RESUMEN

In eukaryotes, RNAs transcribed by RNA Pol II are modified at the 5' end with a 7-methylguanosine (m7G) cap, which is recognized by the nuclear cap binding complex (CBC). The CBC plays multiple important roles in mRNA metabolism, including transcription, splicing, polyadenylation, and export. It promotes mRNA export through direct interaction with a key mRNA export factor, ALYREF, which in turn links the TRanscription and EXport (TREX) complex to the 5' end of mRNA. However, the molecular mechanism for CBC-mediated recruitment of the mRNA export machinery is not well understood. Here, we present the first structure of the CBC in complex with an mRNA export factor, ALYREF. The cryo-EM structure of CBC-ALYREF reveals that the RRM domain of ALYREF makes direct contact with both the NCBP1 and NCBP2 subunits of the CBC. Comparing CBC-ALYREF with other cellular complexes containing CBC and/or ALYREF components provides insights into the coordinated events during mRNA transcription, splicing, and export.

Asunto(s)

Microscopía por Crioelectrón , Complejo Proteico Nuclear de Unión a la Caperuza/metabolismo , Complejo Proteico Nuclear de Unión a la Caperuza/química , Humanos , ARN Mensajero/metabolismo , ARN Mensajero/química , ARN Mensajero/genética , Conformación Proteica , Unión Proteica

RESUMEN

Mutations in SF3B1 are common in many types of cancer, which promotes cancer progression through aberrant RNA splicing. Recently, mRNA nuclear export has been reported to be defective in cells with SF3B1 K700E mutation. However, the mechanism remains unclear. Our study reveals that the K700E mutation in SF3B1 attenuates its interaction with THOC5, an essential component of mRNA nuclear export complex THO. Furthermore, SF3B1 mutation caused reduced binding of THOC5 with some mRNA and inhibited the nuclear export of these mRNA. Interestingly, THOC5 overexpression restores the nuclear export of these mRNA in cells with SF3B1 K700E mutation. Importantly, other types of cancer-associated SF3B1 mutations also inhibited mRNA nuclear export similarly, suggesting that it is common for cancer-associated SF3B1 mutation to inhibit mRNA nuclear export. Our research highlights the critical role of the THOC5-SF3B1 interaction in the regulation of mRNA nuclear export and provides valuable insights into the impact of SF3B1 mutations on mRNA nuclear export.

RESUMEN

Chromosome Region Maintenance 1 (CRM1), also known as Exportin 1 (XPO1), is a protein that is critical for transport of proteins and RNA to the cytoplasm through the nuclear pore complex. CRM1 inhibition with small molecule inhibitors is currently being studied in many cancers, including leukemias, solid organ malignancies and brain tumors. We review the structure of CRM1, its role in nuclear export, the current availability of CRM1 inhibitors, and the role of CRM1 in a number of distinct cellular processes. A deeper understanding of how CRM1 functions in nuclear export as well as other cellular processes may allow for the development of additional novel CRM1 inhibitors.

RESUMEN

Nuclear export of the viral ribonucleoprotein (vRNP) is a critical step in the influenza A virus (IAV) life cycle and may be an effective target for the development of anti-IAV drugs. The host factor ras-related nuclear protein (RAN) is known to participate in the life cycle of several viruses, but its role in influenza virus replication remains unknown. In the present study, we aimed to determine the function of RAN in influenza virus replication using different cell lines and subtype strains. We found that RAN is essential for the nuclear export of vRNP, as it enhances the binding affinity of XPO1 toward the viral nuclear export protein NS2. Depletion of RAN constrained the vRNP complex in the nucleus and attenuated the replication of various subtypes of influenza virus. Using in silico compound screening, we identified that bepotastine could dissociate the RAN-XPO1-vRNP trimeric complex and exhibit potent antiviral activity against influenza virus both in vitro and in vivo. This study demonstrates the important role of RAN in IAV replication and suggests its potential use as an antiviral target.

Asunto(s)

Transporte Activo de Núcleo Celular , Antivirales , Proteína Exportina 1 , Virus de la Influenza A , Carioferinas , Replicación Viral , Proteína de Unión al GTP ran , Replicación Viral/efectos de los fármacos , Humanos , Proteína de Unión al GTP ran/metabolismo , Proteína de Unión al GTP ran/genética , Antivirales/farmacología , Animales , Virus de la Influenza A/efectos de los fármacos , Virus de la Influenza A/fisiología , Carioferinas/metabolismo , Carioferinas/antagonistas & inhibidores , Perros , Receptores Citoplasmáticos y Nucleares/metabolismo , Receptores Citoplasmáticos y Nucleares/genética , Células de Riñón Canino Madin Darby , Proteínas no Estructurales Virales/metabolismo , Proteínas no Estructurales Virales/genética , Ratones , Piperidinas/farmacología , Gripe Humana/virología , Células A549 , Nucleoproteínas/metabolismo , Nucleoproteínas/genética , Células HEK293 , Línea Celular , Núcleo Celular/metabolismo , Ribonucleoproteínas/metabolismo , Ribonucleoproteínas/genética

RESUMEN

MITF, a basic Helix-Loop-Helix Zipper (bHLHZip) transcription factor, plays vital roles in melanocyte development and functions as an oncogene. We perform a genetic screen for suppressors of the Mitf-associated pigmentation phenotype in mice and identify an intragenic Mitf mutation that terminates MITF at the K316 SUMOylation site, leading to loss of the C-end intrinsically disordered region (IDR). The resulting protein is more nuclear but less stable than wild-type MITF and retains DNA-binding ability. As a dimer, it can translocate wild-type and mutant MITF partners into the nucleus, improving its own stability thus ensuring nuclear MITF supply. smFRET analysis shows interactions between K316 SUMOylation and S409 phosphorylation sites across monomers; these interactions largely explain the observed effects. The recurrent melanoma-associated E318K mutation in MITF, which affects K316 SUMOylation, also alters protein regulation in concert with S409. This suggests that residues K316 and S409 of MITF are impacted by SUMOylation and phosphorylation, respectively, mediating effects on nuclear localization and stability through conformational changes. Our work provides a novel mechanism of genetic suppression, and an example of how apparently deleterious mutations lead to normal phenotypes.

RESUMEN

BACKGROUND: DEAD-box RNA helicase 19 A (DDX19A) is overexpressed in cervical squamous cell carcinoma. However, its role in gastric cancer remains unclear. The present study aimed to explore the role and underlying mechanism of DDX19A in the development of gastric cancer. METHODS: The expression of DDX19A in gastric cancer and paracancerous tissues was evaluated through quantitative polymerase chain reaction, western blotting, and immunohistochemical staining. The biological functions of DDX19A in gastric cancer were determined using CCK8, plate colony-forming, and Transwell migration assays. The specific mechanism of DDX19A in gastric cancer cells was studied using western blotting, RNA-binding protein immunoprecipitation, mRNA half-life detection, and nuclear and cytoplasmic RNA isolation. RESULTS: DDX19A was highly expressed in gastric cancer and positively associated with malignant clinicopathological features and poor prognosis. Additionally, DDX19A promoted gastric cancer cell proliferation, migration, and epithelial-mesenchymal transition phenotypes. Mechanistically, DDX19A activated the PI3K/AKT pathway by upregulating phosphatidylinositol-3-kinase (PIK3CA) expression. Furthermore, DDX19A interacted with PIK3CA mRNA, stabilized it, and facilitated its export from the nucleus. CONCLUSIONS: Our study reveals a novel mechanism whereby DDX19A promotes the proliferation and migration of gastric cancer cells by enhancing the stability and nuclear export of PIK3CA mRNA, thereby activating the PI3K/AKT pathway.

RESUMEN

BACKGROUND: The role of selinexor, a targeted inhibitor of exportin 1 (XPO1), in the treatment of cholangiocarcinoma is not yet fully understood. This study conducted comprehensive in vitro and in vivo investigations to elucidate the effects of selinexor on cholangiocarcinoma, with a focus on its mechanistic relationship with the cellular localization of Paternally Expressed Gene 3 (PEG3). METHODS: A patient-derived xenograft (PDX) model was established using samples from a cholangiocarcinoma patient in immunodeficient mice to assess the in vivo effects of selinexor. Additionally, cholangiocarcinoma cell lines HuCC-T1 and BRE were cultured to evaluate selinexor's impact on cell proliferation, invasion, migration, cell cycle, and apoptosis. HuCC-T1 cells were also implanted in immunodeficient mice for further investigation. Immunofluorescence and Western blotting were employed to observe the expression and localization of the PEG3 protein. RESULTS: The results demonstrated that selinexor significantly inhibited tumor growth in the cholangiocarcinoma PDX model and promoted the accumulation of PEG3 protein within the nuclei of tumor cells. In vitro experiments showed that selinexor effectively suppressed cholangiocarcinoma cell proliferation, invasion, and migration, while also impeding the cell cycle and inducing apoptosis. Notably, selinexor markedly facilitated the nuclear accumulation of PEG3 protein in cholangiocarcinoma cells. However, when PEG3 expression was knocked down, the effects of selinexor on cholangiocarcinoma were significantly reversed. CONCLUSION: These findings suggest that selinexor inhibits the progression of cholangiocarcinoma by targeting XPO1 and promoting the nuclear accumulation of PEG3 protein, thereby hindering the cell cycle and inducing apoptosis.

RESUMEN

Dissecting the regulatory mechanisms controlling mammalian transcripts from production to degradation requires quantitative measurements of mRNA flow across the cell. We developed subcellular TimeLapse-seq to measure the rates at which RNAs are released from chromatin, exported from the nucleus, loaded onto polysomes, and degraded within the nucleus and cytoplasm in human and mouse cells. These rates varied substantially, yet transcripts from genes with related functions or targeted by the same transcription factors and RNA-binding proteins flowed across subcellular compartments with similar kinetics. Verifying these associations uncovered a link between DDX3X and nuclear export. For hundreds of RNA metabolism genes, most transcripts with retained introns were degraded by the nuclear exosome, while the remaining molecules were exported with stable cytoplasmic lifespans. Transcripts residing on chromatin for longer had extended poly(A) tails, whereas the reverse was observed for cytoplasmic mRNAs. Finally, machine learning identified molecular features that predicted the diverse life cycles of mRNAs.

Asunto(s)

Núcleo Celular , Cromatina , ARN Helicasas DEAD-box , ARN Mensajero , Animales , Humanos , Ratones , ARN Mensajero/metabolismo , ARN Mensajero/genética , Núcleo Celular/metabolismo , Núcleo Celular/genética , ARN Helicasas DEAD-box/metabolismo , ARN Helicasas DEAD-box/genética , Cromatina/metabolismo , Cromatina/genética , Citoplasma/metabolismo , Citoplasma/genética , Estabilidad del ARN , Transporte Activo de Núcleo Celular , Polirribosomas/metabolismo , Polirribosomas/genética , Aprendizaje Automático , Proteínas de Unión al ARN/metabolismo , Proteínas de Unión al ARN/genética , Exosomas/metabolismo , Exosomas/genética

RESUMEN

Influenza A viruses (IAV) utilize host proteins throughout their life cycle to infect and replicate in their hosts. We previously showed that host adaptive mutations in avian IAV PA help recruit host protein G-Rich RNA Sequence Binding Factor 1 (GRSF1) to the nucleoprotein (NP) 5' untranslated region (UTR), leading to the enhanced nuclear export and translation of NP mRNA. In this study, we evaluated the impact of GRSF1 in the viral life cycle. We rescued and characterized a 2009 pH1N1 virus with a mutated GRSF1 binding site in the 5' UTR of NP mRNA. Mutant viral growth was attenuated relative to pH1N1 wild-type (WT) in mammalian cells. We observed a specific reduction in the NP protein production and cytosolic accumulation of NP mRNAs, indicating a critical role of GRSF1 in the nuclear export of IAV NP mRNAs. Further, in vitro-transcribed mutated NP mRNA was translated less efficiently than WT NP mRNA in transfected cells. Together, these findings show that GRSF1 binding is important for both mRNA nuclear export and translation and affects overall IAV growth. Enhanced association of GRSF1 to NP mRNA by PA mutations leads to rapid virus growth, which could be a key process of mammalian host adaptation of IAV.

Asunto(s)

Transporte Activo de Núcleo Celular , Biosíntesis de Proteínas , ARN Mensajero , ARN Viral , Humanos , ARN Mensajero/genética , ARN Mensajero/metabolismo , ARN Viral/genética , ARN Viral/metabolismo , Animales , Virus de la Influenza A/genética , Virus de la Influenza A/fisiología , Virus de la Influenza A/metabolismo , Replicación Viral , Subtipo H1N1 del Virus de la Influenza A/genética , Subtipo H1N1 del Virus de la Influenza A/metabolismo , Subtipo H1N1 del Virus de la Influenza A/fisiología , Núcleo Celular/metabolismo , Núcleo Celular/virología , Regiones no Traducidas 5'/genética , Proteínas de la Nucleocápside/metabolismo , Proteínas de la Nucleocápside/genética , Células de Riñón Canino Madin Darby , Células HEK293 , Proteínas de Unión al ARN/metabolismo , Proteínas de Unión al ARN/genética , Perros , Gripe Humana/virología , Gripe Humana/metabolismo , Gripe Humana/genética , Mutación , Interacciones Huésped-Patógeno/genética , Proteínas del Núcleo Viral/metabolismo , Proteínas del Núcleo Viral/genética

RESUMEN

Hepatocellular carcinoma (HCC) is one of the most prevalent and deadly tumors; however, its pathogenic mechanism remains largely elusive. In-depth researches are needed to reveal the expression regulatory mechanisms and functions of the RNA-binding protein RALY in HCC. Here, we identify RALY as a highly expressed oncogenic factor that affects HCC cells proliferation both in vitro and in vivo. O-GlcNAcylation of RALY at Ser176 enhances its stability by protecting RALY from TRIM27-mediated ubiquitination, thus maintaining hyper-expression of the RALY protein. Mechanistically, RALY interacts with USP22 messenger RNA, as revealed by RNA immunoprecipitation, to increase their cytoplasmic localization and protein expression, thereby promoting the proliferation of HCC cells. Furthermore, we develop a novel RALY protein degrader based on peptide proteolysis-targeting chimeras, named RALY-PROTAC, which we chemically synthesize by linking a RALY-targeting peptide with the E3 ubiquitin ligase recruitment ligand pomalidomide. In conclusion, our findings demonstrate a novel mechanism by which O-GlcNAcylation/RALY/USP22 mRNA axis aggravates HCC cells proliferation. RALY-PROTACs as degraders of the RALY protein exhibit potential as therapeutic drugs for RALY-overexpressing HCC.

Asunto(s)

Carcinoma Hepatocelular , Proliferación Celular , Neoplasias Hepáticas , Ubiquitina Tiolesterasa , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patología , Carcinoma Hepatocelular/genética , Humanos , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patología , Neoplasias Hepáticas/genética , Ubiquitina Tiolesterasa/metabolismo , Ubiquitina Tiolesterasa/genética , Línea Celular Tumoral , Animales , ARN Mensajero/metabolismo , ARN Mensajero/genética , Ratones , Ratones Desnudos , Ubiquitinación , Transporte Activo de Núcleo Celular

RESUMEN

The model of RNA stability has undergone a transformative shift with the revelation of a cytoplasmic capping activity that means a subset of transcripts are recapped autonomously of their nuclear counterparts. The present study demonstrates nucleo-cytoplasmic shuttling of the mRNA-capping enzyme (CE, also known as RNA guanylyltransferase and 5'-phosphatase; RNGTT), traditionally acknowledged for its nuclear localization and functions, elucidating its contribution to cytoplasmic capping activities. A unique nuclear export sequence in CE mediates XPO1-dependent nuclear export of CE. Notably, during sodium arsenite-induced oxidative stress, cytoplasmic CE (cCE) congregates within stress granules (SGs). Through an integrated approach involving molecular docking and subsequent co-immunoprecipitation, we identify eIF3b, a constituent of SGs, as an interactive associate of CE, implying that it has a potential role in guiding cCE to SGs. We measured the cap status of specific mRNA transcripts from U2OS cells that were non-stressed, stressed and recovered from stress, which indicated that cCE-target transcripts lost their caps during stress but remarkably regained cap stability during the recovery phase. This comprehensive study thus uncovers a novel facet of cytoplasmic CE, which facilitates cellular recovery from stress by maintaining cap homeostasis of target mRNAs.

Asunto(s)

Citoplasma , Homeostasis , ARN Mensajero , Gránulos de Estrés , Humanos , ARN Mensajero/metabolismo , ARN Mensajero/genética , Gránulos de Estrés/metabolismo , Citoplasma/metabolismo , Caperuzas de ARN/metabolismo , Arsenitos/farmacología , Estrés Oxidativo , Transporte Activo de Núcleo Celular , ARN Nucleotidiltransferasas/metabolismo , ARN Nucleotidiltransferasas/genética , Compuestos de Sodio/farmacología , Proteína Exportina 1 , Carioferinas/metabolismo , Carioferinas/genética , Receptores Citoplasmáticos y Nucleares/metabolismo , Receptores Citoplasmáticos y Nucleares/genética , Gránulos Citoplasmáticos/metabolismo , Estabilidad del ARN , Núcleo Celular/metabolismo , Línea Celular Tumoral , Nucleotidiltransferasas

RESUMEN

Promyelocytic leukemia protein (PML), a tumor suppressor protein, plays a key role in cell cycle regulation, apoptosis, senescence and cellular metabolism. Here, we report that PML promotes apoptosis and ferroptosis. Our data showed that PML over-expression inhibited cell proliferation and migration. PML over-expression increased apoptotic cells, nuclear condensation and the loss of mitochondrial membrane potential, accompanied by regulation of Bcl-2 family proteins and reactive oxygen species (ROS) level, suggesting that PML enhanced apoptosis. Meanwhile, PML over-expression not only increased lipid ROS accumulation and Malondialdehyde (MDA) content but also downregulated solute carrier family 7 member 11 (SLC7A11) and glutathione peroxidase 4 (GPX4) expression, indicating that PML enhanced ferroptosis. Additionally, knockdown of p53 attenuated the effect of PML on SLC7A11 and GPX4, and inhibited the increase of lipid ROS and ROS by PML over-expression. Moreover, translocation of PML from nucleus to cytoplasm not only promoted apoptosis and ferroptosis, but also inhibited cell proliferation. Taken together, PML promotes apoptosis and ferroptosis, in which the mediation of p53 and the nuclear export of PML play important roles.

Asunto(s)

Transporte Activo de Núcleo Celular , Sistema de Transporte de Aminoácidos y+ , Apoptosis , Proliferación Celular , Ferroptosis , Fosfolípido Hidroperóxido Glutatión Peroxidasa , Proteína de la Leucemia Promielocítica , Proteína p53 Supresora de Tumor , Humanos , Sistema de Transporte de Aminoácidos y+/metabolismo , Sistema de Transporte de Aminoácidos y+/genética , Línea Celular Tumoral , Movimiento Celular , Núcleo Celular/metabolismo , Potencial de la Membrana Mitocondrial , Fosfolípido Hidroperóxido Glutatión Peroxidasa/metabolismo , Proteína de la Leucemia Promielocítica/metabolismo , Proteína de la Leucemia Promielocítica/genética , Especies Reactivas de Oxígeno/metabolismo , Proteína p53 Supresora de Tumor/metabolismo

RESUMEN

Selinexor, a selective inhibitor of nuclear export (SINE), is gaining recognition beyond oncology for its potential in anti-inflammatory therapy. This review elucidates Selinexor's dual action, highlighting its anti-tumor efficacy in various cancers including hematologic malignancies and solid tumors, and its promising anti-inflammatory effects. In cancer treatment, Selinexor has demonstrated benefits as monotherapy and in combination with other therapeutics, particularly in drug-resistant cases. Its role in enhancing the effectiveness of bone marrow transplants has also been noted. Importantly, the drug's impact on key inflammatory pathways provides a new avenue for the management of conditions like sepsis, viral infections including COVID-19, and chronic inflammatory diseases such as Duchenne Muscular Dystrophy and Parkinson's Disease. The review emphasizes the criticality of managing Selinexor's side effects through diligent dose optimization and patient monitoring. Given the complexities of its broader applications, extensive research is called upon to validate Selinexor's long-term safety and effectiveness, with a keen focus on its integration into clinical practice for a diverse spectrum of disorders.

Asunto(s)

Hidrazinas , Triazoles , Humanos , Hidrazinas/uso terapéutico , Triazoles/uso terapéutico , Triazoles/farmacología , Antiinflamatorios/uso terapéutico , Neoplasias/tratamiento farmacológico , Neoplasias/terapia , Neoplasias/inmunología , COVID-19/inmunología , SARS-CoV-2 , Antineoplásicos/uso terapéutico

RESUMEN

We report the development of a 384-well formatted NanoBRET assay to characterize molecular glues of 14-3-3/client interactions in living cells. The seven isoforms of 14-3-3 are dimeric hub proteins with diverse roles including transcription factor regulation and signal transduction. 14-3-3 interacts with hundreds of client proteins to regulate their function and is therefore an ideal therapeutic target when client selectivity can be achieved. We have developed the NanoBRET system for three 14-3-3σ client proteins CRAF, TAZ, and estrogen receptor α (ERα), which represent three specific binding modes. We have measured stabilization of 14-3-3σ/client complexes by molecular glues with EC50 values between 100 nM and 1 µM in cells, which align with the EC50 values calculated by fluorescence anisotropy in vitro. Developing this NanoBRET system for the hub protein 14-3-3σ allows for a streamlined approach, bypassing multiple optimization steps in the assay development process for other 14-3-3σ clients. The NanoBRET system allows for an assessment of PPI stabilization in a more physiologically relevant, cell-based environment using full-length proteins. The method is applicable to diverse protein-protein interactions (PPIs) and offers a robust platform to explore libraries of compounds for both PPI stabilizers and inhibitors.

Asunto(s)

Proteínas 14-3-3 , Unión Proteica , Proteínas 14-3-3/metabolismo , Proteínas 14-3-3/genética , Humanos , Receptor alfa de Estrógeno/metabolismo , Receptor alfa de Estrógeno/genética , Exorribonucleasas/metabolismo , Exorribonucleasas/genética

RESUMEN

mRNAs continually change their protein partners throughout their lifetimes, yet our understanding of mRNA-protein complex (mRNP) remodeling is limited by a lack of temporal data. Here, we present time-resolved mRNA interactome data by performing pulse metabolic labeling with photoactivatable ribonucleoside in human cells, UVA crosslinking, poly(A)+ RNA isolation, and mass spectrometry. This longitudinal approach allowed the quantification of over 700 RNA binding proteins (RBPs) across ten time points. Overall, the sequential order of mRNA binding aligns well with known functions, subcellular locations, and molecular interactions. However, we also observed RBPs with unexpected dynamics: the transcription-export (TREX) complex recruited posttranscriptionally after nuclear export factor 1 (NXF1) binding, challenging the current view of transcription-coupled mRNA export, and stress granule proteins prevalent in aged mRNPs, indicating roles in late stages of the mRNA life cycle. To systematically identify mRBPs with unknown functions, we employed machine learning to compare mRNA binding dynamics with Gene Ontology (GO) annotations. Our data can be explored at chronology.rna.snu.ac.kr.

Asunto(s)

ARN Mensajero , Proteínas de Unión al ARN , Humanos , ARN Mensajero/metabolismo , ARN Mensajero/genética , Proteínas de Unión al ARN/metabolismo , Proteínas de Unión al ARN/genética , Ribonucleoproteínas/metabolismo , Ribonucleoproteínas/genética , Unión Proteica , Proteínas de Transporte Nucleocitoplasmático/metabolismo , Proteínas de Transporte Nucleocitoplasmático/genética , Células HeLa , Factores de Tiempo , Aprendizaje Automático

RESUMEN

Autographa californica multiple nucleopolyhedrovirus (AcMNPV) Ac93 is highly conserved in all sequenced baculovirus genomes, and it plays important roles in both the nuclear egress of nucleocapsids and the formation of intranuclear microvesicles. In this study, we characterized a cellular CRM1-dependent nuclear export signal (NES) of AcMNPV Ac93. Bioinformatic analysis revealed that AcMNPV Ac93 may contain an NES at amino acids 115-125. Green fluorescent protein (GFP) fused to the NES (GFP:NES) of AcMNPV Ac93 is localized to the cytoplasm of transfected cells. Multiple point mutation analysis demonstrated that NES is important for the nuclear export of GFP:NES. Bimolecular fluorescence complementation experiments and co-immunoprecipitation assays confirmed that Ac93 interacts with Spodoptera frugiperda CRM1 (SfCRM1). However, AcMNPV Ac34 inhibits cellular CRM1-dependent nuclear export of GFP:NES. To determine whether the NES in AcMNPV Ac93 is important for the formation of intranuclear microvesicles, an ac93-null AcMNPV bacmid was constructed; the wild-type and NES-mutated Ac93 were reinserted into the ac93-null AcMNPV bacmid. Immunofluorescence analysis showed that Ac93 and SfCRM1 were predominantly colocalized at intranuclear microvesicles in infected cells, while the construct containing point mutations at residues 123 and 125 of Ac93 resulted in a defect in budded virus production and the abolishment of intranuclear microvesicles. Together, these data demonstrate that Ac93 contains a functional NES, which is required for the production of progeny viruses and the formation of intranuclear microvesicles.IMPORTANCEAutographa californica multiple nucleopolyhedrovirus (AcMNPV) Ac93 is important for the formation of intranuclear microvesicles. However, how the baculovirus manipulates Ac93 for the formation of intranuclear microvesicles is unclear. In this study, we identified a nuclear export signal (NES) at amino acids 115-125 of AcMNPV Ac93. Our results showed that the NES is required for the interaction between Ac93 and Spodoptera frugiperda CRM1 (SfCRM1). However, AcMNPV Ac34 inhibits the nuclear export of green fluorescent protein fused to the NES. Our analysis revealed that Ac93 and SfCRM1 were predominantly colocalized at intranuclear microvesicles in AcMNPV-infected cells. Together, our results indicate that Ac93 participates in the formation of intranuclear microvesicles via the Ac93 NES-mediated CRM1 pathway.

Asunto(s)

Transporte Activo de Núcleo Celular , Señales de Exportación Nuclear , Nucleopoliedrovirus , Proteínas Virales , Animales , Núcleo Celular/metabolismo , Núcleo Celular/virología , Proteína Exportina 1 , Proteínas Fluorescentes Verdes/metabolismo , Proteínas Fluorescentes Verdes/genética , Carioferinas/metabolismo , Nucleopoliedrovirus/metabolismo , Nucleopoliedrovirus/fisiología , Nucleopoliedrovirus/genética , Receptores Citoplasmáticos y Nucleares/metabolismo , Receptores Citoplasmáticos y Nucleares/genética , Células Sf9 , Spodoptera/virología , Proteínas Virales/química , Proteínas Virales/genética , Proteínas Virales/metabolismo

RESUMEN

Cancer metastasis is the major cause of death in patients with breast cancer (BC). The liver is a common site of breast cancer metastasis, and the 5-year survival rate of patients with breast cancer liver metastases (BCLMs) is only about 8.5 %. CircRNAs are involved in a variety of cancer-related pathological behaviors, and their unique structure and resistance to RNA degradation enable them to serve as ideal diagnostic biomarkers and therapeutic targets. Therefore, it is important to investigate the role and molecular mechanism of circRNAs in cancer metastasis. CircLIFR-007 was identified as a critical circular RNA in BC metastasis by circRNAs microarray and qRT-PCR experiment. Cell function assays were performed to explore the effect of circLIFR-007 in breast cancer cells. Experiments in vivo validated the function of circLIFR-007. Several molecular assays were performed to investigate the underlying mechanisms. We found that circLIFR-007 acted as a negative controller in breast cancer liver metastasis. CircLIFR-007 upregulates the phosphorylation level of YAP by exporting hnRNPA1 to promote the combination between hnRNPA1 and YAP in the cytoplasm. Overexpression of circLIFR-007 suppressed the expression of liver metastasis-related proteins, SREBF1 and SNAI1, which were regulated by transcription factor YAP. Functionally, circLIFR-007 inhibits the proliferation and metastasis of breast cancer cells both in vivo and in vitro.

Asunto(s)

Neoplasias de la Mama , Ribonucleoproteína Nuclear Heterogénea A1 , Neoplasias Hepáticas , ARN Circular , Factores de Transcripción , Proteínas Señalizadoras YAP , Humanos , Neoplasias de la Mama/patología , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/genética , Neoplasias Hepáticas/secundario , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/genética , Femenino , Proteínas Señalizadoras YAP/metabolismo , Fosforilación , Animales , Ribonucleoproteína Nuclear Heterogénea A1/metabolismo , Ribonucleoproteína Nuclear Heterogénea A1/genética , ARN Circular/genética , ARN Circular/metabolismo , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Ratones , Línea Celular Tumoral , Regulación Neoplásica de la Expresión Génica , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Transporte Activo de Núcleo Celular , Ratones Desnudos , Proliferación Celular , Ratones Endogámicos BALB C , Células MCF-7

RESUMEN

Combination antiretroviral therapy (ART) suppresses viral replication to undetectable levels, reduces mortality and morbidity, and improves the quality of life of people living with HIV (PWH). However, ART cannot cure HIV infection because it is unable to eliminate latently infected cells. HIV latency may be regulated by different HIV transcription mechanisms, such as blocks to initiation, elongation, and post-transcriptional processes. Several latency-reversing (LRA) and -promoting agents (LPA) have been investigated in clinical trials aiming to eliminate or reduce the HIV reservoir. However, none of these trials has shown a conclusive impact on the HIV reservoir. Here, we review the cellular and viral factors that regulate HIV-1 transcription, the potential pharmacological targets and genetic and epigenetic editing techniques that have been or might be evaluated to disrupt HIV-1 latency, the role of miRNA in post-transcriptional regulation of HIV-1, and the differences between the mechanisms regulating HIV-1 and HIV-2 expression.

RESUMEN

Exportin 1 (XPO1) is the major karyopherin-ß nuclear receptor mediating the nuclear export of hundreds of proteins and some classes of RNA and regulates several critical processes in the cell, including cell-cycle progression, transcription and translation. Viruses have co-opted XPO1 to promote nucleocytoplasmic transport of viral proteins and RNA. Maize mosaic virus (MMV) is a plant-infecting rhabdovirus transmitted in a circulative propagative manner by the corn planthopper, Peregrinus maidis. MMV replicates in the nucleus of plant and insect hosts, and it remains unknown whether MMV co-opts P. maidis XPO1 (PmXPO1) to complete its life cycle. Because XPO1 plays multiple regulatory roles in cell functions and virus infection, we hypothesized that RNAi-mediated silencing of XPO1 would negatively affect MMV accumulation and insect physiology. Although PmXPO1 expression was not modulated during MMV infection, PmXPO1 knockdown negatively affected MMV accumulation in P. maidis at 12 and 15 days after microinjection. Likewise, PmXPO1 knockdown negatively affected P. maidis survival and reproduction. PmXPO1 exhibited tissue-specific expression patterns with higher expression in the ovaries compared with the guts of adult females. Survival rate was significantly lower for PmXPO1 knockdown females, compared with controls, but no effect was observed for males. PmXPO1 knockdown experiments revealed a role for PmXPO1 in ovary function and egg production. Oviposition and egg hatch on plants were dramatically reduced in females treated with dsRNA PmXPO1. These results suggest that PmXPO1 is a positive regulator of P. maidis reproduction and that it plays a proviral role in the insect vector supporting MMV infection.

Asunto(s)

Proteína Exportina 1 , Hemípteros , Insectos Vectores , Carioferinas , Ovario , Interferencia de ARN , Receptores Citoplasmáticos y Nucleares , Animales , Femenino , Hemípteros/virología , Hemípteros/genética , Hemípteros/crecimiento & desarrollo , Carioferinas/metabolismo , Carioferinas/genética , Ovario/virología , Ovario/metabolismo , Ovario/crecimiento & desarrollo , Receptores Citoplasmáticos y Nucleares/metabolismo , Receptores Citoplasmáticos y Nucleares/genética , Insectos Vectores/virología , Insectos Vectores/genética , Rhabdoviridae/fisiología , Proteínas de Insectos/metabolismo , Proteínas de Insectos/genética , Zea mays/virología , Zea mays/genética , Técnicas de Silenciamiento del Gen