPR-Cas9-mediated introduction of pfcpsf3 mutations into parental lines recapitulated AN3661 resistance. PfCPSF3 homology models placed these mutations in the active site, where AN3661 is predicted to bind. Transcripts for three trophozoite-expressed genes were lost in AN3661-treated trophozoites, which was not observed in parasites selected or engineered for AN3661 resistance. Our results identify the pre-mRNA processing factor PfCPSF3 as a promising antimalarial drug target.


Assuntos
Antimaláricos/farmacologia , Compostos de Boro/farmacologia , Fator de Especificidade de Clivagem e Poliadenilação/química , Plasmodium falciparum/efeitos dos fármacos , Proteínas de Protozoários/química , RNA Mensageiro/genética , Sequência de Aminoácidos , Animais , Antimaláricos/síntese química , Compostos de Boro/síntese química , Sistemas CRISPR-Cas , Domínio Catalítico , Fator de Especificidade de Clivagem e Poliadenilação/antagonistas & inibidores , Fator de Especificidade de Clivagem e Poliadenilação/genética , Fator de Especificidade de Clivagem e Poliadenilação/metabolismo , Resistência a Medicamentos/genética , Eritrócitos/efeitos dos fármacos , Eritrócitos/parasitologia , Edição de Genes/métodos , Humanos , Malária/tratamento farmacológico , Malária/parasitologia , Malária Falciparum/tratamento farmacológico , Malária Falciparum/parasitologia , Camundongos , Simulação de Acoplamento Molecular , Mutação , Plasmodium berghei/efeitos dos fármacos , Plasmodium berghei/genética , Plasmodium berghei/crescimento & desenvolvimento , Plasmodium berghei/metabolismo , Plasmodium falciparum/genética , Plasmodium falciparum/crescimento & desenvolvimento , Plasmodium falciparum/metabolismo , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Estrutura Secundária de Proteína , Proteínas de Protozoários/antagonistas & inibidores , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , RNA Mensageiro/metabolismo , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Trofozoítos/efeitos dos fármacos , Trofozoítos/genética , Trofozoítos/crescimento & desenvolvimento , Trofozoítos/metabolismo
6.
EMBO Mol Med ; 9(3): 385-394, 2017 03.
Artigo em Inglês | MEDLINE | ID: mdl-28148555

RESUMO

Toxoplasma gondii is an important food and waterborne pathogen causing toxoplasmosis, a potentially severe disease in immunocompromised or congenitally infected humans. Available therapeutic agents are limited by suboptimal efficacy and frequent side effects that can lead to treatment discontinuation. Here we report that the benzoxaborole AN3661 had potent in vitro activity against T. gondii Parasites selected to be resistant to AN3661 had mutations in TgCPSF3, which encodes a homologue of cleavage and polyadenylation specificity factor subunit 3 (CPSF-73 or CPSF3), an endonuclease involved in mRNA processing in eukaryotes. Point mutations in TgCPSF3 introduced into wild-type parasites using the CRISPR/Cas9 system recapitulated the resistance phenotype. Importantly, mice infected with T. gondii and treated orally with AN3661 did not develop any apparent illness, while untreated controls had lethal infections. Therefore, TgCPSF3 is a promising novel target of T. gondii that provides an opportunity for the development of anti-parasitic drugs.


Assuntos
Antiprotozoários/farmacologia , Compostos de Boro/farmacologia , Fator de Especificidade de Clivagem e Poliadenilação/antagonistas & inibidores , Toxoplasma/efeitos dos fármacos , Toxoplasma/enzimologia , Toxoplasmose/tratamento farmacológico , Administração Oral , Animais , Antiprotozoários/administração & dosagem , Compostos de Boro/administração & dosagem , Modelos Animais de Doenças , Resistência a Medicamentos , Camundongos , Testes de Sensibilidade Parasitária , Mutação Puntual , Análise de Sobrevida
7.
J Biosci ; 42(3): 417-425, 2017 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-29358555

RESUMO

Cleavage and polyadenylation specificity factor 1 (CPSF1), a member of CPSF complex, has been reported to play a key role in pre-mRNA 3'-end formation, but its possible role in ovarian cancer remains unclear. In the present study, we found the mRNA level of CPSF1 was overexpressed in ovarian cancer tissues using Oncomine Cancer Microarray database. Then the loss-of-function assays, including CCK-8, colony formation and flow cytometry assays, were performed to determine the effects of CPSF1 on cell viability, proliferation, cell cycle and apoptosis of human ovarian cancer cell lines (SKOV-3 and OVCAR-3). The results indicated that depletion of CPSF1 suppressed cell viability, impaired colony formation ability, induced cell cycle arrest at G0/G1 phase and promoted cell apoptosis in ovarian cancer cells. Furthermore, knockdown of CPSF1 upregulated the expression of cleaved caspase-3 and PARP and downregulated CDK4/cyclin D1 expression. These data suggested that CPSF1 could promote ovarian cancer cell growth and proliferation in vitro and its depletion might serve as a potential therapeutic target for human ovarian cancer.


Assuntos
Fator de Especificidade de Clivagem e Poliadenilação/genética , Células Epiteliais/metabolismo , Pontos de Checagem da Fase G1 do Ciclo Celular/genética , Regulação Neoplásica da Expressão Gênica , Apoptose/genética , Caspase 3/genética , Caspase 3/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Fator de Especificidade de Clivagem e Poliadenilação/antagonistas & inibidores , Fator de Especificidade de Clivagem e Poliadenilação/metabolismo , Ciclina D1/genética , Ciclina D1/metabolismo , Quinase 4 Dependente de Ciclina/genética , Quinase 4 Dependente de Ciclina/metabolismo , Células Epiteliais/patologia , Feminino , Humanos , Ovário/metabolismo , Ovário/patologia , Poli(ADP-Ribose) Polimerases/genética , Poli(ADP-Ribose) Polimerases/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Transdução de Sinais
8.
J Mol Model ; 20(3): 2142, 2014 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-24562912

RESUMO

Inhibition of CPSF30 function by the effector domain of influenza A virus of non-structural protein 1 (NS1A) protein plays a critical role in the suppression of host key antiviral response. The CPSF30-binding site of NS1A appears to be a very attractive target for the development of new drugs against influenza A virus. In this study, structure-based molecular docking was utilized to screen more than 30,000 compounds from a Traditional Chinese Medicine (TCM) database. Four drug-like compounds were selected as potential inhibitors for the CPSF30-binding site of NS1A. Docking conformation analysis results showed that these potential inhibitors could bind to the CPSF30-binding site with strong hydrophobic interactions and weak hydrogen bonds. Molecular dynamics simulations and MM-PBSA calculations suggested that two of the inhibitors, compounds 32056 and 31674, could stably bind to the CPSF30-binding site with high binding free energy. These two compounds could be modified to achieve higher binding affinity, so that they may be used as potential leads in the development of new anti-influenza drugs.


Assuntos
Antivirais/química , Fator de Especificidade de Clivagem e Poliadenilação/química , Medicina Tradicional Chinesa , Proteínas não Estruturais Virais/química , Algoritmos , Antivirais/metabolismo , Antivirais/farmacologia , Sítios de Ligação , Fator de Especificidade de Clivagem e Poliadenilação/antagonistas & inibidores , Fator de Especificidade de Clivagem e Poliadenilação/metabolismo , Bases de Dados Factuais , Interações Hospedeiro-Patógeno/efeitos dos fármacos , Humanos , Vírus da Influenza A Subtipo H1N1/efeitos dos fármacos , Vírus da Influenza A Subtipo H1N1/metabolismo , Vírus da Influenza A Subtipo H1N1/fisiologia , Influenza Humana/prevenção & controle , Influenza Humana/virologia , Cinética , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Estrutura Molecular , Ligação Proteica/efeitos dos fármacos , Estrutura Terciária de Proteína , Proteínas não Estruturais Virais/metabolismo
9.
Oncogene ; 28(1): 41-51, 2009 Jan 08.
Artigo em Inglês | MEDLINE | ID: mdl-18806823

RESUMO

CSR1 (cellular stress response 1), a newly characterized tumor-suppressor gene, undergoes hypermethylation in over 30% of prostate cancers. Re-expression of CSR1 inhibits cell growth and induces cell death, but the mechanism by which CSR1 suppresses tumor growth is not clear. In this study, we screened a prostate cDNA library using a yeast two-hybrid system and found that the cleavage and polyadenylation-specific factor 3 (CPSF3), an essential component for converting heteronuclear RNA to mRNA, binds with high affinity to the CSR1 C terminus. Further analyses determined that the binding motifs for CPSF3 are located between amino acids 440 and 543. The interaction between CSR1 and CPSF3 induced CPSF3 translocation from the nucleus to the cytoplasm, resulting in inhibition of polyadenylation both in vitro and in vivo. Downregulation of CPSF3 using small interfering RNA induced cell death in a manner similar to CSR1 expression. A CSR1 mutant unable to bind to CPSF3 did not alter CPSF3 subcellular distribution, did not inhibit its polyadenylation activity and did not induce cell death. In summary, CSR1 appears to induce cell death through a novel mechanism by hijacking a critical RNA processing enzyme.


Assuntos
Apoptose , Fator de Especificidade de Clivagem e Poliadenilação/antagonistas & inibidores , Proteínas de Choque Térmico/metabolismo , Próstata/metabolismo , Receptores Depuradores Classe A/metabolismo , Linhagem Celular , Núcleo Celular/metabolismo , Fator de Especificidade de Clivagem e Poliadenilação/genética , Fator de Especificidade de Clivagem e Poliadenilação/metabolismo , Citoplasma/metabolismo , Biblioteca Gênica , Proteínas de Choque Térmico/genética , Humanos , Masculino , Poliadenilação , Transporte Proteico , RNA Interferente Pequeno/genética , Receptores Depuradores Classe A/genética , Técnicas do Sistema de Duplo-Híbrido
10.
J Mol Biol ; 372(2): 317-30, 2007 Sep 14.
Artigo em Inglês | MEDLINE | ID: mdl-17669424

RESUMO

Gene expression in eukaryotes requires the post-transcriptional cleavage of mRNA precursors into mature mRNAs. The cleavage and polyadenylation specificity factor (CPSF) is critical for this process and its 73 kDa subunit (CPSF-73) mediates cleavage coupled to polyadenylation and histone pre-mRNA processing. Using CPSF-73 over-expression and siRNA-mediated knockdown experiments, this study identifies CPSF-73 as an important regulatory protein that represses the basal transcriptional activity of the HIV-1 LTR promoter. Similar results were found with over-expression of the CPSF-73 homologue RC-68, but not with CPSF 100 kDa subunit (CPSF-100) and RC-74. Chromatin immunoprecipitation assays revealed the physical interaction of CPSF-73 with the HIV-1 LTR promoter. Further experiments revealed indirect CPSF-73 binding to the region between -275 to -110 within the 5' upstream region. Functional assays revealed the importance for the 5' upstream region (-454 to -110) of the LTR for CPSF-73-mediated transcription repression. We also show that HIV-1 Tat protein interacts with CPSF-73 and counteracts its repressive activity on the HIV-1 LTR promoter. Our results clearly show a novel function for CPSF-73 and add another candidate protein for explaining the molecular mechanisms underlying HIV-1 latency.


Assuntos
Fator de Especificidade de Clivagem e Poliadenilação/antagonistas & inibidores , Fator de Especificidade de Clivagem e Poliadenilação/metabolismo , Produtos do Gene tat/metabolismo , Repetição Terminal Longa de HIV/genética , Regiões Promotoras Genéticas/genética , Imunoprecipitação da Cromatina , Fator de Especificidade de Clivagem e Poliadenilação/química , Humanos , Ligação Proteica , Subunidades Proteicas/antagonistas & inibidores , Subunidades Proteicas/química , Subunidades Proteicas/metabolismo , Proteínas Repressoras/antagonistas & inibidores , Proteínas Repressoras/química , Proteínas Repressoras/metabolismo , Latência Viral
11.
Nat Struct Mol Biol ; 14(7): 662-9, 2007 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-17572685

RESUMO

Eukaryotic poly(A) signals direct mRNA 3'-end processing and also pausing and termination of transcription. We show that pausing and termination require the processing factor CPSF, which binds the AAUAAA hexamer of the mammalian poly(A) signal. Pausing does not require the RNA polymerase II C-terminal domain (CTD) or the cleavage stimulation factor, CstF, that binds the CTD. Pull-down experiments show that CPSF binds, principally through its 30-kDa subunit, to the body of the polymerase. CPSF can also bind CstF, but this seems to be mutually exclusive with polymerase binding. We suggest that CPSF, while binding the body of the polymerase, scans for hexamers in the extruding RNA. Any encounter with a hexamer triggers pausing. If the hexamer is part of a functional poly(A) signal, CstF is recruited and binds CPSF, causing it to release the polymerase body and move (with CstF) to the CTD.


Assuntos
Fator de Especificidade de Clivagem e Poliadenilação/metabolismo , RNA Polimerases Dirigidas por DNA/antagonistas & inibidores , Poli A/metabolismo , Sinais de Poliadenilação na Ponta 3' do RNA , Transcrição Gênica , Animais , Linhagem Celular , Fator de Especificidade de Clivagem e Poliadenilação/antagonistas & inibidores , Fator Estimulador de Clivagem/metabolismo , RNA Polimerases Dirigidas por DNA/genética , RNA Polimerases Dirigidas por DNA/metabolismo , Humanos , RNA Mensageiro/química , RNA Mensageiro/metabolismo , Deleção de Sequência
12.
Proc Natl Acad Sci U S A ; 103(41): 14994-5001, 2006 Oct 10.
Artigo em Inglês | MEDLINE | ID: mdl-17008405

RESUMO

Many eukaryotic cells use RNA-directed silencing mechanisms to protect against viruses and transposons and to suppress endogenous gene expression at the posttranscriptional level. RNA silencing also is implicated in epigenetic mechanisms affecting chromosome structure and transcriptional gene silencing. Here, we describe enhanced silencing phenotype (esp) mutants in Arabidopsis thaliana that reveal how proteins associated with RNA processing and 3' end formation can influence RNA silencing. These proteins were a putative DEAH RNA helicase homologue of the yeast PRP2 RNA splicing cofactor and homologues of mRNA 3' end formation proteins CstF64, symplekin/PTA1, and CPSF100. The last two proteins physically associated with the flowering time regulator FY in the 3' end formation complex AtCPSF. The phenotypes of the 3' end formation esp mutants include impaired termination of the transgene transcripts, early flowering, and enhanced silencing of the FCA-beta mRNA. Based on these findings, we propose that the ESP-containing 3' end formation complexes prevent transgene and endogenous mRNAs from entering RNA-silencing pathways. According to this proposal, in the absence of these ESP proteins, these RNAs have aberrant 3' termini. The aberrant RNAs would enter the RNA silencing pathways because they are converted into dsRNA by RNA-dependent RNA polymerases.


Assuntos
Arabidopsis/fisiologia , Flores/genética , Interferência de RNA , Processamento Pós-Transcricional do RNA/genética , RNA de Plantas/antagonistas & inibidores , RNA de Plantas/genética , Sequência de Aminoácidos , Arabidopsis/genética , Proteínas de Arabidopsis , Proteínas de Ciclo Celular/antagonistas & inibidores , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Fator de Especificidade de Clivagem e Poliadenilação/antagonistas & inibidores , Fator de Especificidade de Clivagem e Poliadenilação/genética , Fator Estimulador de Clivagem/antagonistas & inibidores , Fator Estimulador de Clivagem/genética , Endopeptidases/genética , Endopeptidases/metabolismo , Flores/fisiologia , Dados de Sequência Molecular , Mutação , RNA de Plantas/metabolismo , Separase , Homologia de Sequência de Aminoácidos
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