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1.
Mol Cell ; 84(13): 2405-2406, 2024 Jul 11.
Artículo en Inglés | MEDLINE | ID: mdl-38996456

RESUMEN

In this issue of Molecular Cell, Razew et al.1 and Sabath et al.2 assign function to an unexplored module of the Integrator (INT) complex, expanding the toolbox of this genome-wide attenuator of RNA polymerase II (RNAPII) transcription.


Asunto(s)
ARN Polimerasa II , Transcripción Genética , ARN Polimerasa II/metabolismo , ARN Polimerasa II/genética
2.
Proc Natl Acad Sci U S A ; 119(48): e2210532119, 2022 11 29.
Artículo en Inglés | MEDLINE | ID: mdl-36409902

RESUMEN

A hexanucleotide repeat expansion in intron 1 of the C9orf72 gene is the most common genetic cause of amyotrophic lateral sclerosis and frontotemporal dementia, or c9ALS/FTD. The RNA transcribed from the expansion, r(G4C2)exp, causes various pathologies, including intron retention, aberrant translation that produces toxic dipeptide repeat proteins (DPRs), and sequestration of RNA-binding proteins (RBPs) in RNA foci. Here, we describe a small molecule that potently and selectively interacts with r(G4C2)exp and mitigates disease pathologies in spinal neurons differentiated from c9ALS patient-derived induced pluripotent stem cells (iPSCs) and in two c9ALS/FTD mouse models. These studies reveal a mode of action whereby a small molecule diminishes intron retention caused by the r(G4C2)exp and allows the liberated intron to be eliminated by the nuclear RNA exosome, a multi-subunit degradation complex. Our findings highlight the complexity of mechanisms available to RNA-binding small molecules to alleviate disease pathologies and establishes a pipeline for the design of brain penetrant small molecules targeting RNA with novel modes of action in vivo.


Asunto(s)
Exosomas , Demencia Frontotemporal , Animales , Ratones , Demencia Frontotemporal/metabolismo , Proteína C9orf72/genética , Proteína C9orf72/metabolismo , ARN/genética , Exosomas/metabolismo , Barrera Hematoencefálica/metabolismo , Complejo Multienzimático de Ribonucleasas del Exosoma/metabolismo , Encéfalo/metabolismo , Proteínas de Unión al ARN/genética , Proteínas de Unión al ARN/metabolismo , ARN Nuclear
3.
Nucleic Acids Res ; 48(12): 6943-6953, 2020 07 09.
Artículo en Inglés | MEDLINE | ID: mdl-32463452

RESUMEN

ARS2 is a conserved protein centrally involved in both nuclear RNA productive and destructive processes. To map features of ARS2 promoting RNA decay, we utilized two different RNA reporters, one of which depends on direct ARS2 tethering for its degradation. In both cases, ARS2 triggers a degradation phenotype aided by its interaction with the poly(A) tail exosome targeting (PAXT) connection. Interestingly, C-terminal amino acids of ARS2, responsible for binding the RNA 5'cap binding complex (CBC), become dispensable when ARS2 is directly tethered to the reporter RNA. In contrast, the Zinc-finger (ZnF) domain of ARS2 is essential for the decay of both reporters and consistently co-immunoprecipitation analyses reveal a necessity of this domain for the interaction of ARS2 with the PAXT-associated RNA helicase MTR4. Taken together, our results map the domains of ARS2 underlying two essential properties of the protein: its RNP targeting ability and its capacity to recruit the RNA decay machinery.


Asunto(s)
Proteínas Nucleares/genética , ARN Helicasas/genética , Estabilidad del ARN/genética , ARN Mensajero/genética , Complejo Multienzimático de Ribonucleasas del Exosoma/genética , Células HEK293 , Humanos , Complejo Proteico Nuclear de Unión a la Caperuza/genética , Proteínas Nucleares/química , Dominios Proteicos/genética , ARN Helicasas/química , ARN Mensajero/química , ARN Nuclear/química , ARN Nuclear/genética
4.
Nucleic Acids Res ; 46(11): 5455-5469, 2018 06 20.
Artículo en Inglés | MEDLINE | ID: mdl-29659982

RESUMEN

Mammalian gene promoters and enhancers share many properties. They are composed of a unified promoter architecture of divergent transcripton initiation and gene promoters may exhibit enhancer function. However, it is currently unclear how expression strength of a regulatory element relates to its enhancer strength and if the unifying architecture is conserved across Metazoa. Here we investigate the transcription initiation landscape and its associated RNA decay in Drosophila melanogaster. We find that the majority of active gene-distal enhancers and a considerable fraction of gene promoters are divergently transcribed. We observe quantitative relationships between enhancer potential, expression level and core promoter strength, providing an explanation for indirectly related histone modifications that are reflecting expression levels. Lowly abundant unstable RNAs initiated from weak core promoters are key characteristics of gene-distal developmental enhancers, while the housekeeping enhancer strengths of gene promoters reflect their expression strengths. The seemingly separable layer of regulation by gene promoters with housekeeping enhancer potential is also indicated by chromatin interaction data. Our results suggest a unified promoter architecture of many D. melanogaster regulatory elements, that is universal across Metazoa, whose regulatory functions seem to be related to their core promoter elements.


Asunto(s)
Drosophila melanogaster/genética , Elementos de Facilitación Genéticos/genética , Regiones Promotoras Genéticas/genética , Sitio de Iniciación de la Transcripción , Transcripción Genética/genética , Animales , Línea Celular , Código de Histonas/fisiología , ARN/metabolismo , Estabilidad del ARN/genética , Activación Transcripcional/genética
5.
Cell Rep ; 43(1): 113639, 2024 01 23.
Artículo en Inglés | MEDLINE | ID: mdl-38175753

RESUMEN

The nuclear cap-binding complex (CBC) coordinates co-transcriptional maturation, transport, or degradation of nascent RNA polymerase II (Pol II) transcripts. CBC with its partner ARS2 forms mutually exclusive complexes with diverse "effectors" that promote either productive or destructive outcomes. Combining AlphaFold predictions with structural and biochemical validation, we show how effectors NCBP3, NELF-E, ARS2, PHAX, and ZC3H18 form competing binary complexes with CBC and how PHAX, NCBP3, ZC3H18, and other effectors compete for binding to ARS2. In ternary CBC-ARS2 complexes with PHAX, NCBP3, or ZC3H18, ARS2 is responsible for the initial effector recruitment but inhibits their direct binding to the CBC. We show that in vivo ZC3H18 binding to both CBC and ARS2 is required for nuclear RNA degradation. We propose that recruitment of PHAX to CBC-ARS2 can lead, with appropriate cues, to competitive displacement of ARS2 and ZC3H18 from the CBC, thus promoting a productive rather than a degradative RNA fate.


Asunto(s)
Complejo Proteico Nuclear de Unión a la Caperuza , ARN , Unión Competitiva , Complejo Proteico Nuclear de Unión a la Caperuza/química , ARN/genética , ARN Polimerasa II/metabolismo , ARN Nuclear
6.
Nat Commun ; 14(1): 696, 2023 02 08.
Artículo en Inglés | MEDLINE | ID: mdl-36755023

RESUMEN

Dynamic RNA-protein interactions govern the co-transcriptional packaging of RNA polymerase II (RNAPII)-derived transcripts. Yet, our current understanding of this process in vivo primarily stems from steady state analysis. To remedy this, we here conduct temporal-iCLIP (tiCLIP), combining RNAPII transcriptional synchronisation with UV cross-linking of RNA-protein complexes at serial timepoints. We apply tiCLIP to the RNA export adaptor, ALYREF; a component of the Nuclear Exosome Targeting (NEXT) complex, RBM7; and the nuclear cap binding complex (CBC). Regardless of function, all tested factors interact with nascent RNA as it exits RNAPII. Moreover, we demonstrate that the two transesterification steps of pre-mRNA splicing temporally separate ALYREF and RBM7 binding to splicing intermediates, and that exon-exon junction density drives RNA 5'end binding of ALYREF. Finally, we identify underappreciated steps in snoRNA 3'end processing performed by RBM7. Altogether, our data provide a temporal view of RNA-protein interactions during the early phases of transcription.


Asunto(s)
Núcleo Celular , Proteínas de Unión al ARN , Proteínas de Unión al ARN/metabolismo , Núcleo Celular/metabolismo , Precursores del ARN/metabolismo , Empalme del ARN , ARN Polimerasa II/metabolismo , ARN Nucleolar Pequeño/metabolismo
7.
Nat Commun ; 10(1): 3393, 2019 07 29.
Artículo en Inglés | MEDLINE | ID: mdl-31358741

RESUMEN

The nuclear exosome and its essential co-factor, the RNA helicase MTR4, play crucial roles in several RNA degradation pathways. Besides unwinding RNA substrates for exosome-mediated degradation, MTR4 associates with RNA-binding proteins that function as adaptors in different RNA processing and decay pathways. Here, we identify and characterize the interactions of human MTR4 with a ribosome processing adaptor, NVL, and with ZCCHC8, an adaptor involved in the decay of small nuclear RNAs. We show that the unstructured regions of NVL and ZCCHC8 contain short linear motifs that bind the MTR4 arch domain in a mutually exclusive manner. These short sequences diverged from the arch-interacting motif (AIM) of yeast rRNA processing factors. Our results suggest that nuclear exosome adaptors have evolved canonical and non-canonical AIM sequences to target human MTR4 and demonstrate the versatility and specificity with which the MTR4 arch domain can recruit a repertoire of different RNA-binding proteins.


Asunto(s)
ATPasas Asociadas con Actividades Celulares Diversas/metabolismo , Proteínas Portadoras/metabolismo , Núcleo Celular/metabolismo , Exosomas/genética , Proteínas Nucleares/metabolismo , ARN Helicasas/metabolismo , ATPasas Asociadas con Actividades Celulares Diversas/química , ATPasas Asociadas con Actividades Celulares Diversas/genética , Secuencia de Aminoácidos , Sitios de Unión/genética , Proteínas Portadoras/química , Proteínas Portadoras/genética , Cristalografía por Rayos X , Exosomas/metabolismo , Células HeLa , Humanos , Mutación , Proteínas Nucleares/química , Proteínas Nucleares/genética , Unión Proteica , Dominios Proteicos , ARN Helicasas/química , ARN Helicasas/genética , Proteínas de Unión al ARN/química , Proteínas de Unión al ARN/genética , Proteínas de Unión al ARN/metabolismo , Homología de Secuencia de Aminoácido
8.
Nat Commun ; 5: 5336, 2014 Nov 12.
Artículo en Inglés | MEDLINE | ID: mdl-25387874

RESUMEN

Mammalian genomes are pervasively transcribed, yielding a complex transcriptome with high variability in composition and cellular abundance. Although recent efforts have identified thousands of new long non-coding (lnc) RNAs and demonstrated a complex transcriptional repertoire produced by protein-coding (pc) genes, limited progress has been made in distinguishing functional RNA from spurious transcription events. This is partly due to present RNA classification, which is typically based on technical rather than biochemical criteria. Here we devise a strategy to systematically categorize human RNAs by their sensitivity to the ribonucleolytic RNA exosome complex and by the nature of their transcription initiation. These measures are surprisingly effective at correctly classifying annotated transcripts, including lncRNAs of known function. The approach also identifies uncharacterized stable lncRNAs, hidden among a vast majority of unstable transcripts. The predictive power of the approach promises to streamline the functional analysis of known and novel RNAs.


Asunto(s)
Estabilidad del ARN/fisiología , ARN/clasificación , Exosomas/genética , Exosomas/fisiología , Células HeLa , Humanos , Regiones Promotoras Genéticas/genética , Regiones Promotoras Genéticas/fisiología , ARN/fisiología , Estabilidad del ARN/genética , ARN no Traducido/fisiología , Iniciación de la Transcripción Genética/fisiología , Transcripción Genética/fisiología
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