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1.
Cell ; 175(5): 1393-1404.e11, 2018 11 15.
Artículo en Inglés | MEDLINE | ID: mdl-30454648

RESUMEN

Ribonuclease (RNase) P is a ubiquitous ribozyme that cleaves the 5' leader from precursor tRNAs. Here, we report cryo-electron microscopy structures of the human nuclear RNase P alone and in complex with tRNAVal. Human RNase P is a large ribonucleoprotein complex that contains 10 protein components and one catalytic RNA. The protein components form an interlocked clamp that stabilizes the RNA in a conformation optimal for substrate binding. Human RNase P recognizes the tRNA using a double-anchor mechanism through both protein-RNA and RNA-RNA interactions. Structural comparison of the apo and tRNA-bound human RNase P reveals that binding of tRNA induces a local conformational change in the catalytic center, transforming the ribozyme into an active state. Our results also provide an evolutionary model depicting how auxiliary RNA elements in bacterial RNase P, essential for substrate binding, and catalysis, were replaced by the much more complex and multifunctional protein components in higher organisms.


Asunto(s)
Microscopía por Crioelectrón , ARN de Transferencia/química , Ribonucleasa P/química , Sitios de Unión , Evolución Molecular , Células HEK293 , Holoenzimas/química , Humanos , Simulación de Dinámica Molecular , Conformación de Ácido Nucleico , Dominios Proteicos , Estructura Terciaria de Proteína , ARN de Transferencia/metabolismo , Ribonucleasa P/aislamiento & purificación , Ribonucleasa P/metabolismo
2.
Cell ; 172(1-2): 331-343.e13, 2018 01 11.
Artículo en Inglés | MEDLINE | ID: mdl-29290466

RESUMEN

Telomerase maintains chromosome ends from humans to yeasts. Recruitment of yeast telomerase to telomeres occurs through its Ku and Est1 subunits via independent interactions with telomerase RNA (TLC1) and telomeric proteins Sir4 and Cdc13, respectively. However, the structures of the molecules comprising these telomerase-recruiting pathways remain unknown. Here, we report crystal structures of the Ku heterodimer and Est1 complexed with their key binding partners. Two major findings are as follows: (1) Ku specifically binds to telomerase RNA in a distinct, yet related, manner to how it binds DNA; and (2) Est1 employs two separate pockets to bind distinct motifs of Cdc13. The N-terminal Cdc13-binding site of Est1 cooperates with the TLC1-Ku-Sir4 pathway for telomerase recruitment, whereas the C-terminal interface is dispensable for binding Est1 in vitro yet is nevertheless essential for telomere maintenance in vivo. Overall, our results integrate previous models and provide fundamentally valuable structural information regarding telomere biology.


Asunto(s)
Proteínas de Unión al ADN/química , Simulación del Acoplamiento Molecular , Proteínas de Saccharomyces cerevisiae/química , Telomerasa/química , Homeostasis del Telómero , Proteínas de Unión a Telómeros/química , Sitios de Unión , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Unión Proteica , ARN/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas Reguladoras de Información Silente de Saccharomyces cerevisiae/genética , Proteínas Reguladoras de Información Silente de Saccharomyces cerevisiae/metabolismo , Telomerasa/genética , Telomerasa/metabolismo , Proteínas de Unión a Telómeros/genética , Proteínas de Unión a Telómeros/metabolismo
3.
Mol Cell ; 82(21): 4116-4130.e6, 2022 11 03.
Artículo en Inglés | MEDLINE | ID: mdl-36283412

RESUMEN

Pyruvate carboxylase (PC) catalyzes the two-step carboxylation of pyruvate to produce oxaloacetate, playing a key role in the maintenance of metabolic homeostasis in cells. Given its involvement in multiple diseases, PC has been regarded as a potential therapeutic target for obesity, diabetes, and cancer. Albeit acetyl-CoA has been recognized as the allosteric regulator of PC for over 60 years, the underlying mechanism of how acetyl-CoA induces PC activation remains enigmatic. Herein, by using time-resolved cryo-electron microscopy, we have captured the snapshots of PC transitional states during its catalytic cycle. These structures and the biochemical studies reveal that acetyl-CoA stabilizes PC in a catalytically competent conformation, which triggers a cascade of events, including ATP hydrolysis and the long-distance communication between the two reactive centers. These findings provide an integrated picture for PC catalysis and unveil the unique allosteric mechanism of acetyl-CoA in an essential biochemical reaction in all kingdoms of life.


Asunto(s)
Acetil-CoA Carboxilasa , Piruvato Carboxilasa , Humanos , Piruvato Carboxilasa/genética , Piruvato Carboxilasa/metabolismo , Acetilcoenzima A/metabolismo , Regulación Alostérica , Microscopía por Crioelectrón , Conformación Molecular , Acetil-CoA Carboxilasa/metabolismo
4.
Mol Cell ; 82(20): 3810-3825.e8, 2022 10 20.
Artículo en Inglés | MEDLINE | ID: mdl-36108631

RESUMEN

Human mixed-lineage leukemia (MLL) family methyltransferases methylate histone H3 lysine 4 to different methylation states (me1/me2/me3) with distinct functional outputs, but the mechanism underlying the different product specificities of MLL proteins remains unclear. Here, we develop methodologies to quantitatively measure the methylation rate difference between mono-, di-, and tri-methylation steps and demonstrate that MLL proteins possess distinct product specificities in the context of the minimum MLL-RBBP5-ASH2L complex. Comparative structural analyses of MLL complexes by X-ray crystal structures, fluorine-19 nuclear magnetic resonance, and molecular dynamics simulations reveal that the dynamics of two conserved tyrosine residues at the "F/Y (phenylalanine/tyrosine) switch" positions fine-tune the product specificity. The variation in the intramolecular interaction between SET-N and SET-C affects the F/Y switch dynamics, thus determining the product specificities of MLL proteins. These results indicate a modified F/Y switch rule applicable for most SET domain methyltransferases and implicate the functional divergence of MLL proteins.


Asunto(s)
N-Metiltransferasa de Histona-Lisina , Leucemia , Humanos , N-Metiltransferasa de Histona-Lisina/metabolismo , Histonas/metabolismo , Metiltransferasas/genética , Metiltransferasas/metabolismo , Lisina/metabolismo , Flúor/metabolismo , Proteína de la Leucemia Mieloide-Linfoide/metabolismo , Tirosina , Fenilalanina
5.
Nature ; 603(7903): 942-948, 2022 03.
Artículo en Inglés | MEDLINE | ID: mdl-35322232

RESUMEN

Standard first-line chemotherapy results in disease progression and death within one year in most patients with human epidermal growth factor receptor 2 (HER2)-negative gastro-oesophageal adenocarcinoma1-4. Nivolumab plus chemotherapy demonstrated superior overall survival versus chemotherapy at 12-month follow-up in gastric, gastro-oesophageal junction or oesophageal adenocarcinoma in the randomized, global CheckMate 649 phase 3 trial5 (programmed death ligand-1 (PD-L1) combined positive score ≥5 and all randomized patients). On the basis of these results, nivolumab plus chemotherapy is now approved as a first-line treatment for these patients in many countries6. Nivolumab and the cytotoxic T-lymphocyte antigen-4 (CTLA-4) inhibitor ipilimumab have distinct but complementary mechanisms of action that contribute to the restoration of anti-tumour T-cell function and induction of de novo anti-tumour T-cell responses, respectively7-11. Treatment combining 1 mg kg-1 nivolumab with 3 mg kg-1 ipilimumab demonstrated clinically meaningful anti-tumour activity with a manageable safety profile in heavily pre-treated patients with advanced gastro-oesophageal cancer12. Here we report both long-term follow-up results comparing nivolumab plus chemotherapy versus chemotherapy alone and the first results comparing nivolumab plus ipilimumab versus chemotherapy alone from CheckMate 649. After the 24.0-month minimum follow-up, nivolumab plus chemotherapy continued to demonstrate improvement in overall survival versus chemotherapy alone in patients with PD-L1 combined positive score ≥5 (hazard ratio 0.70; 95% confidence interval 0.61, 0.81) and all randomized patients (hazard ratio 0.79; 95% confidence interval 0.71, 0.88). Overall survival in patients with PD-L1 combined positive score ≥ 5 for nivolumab plus ipilimumab versus chemotherapy alone did not meet the prespecified boundary for significance. No new safety signals were identified. Our results support the continued use of nivolumab plus chemotherapy as standard first-line treatment for advanced gastro-oesophageal adenocarcinoma.


Asunto(s)
Adenocarcinoma , Protocolos de Quimioterapia Combinada Antineoplásica , Neoplasias Esofágicas , Neoplasias Gástricas , Adenocarcinoma/tratamiento farmacológico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Antígeno B7-H1 , Neoplasias Esofágicas/tratamiento farmacológico , Unión Esofagogástrica , Estudios de Seguimiento , Humanos , Ipilimumab/efectos adversos , Ipilimumab/uso terapéutico , Nivolumab/efectos adversos , Nivolumab/uso terapéutico , Neoplasias Gástricas/tratamiento farmacológico
6.
Mol Cell ; 75(2): 238-251.e5, 2019 07 25.
Artículo en Inglés | MEDLINE | ID: mdl-31348879

RESUMEN

BRCT domains support myriad protein-protein interactions involved in genome maintenance. Although di-BRCT recognition of phospho-proteins is well known to support the genotoxic response, whether multi-BRCT domains can acquire distinct structures and functions is unclear. Here we present the tetra-BRCT structures from the conserved yeast protein Rtt107 in free and ligand-bound forms. The four BRCT repeats fold into a tetrahedral structure that recognizes unmodified ligands using a bi-partite mechanism, suggesting repeat origami enabling function acquisition. Functional studies show that Rtt107 binding of partner proteins of diverse activities promotes genome replication and stability in both distinct and concerted manners. A unified theme is that tetra- and di-BRCT domains of Rtt107 collaborate to recruit partner proteins to chromatin. Our work thus illustrates how a master regulator uses two types of BRCT domains to recognize distinct genome factors and direct them to chromatin for constitutive genome protection.


Asunto(s)
Inestabilidad Genómica/genética , Proteínas Nucleares/ultraestructura , Dominios y Motivos de Interacción de Proteínas/genética , Proteínas de Saccharomyces cerevisiae/ultraestructura , Saccharomyces cerevisiae/genética , Cromatina/genética , Daño del ADN/genética , Ligandos , Proteínas Nucleares/química , Proteínas Nucleares/genética , Fosforilación , Unión Proteica , Dominios Proteicos/genética , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/genética
7.
Development ; 150(20)2023 10 15.
Artículo en Inglés | MEDLINE | ID: mdl-36975381

RESUMEN

Methionine is important for intestinal development and homeostasis in various organisms. However, the underlying mechanisms are poorly understood. Here, we demonstrate that the methionine adenosyltransferase gene Mat2a is essential for intestinal development and that the metabolite S-adenosyl-L-methionine (SAM) plays an important role in intestinal homeostasis. Intestinal epithelial cell (IEC)-specific knockout of Mat2a exhibits impaired intestinal development and neonatal lethality. Mat2a deletion in the adult intestine reduces cell proliferation and triggers IEC apoptosis, leading to severe intestinal epithelial atrophy and intestinal inflammation. Mechanistically, we reveal that SAM maintains the integrity of differentiated epithelium and protects IECs from apoptosis by suppressing the expression of caspases 3 and 8 and their activation. SAM supplementation improves the defective intestinal epithelium and reduces inflammatory infiltration sequentially. In conclusion, our study demonstrates that methionine metabolism and its intermediate metabolite SAM play essential roles in intestinal development and homeostasis in mice.


Asunto(s)
Metionina Adenosiltransferasa , S-Adenosilmetionina , Ratones , Animales , S-Adenosilmetionina/metabolismo , Metionina Adenosiltransferasa/genética , Metionina Adenosiltransferasa/metabolismo , Mucosa Intestinal/metabolismo , Metionina , Suplementos Dietéticos
8.
Nature ; 579(7797): 152-157, 2020 03.
Artículo en Inglés | MEDLINE | ID: mdl-32076264

RESUMEN

GPR52 is a class-A orphan G-protein-coupled receptor that is highly expressed in the brain and represents a promising therapeutic target for the treatment of Huntington's disease and several psychiatric disorders1,2. Pathological malfunction of GPR52 signalling occurs primarily through the heterotrimeric Gs protein2, but it is unclear how GPR52 and Gs couple for signal transduction and whether a native ligand or other activating input is required. Here we present the high-resolution structures of human GPR52 in three states: a ligand-free state, a Gs-coupled self-activation state and a potential allosteric ligand-bound state. Together, our structures reveal that extracellular loop 2 occupies the orthosteric binding pocket and operates as a built-in agonist, conferring an intrinsically high level of basal activity to GPR523. A fully active state is achieved when Gs is coupled to GPR52 in the absence of an external agonist. The receptor also features a side pocket for ligand binding. These insights into the structure and function of GPR52 could improve our understanding of other self-activated GPCRs, enable the identification of endogenous and tool ligands, and guide drug discovery efforts that target GPR52.


Asunto(s)
Receptores Acoplados a Proteínas G/química , Receptores Acoplados a Proteínas G/metabolismo , Regulación Alostérica , Sitio Alostérico , Secuencias de Aminoácidos , Secuencia de Aminoácidos , Apoproteínas/agonistas , Apoproteínas/química , Apoproteínas/metabolismo , Sitios de Unión , Microscopía por Crioelectrón , Cristalografía por Rayos X , Subunidades alfa de la Proteína de Unión al GTP Gs/química , Subunidades alfa de la Proteína de Unión al GTP Gs/metabolismo , Subunidades alfa de la Proteína de Unión al GTP Gs/ultraestructura , Humanos , Ligandos , Modelos Moleculares , Receptores Acoplados a Proteínas G/agonistas , Receptores Acoplados a Proteínas G/ultraestructura
9.
Nucleic Acids Res ; 52(2): 953-966, 2024 Jan 25.
Artículo en Inglés | MEDLINE | ID: mdl-38055835

RESUMEN

Nuclear respiratory factor 1 (NRF1) regulates the expression of genes that are vital for mitochondrial biogenesis, respiration, and various other cellular processes. While NRF1 has been reported to bind specifically to GC-rich promoters as a homodimer, the precise molecular mechanism governing its recognition of target gene promoters has remained elusive. To unravel the recognition mechanism, we have determined the crystal structure of the NRF1 homodimer bound to an ATGCGCATGCGCAT dsDNA. In this complex, NRF1 utilizes a flexible linker to connect its dimerization domain (DD) and DNA binding domain (DBD). This configuration allows one NRF1 monomer to adopt a U-turn conformation, facilitating the homodimer to specifically bind to the two TGCGC motifs in the GCGCATGCGC consensus sequence from opposite directions. Strikingly, while the NRF1 DBD alone could also bind to the half-site (TGCGC) DNA of the consensus sequence, the cooperativity between DD and DBD is essential for the binding of the intact GCGCATGCGC sequence and the transcriptional activity of NRF1. Taken together, our results elucidate the molecular mechanism by which NRF1 recognizes specific DNA sequences in the promoters to regulate gene expression.


Asunto(s)
ADN , Factor Nuclear 1 de Respiración , Humanos , Secuencia de Bases , ADN/metabolismo , Proteínas de Unión al ADN/genética , Factor Nuclear 1 de Respiración/genética , Factor Nuclear 1 de Respiración/metabolismo , Regiones Promotoras Genéticas
10.
Traffic ; 24(9): 384-396, 2023 09.
Artículo en Inglés | MEDLINE | ID: mdl-37341018

RESUMEN

Iron homeostasis is crucial for maintaining proper cellular function, and its disruption is considered one of the pathogenic mechanisms underlying musculoskeletal diseases. Under conditions of oxidative stress, the accumulation of cellular iron overload and lipid peroxidation can lead to ferroptosis. Extracellular vesicles (EVs), serving as mediators in the cell-to-cell communication, play an important role in regulating the outcome of cell ferroptosis. Growing evidence has proven that EV biogenesis and secretion are tightly associated with cellular iron export. Furthermore, different sources of EVs deliver diverse cargoes to bring about phenotypic changes in the recipient cells, either activating or inhibiting ferroptosis. Thus, delivering therapies targeting ferroptosis through EVs may hold significant potential for treating musculoskeletal diseases. This review aims to summarize current knowledge on the role of EVs in iron homeostasis and ferroptosis, as well as their therapeutic applications in musculoskeletal diseases, and thereby provide valuable insights for both research and clinical practice.


Asunto(s)
Vesículas Extracelulares , Ferroptosis , Enfermedades Musculoesqueléticas , Humanos , Hierro , Enfermedades Musculoesqueléticas/terapia , Homeostasis
11.
J Biol Chem ; 300(3): 105776, 2024 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-38382670

RESUMEN

The CCAAT/enhancer-binding proteins (C/EBPs) constitute a family of pivotal transcription factors involved in tissue development, cellular function, proliferation, and differentiation. NFIL3, as one of them, plays an important role in regulating immune cell differentiation, circadian clock system, and neural regeneration, yet its specific DNA recognition mechanism remains enigmatic. In this study, we showed by the ITC binding experiments that NFIL3 prefers to bind to the TTACGTAA DNA motif. Our structural studies revealed that the α-helical NFIL3 bZIP domain dimerizes through its leucine zipper region, and binds to DNA via its basic region. The two basic regions of the NFIL3 bZIP dimer were pushed apart upon binding to DNA, facilitating the snug accommodation of the two basic regions within the major grooves of the DNA. Remarkably, our binding and structural data also revealed that both NFIL3 and C/EBPα/ß demonstrate a shared preference for the TTACGTAA sequence. Furthermore, our study revealed that disease-associated mutations within the NFIL3 bZIP domain result in either reduction or complete disruption of its DNA binding ability. These discoveries not only provide valuable insights into the DNA binding mechanisms of NFIL3 but also elucidate the causal role of NFIL3 mutations in disease pathogenesis.


Asunto(s)
Factores de Transcripción con Cremalleras de Leucina de Carácter Básico , ADN , Secuencia de Aminoácidos , Secuencia de Bases , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/genética , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/metabolismo , Proteínas Potenciadoras de Unión a CCAAT/metabolismo , ADN/metabolismo , Factores de Transcripción/metabolismo , Humanos
12.
J Biol Chem ; 300(3): 105729, 2024 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-38336296

RESUMEN

RNase P and RNase mitochondrial RNA processing (MRP) are ribonucleoproteins (RNPs) that consist of a catalytic RNA and a varying number of protein cofactors. RNase P is responsible for precursor tRNA maturation in all three domains of life, while RNase MRP, exclusive to eukaryotes, primarily functions in rRNA biogenesis. While eukaryotic RNase P is associated with more protein cofactors and has an RNA subunit with fewer auxiliary structural elements compared to its bacterial cousin, the double-anchor precursor tRNA recognition mechanism has remarkably been preserved during evolution. RNase MRP shares evolutionary and structural similarities with RNase P, preserving the catalytic core within the RNA moiety inherited from their common ancestor. By incorporating new protein cofactors and RNA elements, RNase MRP has established itself as a distinct RNP capable of processing ssRNA substrates. The structural information on RNase P and MRP helps build an evolutionary trajectory, depicting how emerging protein cofactors harmonize with the evolution of RNA to shape different functions for RNase P and MRP. Here, we outline the structural and functional relationship between RNase P and MRP to illustrate the coevolution of RNA and protein cofactors, a key driver for the extant, diverse RNP world.


Asunto(s)
Endorribonucleasas , Evolución Molecular , Subunidades de Proteína , ARN Catalítico , Ribonucleasa P , Coenzimas , Endorribonucleasas/química , Endorribonucleasas/metabolismo , Subunidades de Proteína/química , Subunidades de Proteína/metabolismo , Ribonucleasa P/química , Ribonucleasa P/metabolismo , Procesamiento Postranscripcional del ARN , ARN Catalítico/genética , ARN Catalítico/metabolismo , ARN de Transferencia/genética , ARN de Transferencia/metabolismo , Especificidad por Sustrato , Eucariontes/enzimología
13.
N Engl J Med ; 386(5): 449-462, 2022 02 03.
Artículo en Inglés | MEDLINE | ID: mdl-35108470

RESUMEN

BACKGROUND: First-line chemotherapy for advanced esophageal squamous-cell carcinoma results in poor outcomes. The monoclonal antibody nivolumab has shown an overall survival benefit over chemotherapy in previously treated patients with advanced esophageal squamous-cell carcinoma. METHODS: In this open-label, phase 3 trial, we randomly assigned adults with previously untreated, unresectable advanced, recurrent, or metastatic esophageal squamous-cell carcinoma in a 1:1:1 ratio to receive nivolumab plus chemotherapy, nivolumab plus the monoclonal antibody ipilimumab, or chemotherapy. The primary end points were overall survival and progression-free survival, as determined by blinded independent central review. Hierarchical testing was performed first in patients with tumor-cell programmed death ligand 1 (PD-L1) expression of 1% or greater and then in the overall population (all randomly assigned patients). RESULTS: A total of 970 patients underwent randomization. At a 13-month minimum follow-up, overall survival was significantly longer with nivolumab plus chemotherapy than with chemotherapy alone, both among patients with tumor-cell PD-L1 expression of 1% or greater (median, 15.4 vs. 9.1 months; hazard ratio, 0.54; 99.5% confidence interval [CI], 0.37 to 0.80; P<0.001) and in the overall population (median, 13.2 vs. 10.7 months; hazard ratio, 0.74; 99.1% CI, 0.58 to 0.96; P = 0.002). Overall survival was also significantly longer with nivolumab plus ipilimumab than with chemotherapy among patients with tumor-cell PD-L1 expression of 1% or greater (median, 13.7 vs. 9.1 months; hazard ratio, 0.64; 98.6% CI, 0.46 to 0.90; P = 0.001) and in the overall population (median, 12.7 vs. 10.7 months; hazard ratio, 0.78; 98.2% CI, 0.62 to 0.98; P = 0.01). Among patients with tumor-cell PD-L1 expression of 1% or greater, a significant progression-free survival benefit was also seen with nivolumab plus chemotherapy over chemotherapy alone (hazard ratio for disease progression or death, 0.65; 98.5% CI, 0.46 to 0.92; P = 0.002) but not with nivolumab plus ipilimumab as compared with chemotherapy. The incidence of treatment-related adverse events of grade 3 or 4 was 47% with nivolumab plus chemotherapy, 32% with nivolumab plus ipilimumab, and 36% with chemotherapy alone. CONCLUSIONS: Both first-line treatment with nivolumab plus chemotherapy and first-line treatment with nivolumab plus ipilimumab resulted in significantly longer overall survival than chemotherapy alone in patients with advanced esophageal squamous-cell carcinoma, with no new safety signals identified. (Funded by Bristol Myers Squibb and Ono Pharmaceutical; CheckMate 648 ClinicalTrials.gov number, NCT03143153.).


Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Carcinoma de Células Escamosas/tratamiento farmacológico , Neoplasias Esofágicas/tratamiento farmacológico , Inhibidores de Puntos de Control Inmunológico/administración & dosificación , Ipilimumab/administración & dosificación , Nivolumab/administración & dosificación , Adulto , Anciano , Anciano de 80 o más Años , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Antígeno B7-H1/antagonistas & inhibidores , Carcinoma de Células Escamosas/mortalidad , Neoplasias Esofágicas/mortalidad , Femenino , Humanos , Inhibidores de Puntos de Control Inmunológico/efectos adversos , Ipilimumab/efectos adversos , Masculino , Persona de Mediana Edad , Nivolumab/efectos adversos , Supervivencia sin Progresión , Análisis de Supervivencia
14.
Plant Physiol ; 194(4): 2165-2182, 2024 Mar 29.
Artículo en Inglés | MEDLINE | ID: mdl-37995374

RESUMEN

N6-methyladenosine (m6A) in mRNA and 5-methylcytosine (5mC) in DNA have critical functions for regulating gene expression and modulating plant growth and development. However, the interplay between m6A and 5mC is an elusive territory and remains unclear mechanistically in plants. We reported an occurrence of crosstalk between m6A and 5mC in maize (Zea mays) via the interaction between mRNA adenosine methylase (ZmMTA), the core component of the m6A methyltransferase complex, and decrease in DNA methylation 1 (ZmDDM1), a key chromatin-remodeling factor that regulates DNA methylation. Genes with m6A modification were coordinated with a much higher level of DNA methylation than genes without m6A modification. Dysfunction of ZmMTA caused severe arrest during maize embryogenesis and endosperm development, leading to a significant decrease in CHH methylation in the 5' region of m6A-modified genes. Instead, loss of function of ZmDDM1 had no noteworthy effects on ZmMTA-related activity. This study establishes a direct link between m6A and 5mC during maize kernel development and provides insights into the interplay between RNA modification and DNA methylation.


Asunto(s)
Metilación de ADN , Zea mays , Metilación de ADN/genética , Zea mays/genética , Zea mays/metabolismo , Metilación de ARN , Metiltransferasas/genética , Metiltransferasas/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , ARN/metabolismo
15.
Nature ; 573(7774): 445-449, 2019 09.
Artículo en Inglés | MEDLINE | ID: mdl-31485071

RESUMEN

Methyltransferases of the mixed-lineage leukaemia (MLL) family-which include MLL1, MLL2, MLL3, MLL4, SET1A and SET1B-implement methylation of histone H3 on lysine 4 (H3K4), and have critical and distinct roles in the regulation of transcription in haematopoiesis, adipogenesis and development1-6. The C-terminal catalytic SET (Su(var.)3-9, enhancer of zeste and trithorax) domains of MLL proteins are associated with a common set of regulatory factors (WDR5, RBBP5, ASH2L and DPY30) to achieve specific activities7-9. Current knowledge of the regulation of MLL activity is limited to the catalysis of histone H3 peptides, and how H3K4 methyl marks are deposited on nucleosomes is poorly understood. H3K4 methylation is stimulated by mono-ubiquitination of histone H2B on lysine 120 (H2BK120ub1), a prevalent histone H2B mark that disrupts chromatin compaction and favours open chromatin structures, but the underlying mechanism remains unknown10-12. Here we report cryo-electron microscopy structures of human MLL1 and MLL3 catalytic modules associated with nucleosome core particles that contain H2BK120ub1 or unmodified H2BK120. These structures demonstrate that the MLL1 and MLL3 complexes both make extensive contacts with the histone-fold and DNA regions of the nucleosome; this allows ease of access to the histone H3 tail, which is essential for the efficient methylation of H3K4. The H2B-conjugated ubiquitin binds directly to RBBP5, orienting the association between MLL1 or MLL3 and the nucleosome. The MLL1 and MLL3 complexes display different structural organizations at the interface between the WDR5, RBBP5 and MLL1 (or the corresponding MLL3) subunits, which accounts for the opposite roles of WDR5 in regulating the activity of the two enzymes. These findings transform our understanding of the structural basis for the regulation of MLL activity at the nucleosome level, and highlight the pivotal role of nucleosome regulation in histone-tail modification.


Asunto(s)
Proteínas de Unión al ADN/química , N-Metiltransferasa de Histona-Lisina/metabolismo , Modelos Moleculares , Proteína de la Leucemia Mieloide-Linfoide/química , Nucleosomas/química , Nucleosomas/metabolismo , Microscopía por Crioelectrón , Proteínas de Unión al ADN/metabolismo , Activación Enzimática/genética , Regulación Enzimológica de la Expresión Génica/genética , N-Metiltransferasa de Histona-Lisina/química , Histonas/química , Histonas/metabolismo , Humanos , Metilación , Proteína de la Leucemia Mieloide-Linfoide/metabolismo , Estructura Cuaternaria de Proteína
16.
Mol Cell ; 65(5): 801-817.e4, 2017 Mar 02.
Artículo en Inglés | MEDLINE | ID: mdl-28216226

RESUMEN

Telomeres employ TRF2 to protect chromosome ends from activating the DNA damage sensor MRE11-RAD50-NBS1 (MRN), thereby repressing ATM-dependent DNA damage checkpoint responses. How TRF2 prevents MRN activation at dysfunctional telomeres is unclear. Here, we show that the phosphorylation status of NBS1 determines the repair pathway choice of dysfunctional telomeres. The crystal structure of the TRF2-NBS1 complex at 3.0 Å resolution shows that the NBS1 429YQLSP433 motif interacts specifically with the TRF2TRFH domain. Phosphorylation of NBS1 serine 432 by CDK2 in S/G2 dissociates NBS1 from TRF2, promoting TRF2-Apollo/SNM1B complex formation and the protection of leading-strand telomeres. Classical-NHEJ-mediated repair of telomeres lacking TRF2 requires phosphorylated NBS1S432 to activate ATM, while interaction of de-phosphorylated NBS1S432 with TRF2 promotes alternative-NHEJ repair of telomeres lacking POT1-TPP1. Our work advances understanding of how the TRF2TRFH domain orchestrates telomere end protection and reveals how the phosphorylation status of the NBS1S432 dictates repair pathway choice of dysfunctional telomeres.


Asunto(s)
Proteínas de Ciclo Celular/metabolismo , Roturas del ADN de Doble Cadena , Reparación del ADN por Unión de Extremidades , Proteínas Nucleares/metabolismo , Telómero/metabolismo , Proteína 2 de Unión a Repeticiones Teloméricas/metabolismo , Aminopeptidasas/genética , Aminopeptidasas/metabolismo , Proteínas de la Ataxia Telangiectasia Mutada/genética , Proteínas de la Ataxia Telangiectasia Mutada/metabolismo , Sitios de Unión , Proteínas de Ciclo Celular/química , Proteínas de Ciclo Celular/genética , Quinasa 2 Dependiente de la Ciclina/genética , Quinasa 2 Dependiente de la Ciclina/metabolismo , Enzimas Reparadoras del ADN/genética , Enzimas Reparadoras del ADN/metabolismo , Proteínas de Unión al ADN , Dipeptidil-Peptidasas y Tripeptidil-Peptidasas/genética , Dipeptidil-Peptidasas y Tripeptidil-Peptidasas/metabolismo , Exodesoxirribonucleasas , Fase G1 , Fase G2 , Células HCT116 , Humanos , Proteínas Inhibidoras de la Apoptosis/genética , Proteínas Inhibidoras de la Apoptosis/metabolismo , Modelos Moleculares , Proteínas Nucleares/química , Proteínas Nucleares/genética , Fosforilación , Unión Proteica , Dominios y Motivos de Interacción de Proteínas , Fase S , Serina Proteasas/genética , Serina Proteasas/metabolismo , Complejo Shelterina , Relación Estructura-Actividad , Telómero/genética , Telómero/patología , Proteínas de Unión a Telómeros/genética , Proteínas de Unión a Telómeros/metabolismo , Proteína 2 de Unión a Repeticiones Teloméricas/química , Proteína 2 de Unión a Repeticiones Teloméricas/genética
17.
Mol Cell ; 65(5): 818-831.e5, 2017 Mar 02.
Artículo en Inglés | MEDLINE | ID: mdl-28216227

RESUMEN

Telomeric repeat binding factor 1 (TRF1) is essential to the maintenance of telomere chromatin structure and integrity. However, how telomere integrity is maintained, especially in response to damage, remains poorly understood. Here, we identify Nek7, a member of the Never in Mitosis Gene A (NIMA) kinase family, as a regulator of telomere integrity. Nek7 is recruited to telomeres and stabilizes TRF1 at telomeres after damage in an ATM activation-dependent manner. Nek7 deficiency leads to telomere aberrations, long-lasting γH2AX and 53BP1 foci, and augmented cell death upon oxidative telomeric DNA damage. Mechanistically, Nek7 interacts with and phosphorylates TRF1 on Ser114, which prevents TRF1 from binding to Fbx4, an Skp1-Cul1-F box E3 ligase subunit, thereby alleviating proteasomal degradation of TRF1, leading to a stable association of TRF1 with Tin2 to form a shelterin complex. Our data reveal a mechanism of efficient protection of telomeres from damage through Nek7-dependent stabilization of TRF1.


Asunto(s)
Daño del ADN , Quinasas Relacionadas con NIMA/metabolismo , Estrés Oxidativo , Proteínas de Unión a Telómeros/metabolismo , Telómero/enzimología , Proteínas de la Ataxia Telangiectasia Mutada/genética , Proteínas de la Ataxia Telangiectasia Mutada/metabolismo , Sitios de Unión , Proteínas F-Box/genética , Proteínas F-Box/metabolismo , Células HEK293 , Células HeLa , Histonas/metabolismo , Humanos , Quinasas Relacionadas con NIMA/genética , Fosforilación , Complejo de la Endopetidasa Proteasomal/metabolismo , Unión Proteica , Estabilidad Proteica , Interferencia de ARN , Complejo Shelterina , Telómero/genética , Telómero/efectos de la radiación , Proteínas de Unión a Telómeros/genética , Factores de Tiempo , Transfección , Proteína 1 de Unión al Supresor Tumoral P53/genética , Proteína 1 de Unión al Supresor Tumoral P53/metabolismo , Ubiquitinación
18.
PLoS Genet ; 18(7): e1010308, 2022 07.
Artículo en Inglés | MEDLINE | ID: mdl-35849625

RESUMEN

The conserved shelterin complex caps chromosome ends to protect telomeres and regulate telomere replication. In fission yeast Schizosaccharomyces pombe, shelterin consists of telomeric single- and double-stranded DNA-binding modules Pot1-Tpz1 and Taz1-Rap1 connected by Poz1, and a specific component Ccq1. While individual structures of the two DNA-binding OB folds of Pot1 (Pot1OB1-GGTTAC and Pot1OB2-GGTTACGGT) are available, structural insight into recognition of telomeric repeats with spacers by the complete DNA-binding domain (Pot1DBD) remains an open question. Moreover, structural information about the Tpz1-Ccq1 interaction requires to be revealed for understanding how the specific component Ccq1 of S. pombe shelterin is recruited to telomeres to function as an interacting hub. Here, we report the crystal structures of Pot1DBD-single-stranded-DNA, Pot1372-555-Tpz1185-212 and Tpz1425-470-Ccq1123-439 complexes and propose an integrated model depicting the assembly mechanism of the shelterin complex at telomeres. The structure of Pot1DBD-DNA unveils how Pot1 recognizes S. pombe degenerate telomeric sequences. Our analyses of Tpz1-Ccq1 reveal structural basis for the essential role of the Tpz1-Ccq1 interaction in telomere recruitment of Ccq1 that is required for telomere maintenance and telomeric heterochromatin formation. Overall, our findings provide valuable structural information regarding interactions within fission yeast shelterin complex at 3' ss telomeric overhang.


Asunto(s)
Proteínas de Schizosaccharomyces pombe , Schizosaccharomyces , Telomerasa , Proteínas Portadoras/genética , ADN de Cadena Simple , Unión Proteica , Schizosaccharomyces/genética , Schizosaccharomyces/metabolismo , Proteínas de Schizosaccharomyces pombe/metabolismo , Complejo Shelterina , Telomerasa/genética , Telómero/genética , Telómero/metabolismo , Proteínas de Unión a Telómeros/genética , Proteínas de Unión a Telómeros/metabolismo
19.
Semin Cancer Biol ; 95: 52-74, 2023 10.
Artículo en Inglés | MEDLINE | ID: mdl-37473825

RESUMEN

Head and neck tumors (HNTs) constitute a multifaceted ensemble of pathologies that primarily involve regions such as the oral cavity, pharynx, and nasal cavity. The intricate anatomical structure of these regions poses considerable challenges to efficacious treatment strategies. Despite the availability of myriad treatment modalities, the overall therapeutic efficacy for HNTs continues to remain subdued. In recent years, the deployment of artificial intelligence (AI) in healthcare practices has garnered noteworthy attention. AI modalities, inclusive of machine learning (ML), neural networks (NNs), and deep learning (DL), when amalgamated into the holistic management of HNTs, promise to augment the precision, safety, and efficacy of treatment regimens. The integration of AI within HNT management is intricately intertwined with domains such as medical imaging, bioinformatics, and medical robotics. This article intends to scrutinize the cutting-edge advancements and prospective applications of AI in the realm of HNTs, elucidating AI's indispensable role in prevention, diagnosis, treatment, prognostication, research, and inter-sectoral integration. The overarching objective is to stimulate scholarly discourse and invigorate insights among medical practitioners and researchers to propel further exploration, thereby facilitating superior therapeutic alternatives for patients.


Asunto(s)
Inteligencia Artificial , Neoplasias de Cabeza y Cuello , Humanos , Aprendizaje Automático , Redes Neurales de la Computación , Neoplasias de Cabeza y Cuello/diagnóstico , Neoplasias de Cabeza y Cuello/terapia , Diagnóstico por Imagen/métodos
20.
BMC Genomics ; 25(1): 231, 2024 Mar 02.
Artículo en Inglés | MEDLINE | ID: mdl-38431573

RESUMEN

BACKGROUND: Telomere length is a critical metric linked to aging, health, and disease. Currently, the exploration of target proteins related to telomere length is usually limited to the context of aging and specific diseases, which limits the discovery of more relevant drug targets. This study integrated large-scale plasma cis-pQTLs data and telomere length GWAS datasets. We used Mendelian randomization(MR) to identify drug target proteins for telomere length, providing essential clues for future precision therapy and targeted drug development. METHODS: Using plasma cis-pQTLs data from a previous GWAS study (3,606 Pqtls associated with 2,656 proteins) and a GWAS dataset of telomere length (sample size: 472,174; GWAS ID: ieu-b-4879) from UK Biobank, using MR, external validation, and reverse causality testing, we identified essential drug target proteins for telomere length. We also performed co-localization, Phenome-wide association studies and enrichment analysis, protein-protein interaction network construction, search for existing intervening drugs, and potential drug/compound prediction for these critical targets to strengthen and expand our findings. RESULTS: After Bonferron correction (p < 0.05/734), RPN1 (OR: 0.96; 95%CI: (0.95, 0.97)), GDI2 (OR: 0.94; 95%CI: (0.92, 0.96)), NT5C (OR: 0.97; 95%CI: (0.95, 0.98)) had a significant negative causal association with telomere length; TYRO3 (OR: 1.11; 95%CI: (1.09, 1.15)) had a significant positive causal association with telomere length. GDI2 shared the same genetic variants with telomere length (coloc.abf-PPH 4 > 0.8). CONCLUSION: Genetically determined plasma RPN1, GDI2, NT5C, and TYRO3 have significant causal effects on telomere length and can potentially be drug targets. Further exploration of the role and mechanism of these proteins/genes in regulating telomere length is needed.


Asunto(s)
Sistemas de Liberación de Medicamentos , Estudio de Asociación del Genoma Completo , Fenómica , Telómero/genética , Análisis de la Aleatorización Mendeliana
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