RESUMO
During the last ten years, developments in cryo-electron microscopy have transformed our understanding of eukaryotic ribosome assembly. As a result, the field has advanced from a list of the vast array of ribosome assembly factors toward an emerging molecular movie in which individual frames are represented by structures of stable ribosome assembly intermediates with complementary biochemical and genetic data. In this review, we discuss the mechanisms driving the assembly of yeast and human small and large ribosomal subunits. A particular emphasis is placed on the most recent findings that illustrate key concepts of ribosome assembly, such as folding of preribosomal RNA, the enforced chronology of assembly, enzyme-mediated irreversible transitions, and proofreading of preribosomal particles.
Assuntos
Microscopia Crioeletrônica , Proteínas Ribossômicas , Ribossomos , Humanos , Ribossomos/metabolismo , Ribossomos/ultraestrutura , Ribossomos/química , Ribossomos/genética , Proteínas Ribossômicas/metabolismo , Proteínas Ribossômicas/química , Proteínas Ribossômicas/genética , RNA Ribossômico/metabolismo , RNA Ribossômico/química , RNA Ribossômico/genética , Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/genética , Modelos Moleculares , Células Eucarióticas/metabolismo , Células Eucarióticas/ultraestrutura , Dobramento de RNA , Subunidades Ribossômicas Menores de Eucariotos/metabolismo , Subunidades Ribossômicas Menores de Eucariotos/química , Subunidades Ribossômicas Menores de Eucariotos/genética , Subunidades Ribossômicas Menores de Eucariotos/ultraestrutura , AnimaisRESUMO
Cellular senescence is an irreversible state of cell-cycle arrest induced by various stresses, including aberrant oncogene activation, telomere shortening, and DNA damage. Through a genome-wide screen, we discovered a conserved small nucleolar RNA (snoRNA), SNORA13, that is required for multiple forms of senescence in human cells and mice. Although SNORA13 guides the pseudouridylation of a conserved nucleotide in the ribosomal decoding center, loss of this snoRNA minimally impacts translation. Instead, we found that SNORA13 negatively regulates ribosome biogenesis. Senescence-inducing stress perturbs ribosome biogenesis, resulting in the accumulation of free ribosomal proteins (RPs) that trigger p53 activation. SNORA13 interacts directly with RPL23, decreasing its incorporation into maturing 60S subunits and, consequently, increasing the pool of free RPs, thereby promoting p53-mediated senescence. Thus, SNORA13 regulates ribosome biogenesis and the p53 pathway through a non-canonical mechanism distinct from its role in guiding RNA modification. These findings expand our understanding of snoRNA functions and their roles in cellular signaling.
RESUMO
Ribosomes, which synthesize the proteins of a cell, comprise ribosomal RNA and ribosomal proteins, which coassemble hierarchically during a process termed ribosome biogenesis. Historically, biochemical and molecular biology approaches have revealed how preribosomal particles form and mature in consecutive steps, starting in the nucleolus and terminating after nuclear export into the cytoplasm. However, only recently, due to the revolution in cryo-electron microscopy, could pseudoatomic structures of different preribosomal particles be obtained. Together with in vitro maturation assays, these findings shed light on how nascent ribosomes progress stepwise along a dynamic biogenesis pathway. Preribosomes assemble gradually, chaperoned by a myriad of assembly factors and small nucleolar RNAs, before they reach maturity and enter translation. This information will lead to a better understanding of how ribosome synthesis is linked to other cellular pathways in humans and how it can cause diseases, including cancer, if disturbed.
Assuntos
Eucariotos/metabolismo , RNA Ribossômico/metabolismo , Proteínas Ribossômicas/metabolismo , Ribossomos/metabolismo , Nucléolo Celular/metabolismo , Microscopia Crioeletrônica , Humanos , Biogênese de Organelas , Multimerização ProteicaRESUMO
The synthesis of new ribosomes begins during transcription of the rRNA and is widely assumed to follow an orderly 5' to 3' gradient. To visualize co-transcriptional assembly of ribosomal protein-RNA complexes in real time, we developed a single-molecule platform that simultaneously monitors transcription and protein association with the elongating transcript. Unexpectedly, the early assembly protein uS4 binds newly made pre-16S rRNA only transiently, likely due to non-native folding of the rRNA during transcription. Stable uS4 binding became more probable only in the presence of additional ribosomal proteins that bind upstream and downstream of protein uS4 by allowing productive assembly intermediates to form earlier. We propose that dynamic sampling of elongating RNA by multiple proteins overcomes heterogeneous RNA folding, preventing assembly bottlenecks and initiating assembly within the transcription time window. This may be a common feature of transcription-coupled RNP assembly.
Assuntos
Ribonucleoproteínas/metabolismo , Transcrição Gênica , Fluorescência , Modelos Biológicos , Ligação Proteica , Estabilidade Proteica , Precursores de RNA/biossíntese , Precursores de RNA/química , Precursores de RNA/genética , Precursores de RNA/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteínas Ribossômicas/metabolismo , Ribossomos/metabolismo , Elongação da Transcrição GenéticaRESUMO
Ribosome biogenesis is a complex and highly energy-demanding process that requires the concerted action of all three nuclear RNA polymerases (Pol I-III) in eukaryotes. The three largest ribosomal RNAs (rRNAs) originate from a precursor transcript (pre-rRNA) that is encoded by multicopy genes located in the nucleolus. Transcription of these rRNA genes (rDNA) by Pol I is the key regulation step in ribosome production and is tightly controlled by an intricate network of signaling pathways and epigenetic mechanisms. In this article, we give an overview of the composition of the basal Pol I machinery and rDNA chromatin. We discuss rRNA gene regulation in response to environmental signals and developmental cues and focus on perturbations occurring in diseases linked to either excessive or limited rRNA levels. Finally, we discuss the emerging view that rDNA integrity and activity may be involved in the aging process.
Assuntos
RNA Polimerase I/genética , RNA Polimerase I/metabolismo , Envelhecimento/genética , Envelhecimento/metabolismo , Animais , Diferenciação Celular/genética , Diferenciação Celular/fisiologia , Cromatina/genética , Cromatina/metabolismo , DNA Ribossômico/genética , DNA Ribossômico/metabolismo , Epigênese Genética , Humanos , Modelos Biológicos , Família Multigênica , Neoplasias/genética , Neoplasias/metabolismo , Células-Tronco Pluripotentes/citologia , Células-Tronco Pluripotentes/metabolismo , RNA Ribossômico/genética , RNA Ribossômico/metabolismo , Ribossomos/genética , Ribossomos/metabolismo , Transdução de Sinais , Transcrição GênicaRESUMO
Mdn1 is an essential AAA (ATPase associated with various activities) protein that removes assembly factors from distinct precursors of the ribosomal 60S subunit. However, Mdn1's large size (â¼5,000 amino acid [aa]) and its limited homology to other well-studied proteins have restricted our understanding of its remodeling function. Here, we present structures for S. pombe Mdn1 in the presence of AMPPNP at up to â¼4 Å or ATP plus Rbin-1, a chemical inhibitor, at â¼8 Å resolution. These data reveal that Mdn1's MIDAS domain is tethered to its ring-shaped AAA domain through an â¼20 nm long structured linker and a flexible â¼500 aa Asp/Glu-rich motif. We find that the MIDAS domain, which also binds other ribosome-assembly factors, docks onto the AAA ring in a nucleotide state-specific manner. Together, our findings reveal how conformational changes in the AAA ring can be directly transmitted to the MIDAS domain and thereby drive the targeted release of assembly factors from ribosomal 60S-subunit precursors.
Assuntos
ATPases Associadas a Diversas Atividades Celulares/química , Simulação de Dinâmica Molecular , Proteínas de Schizosaccharomyces pombe/química , Schizosaccharomyces/enzimologia , Motivos de Aminoácidos , Animais , Sítios de Ligação , Microscopia Crioeletrônica , Biogênese de Organelas , Ligação Proteica , Subunidades Ribossômicas Maiores de Eucariotos/metabolismo , Células Sf9 , SpodopteraRESUMO
Eukaryotic 60S ribosomal subunits are comprised of three rRNAs and â¼50 ribosomal proteins. The initial steps of their formation take place in the nucleolus, but, owing to a lack of structural information, this process is poorly understood. Using cryo-EM, we solved structures of early 60S biogenesis intermediates at 3.3 Å to 4.5 Å resolution, thereby providing insights into their sequential folding and assembly pathway. Besides revealing distinct immature rRNA conformations, we map 25 assembly factors in six different assembly states. Notably, the Nsa1-Rrp1-Rpf1-Mak16 module stabilizes the solvent side of the 60S subunit, and the Erb1-Ytm1-Nop7 complex organizes and connects through Erb1's meandering N-terminal extension, eight assembly factors, three ribosomal proteins, and three 25S rRNA domains. Our structural snapshots reveal the order of integration and compaction of the six major 60S domains within early nucleolar 60S particles developing stepwise from the solvent side around the exit tunnel to the central protuberance.
Assuntos
Chaetomium/química , Biogênese de Organelas , Subunidades Ribossômicas Maiores de Eucariotos/química , Chaetomium/citologia , Microscopia Crioeletrônica , Redes e Vias Metabólicas , Modelos Moleculares , Dobramento de RNA , Ribonucleoproteínas/químicaRESUMO
All cellular proteins are synthesized by ribosomes, whose biogenesis in eukaryotes is a complex multi-step process completed within minutes. Several chemical inhibitors of ribosome function are available and used as tools or drugs. By contrast, we lack potent validated chemical probes to analyze the dynamics of eukaryotic ribosome assembly. Here, we combine chemical and genetic approaches to discover ribozinoindoles (or Rbins), potent and reversible triazinoindole-based inhibitors of eukaryotic ribosome biogenesis. Analyses of Rbin sensitivity and resistance conferring mutations in fission yeast, along with biochemical assays with recombinant proteins, provide evidence that Rbins' physiological target is Midasin, an essential â¼540-kDa AAA+ (ATPases associated with diverse cellular activities) protein. Using Rbins to acutely inhibit or activate Midasin function, in parallel experiments with inhibitor-sensitive or inhibitor-resistant cells, we uncover Midasin's role in assembling Nsa1 particles, nucleolar precursors of the 60S subunit. Together, our findings demonstrate that Rbins are powerful probes for eukaryotic ribosome assembly.
Assuntos
Adenosina Trifosfatases/antagonistas & inibidores , Inibidores Enzimáticos/farmacologia , Indóis/farmacologia , Subunidades Ribossômicas Maiores de Eucariotos/efeitos dos fármacos , Subunidades Ribossômicas Maiores de Eucariotos/metabolismo , Proteínas de Schizosaccharomyces pombe/antagonistas & inibidores , Triazinas/farmacologia , Inibidores Enzimáticos/química , Inibidores Enzimáticos/isolamento & purificação , Indóis/química , Indóis/isolamento & purificação , Schizosaccharomyces/efeitos dos fármacos , Schizosaccharomyces/metabolismo , Relação Estrutura-Atividade , Triazinas/química , Triazinas/isolamento & purificaçãoRESUMO
The nucleolus is the largest biomolecular condensate and facilitates transcription, processing, and assembly of ribosomal RNA (rRNA). Although nucleolar function is thought to require multiphase liquid-like properties, nucleolar fluidity and its connection to the highly coordinated transport and biogenesis of ribosomal subunits are poorly understood. Here, we use quantitative imaging, mathematical modeling, and pulse-chase nucleotide labeling to examine nucleolar material properties and rRNA dynamics. The mobility of rRNA is several orders of magnitude slower than that of nucleolar proteins, with rRNA steadily moving away from the transcriptional sites in a slow (â¼1 Å/s), radially directed fashion. This constrained but directional mobility, together with polymer physics-based calculations, suggests that nascent rRNA forms an entangled gel, whose constant production drives outward flow. We propose a model in which progressive maturation of nascent rRNA reduces its initial entanglement, fluidizing the nucleolar periphery to facilitate the release of assembled pre-ribosomal particles.
Assuntos
RNA Ribossômico , RNA , RNA/genética , RNA Ribossômico/genética , Condensados Biomoleculares , Nucléolo Celular/genética , Proteínas Nucleares/genéticaRESUMO
Ribosome biogenesis takes place in the nucleolus, a nuclear membrane-less organelle. Although well studied, it remains unknown how nascent ribosomal subunits separate from the central chromatin compartment and move to the outer granular component, where maturation occurs. We find that the Schizosaccharomyces pombe nucleophosmin-like protein Fkbp39 localizes to rDNA sites encoding the 60S subunit rRNA, and this localization contributes to its specific association with nascent 60S subunits. Fkbp39 dissociates from chromatin to bind nascent 60S subunits, causing the latter to partition away from chromatin and from nascent 40S subunits through liquid-liquid phase separation. In vivo, Fkbp39 binding directs the translocation of nascent 60S subunits toward the nucleophosmin-rich granular component. This process increases the efficiency of 60S subunit assembly, facilitating the incorporation of 60S RNA domain III. Thus, chromatin localization determines the specificity of nucleophosmin in sorting nascent ribosomal subunits and coordinates their movement into specialized assembly compartments within the nucleolus.
Assuntos
Cromatina , Schizosaccharomyces , Cromatina/genética , Nucleofosmina , Nucléolo Celular/genética , Membrana Nuclear , Schizosaccharomyces/genética , Ribossomos/genéticaRESUMO
Ribosomal RNAs (rRNAs) are the most abundant cellular RNAs, and their synthesis from rDNA repeats by RNA polymerase I accounts for the bulk of all transcription. Despite substantial variation in rRNA transcription rates across cell types, little is known about cell-type-specific factors that bind rDNA and regulate rRNA transcription to meet tissue-specific needs. Using hematopoiesis as a model system, we mapped about 2,200 ChIP-seq datasets for 250 transcription factors (TFs) and chromatin proteins to human and mouse rDNA and identified robust binding of multiple TF families to canonical TF motifs on rDNA. Using a 47S-FISH-Flow assay developed for nascent rRNA quantification, we demonstrated that targeted degradation of C/EBP alpha (CEBPA), a critical hematopoietic TF with conserved rDNA binding, caused rapid reduction in rRNA transcription due to reduced RNA Pol I occupancy. Our work identifies numerous potential rRNA regulators and provides a template for dissection of TF roles in rRNA transcription.
Assuntos
RNA Polimerase I , Fatores de Transcrição , Humanos , Camundongos , Animais , RNA Polimerase I/genética , RNA Polimerase I/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , RNA Ribossômico/genética , Transcrição Gênica , DNA Ribossômico/genética , RNA , CromatinaRESUMO
ADP-ribosylation (ADPRylation) is a post-translational modification of proteins catalyzed by ADP-ribosyl transferase (ART) enzymes, including nuclear PARPs (e.g., PARP1 and PARP2). Historically, studies of ADPRylation and PARPs have focused on DNA damage responses in cancers, but more recent studies elucidate diverse roles in a broader array of biological processes. Here, we summarize the expanding array of molecular mechanisms underlying the biological functions of nuclear PARPs with a focus on PARP1, the founding member of the family. This includes roles in DNA repair, chromatin regulation, gene expression, ribosome biogenesis, and RNA biology. We also present new concepts in PARP1-dependent regulation, including PAR-dependent post-translational modifications, "ADPR spray," and PAR-mediated biomolecular condensate formation. Moreover, we review advances in the therapeutic mechanisms of PARP inhibitors (PARPi) as well as the progress on the mechanisms of PARPi resistance. Collectively, the recent progress in the field has yielded new insights into the expanding universe of PARP1-mediated molecular and therapeutic mechanisms in a variety of biological processes.
Assuntos
ADP-Ribosilação , Reparo do DNA , Cromatina/genética , Dano ao DNA , Poli(ADP-Ribose) Polimerase-1/genética , Poli(ADP-Ribose) Polimerase-1/metabolismo , Processamento de Proteína Pós-Traducional , RNA/metabolismoRESUMO
Ribosome biogenesis is a fundamental multi-step cellular process that culminates in the formation of ribosomal subunits, whose production and modification are regulated by numerous biogenesis factors. In this study, we analyze physiologic prokaryotic ribosome biogenesis by isolating bona fide pre-50S subunits from an Escherichia coli strain with the biogenesis factor ObgE, affinity tagged at its native gene locus. Our integrative structural approach reveals a network of interacting biogenesis factors consisting of YjgA, RluD, RsfS, and ObgE on the immature pre-50S subunit. In addition, our study provides mechanistic insight into how the GTPase ObgE, in concert with other biogenesis factors, facilitates the maturation of the 50S functional core and reveals both conserved and divergent evolutionary features of ribosome biogenesis between prokaryotes and eukaryotes.
Assuntos
Proteínas de Escherichia coli , Evolução Molecular , Loci Gênicos , Hidroliases , Proteínas Monoméricas de Ligação ao GTP , Subunidades Ribossômicas Maiores de Bactérias , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Hidroliases/química , Hidroliases/genética , Hidroliases/metabolismo , Proteínas Monoméricas de Ligação ao GTP/química , Proteínas Monoméricas de Ligação ao GTP/genética , Proteínas Monoméricas de Ligação ao GTP/metabolismo , Subunidades Ribossômicas Maiores de Bactérias/química , Subunidades Ribossômicas Maiores de Bactérias/genética , Subunidades Ribossômicas Maiores de Bactérias/metabolismoRESUMO
Ribosome assembly is catalyzed by numerous trans-acting factors and coupled with irreversible pre-rRNA processing, driving the pathway toward mature ribosomal subunits. One decisive step early in this progression is removal of the 5' external transcribed spacer (5'-ETS), an RNA extension at the 18S rRNA that is integrated into the huge 90S pre-ribosome structure. Upon endo-nucleolytic cleavage at an internal site, A1, the 5'-ETS is separated from the 18S rRNA and degraded. Here we present biochemical and cryo-electron microscopy analyses that depict the RNA exosome, a major 3'-5' exoribonuclease complex, in a super-complex with the 90S pre-ribosome. The exosome is docked to the 90S through its co-factor Mtr4 helicase, a processive RNA duplex-dismantling helicase, which strategically positions the exosome at the base of 5'-ETS helices H9-H9', which are dislodged in our 90S-exosome structures. These findings suggest a direct role of the exosome in structural remodeling of the 90S pre-ribosome to drive eukaryotic ribosome synthesis.
Assuntos
RNA Helicases DEAD-box/química , Endorribonucleases/química , Exonucleases/química , Complexo Multienzimático de Ribonucleases do Exossomo/ultraestrutura , RNA Ribossômico 18S/química , Ribossomos/ultraestrutura , Proteínas de Saccharomyces cerevisiae/química , Saccharomyces cerevisiae/genética , Sítios de Ligação , Microscopia Crioeletrônica , RNA Helicases DEAD-box/genética , RNA Helicases DEAD-box/metabolismo , Endorribonucleases/genética , Endorribonucleases/metabolismo , Exonucleases/genética , Exonucleases/metabolismo , Complexo Multienzimático de Ribonucleases do Exossomo/genética , Complexo Multienzimático de Ribonucleases do Exossomo/metabolismo , Modelos Moleculares , Ligação Proteica , Biossíntese de Proteínas , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Domínios e Motivos de Interação entre Proteínas , Estabilidade de RNA , RNA Ribossômico 18S/genética , RNA Ribossômico 18S/metabolismo , Ribossomos/genética , Ribossomos/metabolismo , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismoRESUMO
It is unknown how ribosomal gene (rDNA) arrays from multiple chromosomal nucleolar organizers (NORs) partition within human nucleoli. Exploration of this paradigm for chromosomal organization is complicated by the shared DNA sequence composition of five NOR-bearing acrocentric chromosome p-arms. Here, we devise a methodology for genetic manipulation of individual NORs. Efficient "scarless" genome editing of rDNA repeats is achieved on "poised" human NORs held within monochromosomal cell hybrids. Subsequent transfer to human cells introduces "active" NORs yielding readily discernible functional customized ribosomes. We reveal that ribosome biogenesis occurs entirely within constrained territories, tethered to individual NORs inside a larger nucleolus.
Assuntos
Nucléolo Celular/metabolismo , Região Organizadora do Nucléolo/genética , Região Organizadora do Nucléolo/metabolismo , Ribossomos/metabolismo , Sequência de Bases , Linhagem Celular , Nucléolo Celular/genética , Cromossomos/metabolismo , Edição de Genes , Humanos , Ribossomos/genéticaRESUMO
In contrast to the bacterial translation machinery, mitoribosomes and mitochondrial translation factors are highly divergent in terms of composition and architecture. There is increasing evidence that the biogenesis of mitoribosomes is an intricate pathway, involving many assembly factors. To better understand this process, we investigated native assembly intermediates of the mitoribosomal large subunit from the human parasite Trypanosoma brucei using cryo-electron microscopy. We identify 28 assembly factors, 6 of which are homologous to bacterial and eukaryotic ribosome assembly factors. They interact with the partially folded rRNA by specifically recognizing functionally important regions such as the peptidyltransferase center. The architectural and compositional comparison of the assembly intermediates indicates a stepwise modular assembly process, during which the rRNA folds toward its mature state. During the process, several conserved GTPases and a helicase form highly intertwined interaction networks that stabilize distinct assembly intermediates. The presented structures provide general insights into mitoribosomal maturation.
Assuntos
Ribossomos Mitocondriais/química , RNA Ribossômico/metabolismo , Subunidades Ribossômicas Maiores/química , Trypanosoma brucei brucei/metabolismo , Microscopia Crioeletrônica , RNA Helicases DEAD-box/química , RNA Helicases DEAD-box/metabolismo , GTP Fosfo-Hidrolases/química , GTP Fosfo-Hidrolases/genética , GTP Fosfo-Hidrolases/metabolismo , Ribossomos Mitocondriais/metabolismo , Modelos Moleculares , Conformação de Ácido Nucleico , RNA Ribossômico/química , Proteínas Ribossômicas/química , Proteínas Ribossômicas/genética , Proteínas Ribossômicas/metabolismo , Subunidades Ribossômicas Maiores/metabolismo , Trypanosoma brucei brucei/genéticaRESUMO
Ribosome assembly is driven by numerous assembly factors, including the Rix1 complex and the AAA ATPase Rea1. These two assembly factors catalyze 60S maturation at two distinct states, triggering poorly understood large-scale structural transitions that we analyzed by cryo-electron microscopy. Two nuclear pre-60S intermediates were discovered that represent previously unknown states after Rea1-mediated removal of the Ytm1-Erb1 complex and reveal how the L1 stalk develops from a pre-mature nucleolar to a mature-like nucleoplasmic state. A later pre-60S intermediate shows how the central protuberance arises, assisted by the nearby Rix1-Rea1 machinery, which was solved in its pre-ribosomal context to molecular resolution. This revealed a Rix12-Ipi32 tetramer anchored to the pre-60S via Ipi1, strategically positioned to monitor this decisive remodeling. These results are consistent with a general underlying principle that temporarily stabilized immature RNA domains are successively remodeled by assembly factors, thereby ensuring failsafe assembly progression.
Assuntos
Subunidades Ribossômicas Maiores de Eucariotos/química , Subunidades Ribossômicas Maiores de Eucariotos/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , ATPases Associadas a Diversas Atividades Celulares/genética , ATPases Associadas a Diversas Atividades Celulares/metabolismo , Nucléolo Celular/genética , Nucléolo Celular/metabolismo , Microscopia Crioeletrônica , Escherichia coli/genética , Modelos Moleculares , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Subunidades Ribossômicas Maiores de Eucariotos/genética , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genéticaRESUMO
ADP-ribosylation (ADPRylation) is a posttranslational modification of proteins discovered nearly six decades ago, but many important questions remain regarding its molecular functions and biological roles, as well as the activity of the ADP-ribose (ADPR) transferase enzymes (PARP family members) that catalyze it. Growing evidence indicates that PARP-mediated ADPRylation events are key regulators of the protein biosynthetic pathway, leading from rDNA transcription and ribosome biogenesis to mRNA synthesis, processing, and translation. In this review we describe the role of PARP proteins and ADPRylation in all facets of this pathway. PARP-1 and its enzymatic activity are key regulators of rDNA transcription, which is a critical step in ribosome biogenesis. An emerging role of PARPs in alternative splicing of mRNAs, as well as direct ADPRylation of mRNAs, highlight the role of PARP members in RNA processing. Furthermore, PARP activity, stimulated by cellular stresses, such as viral infections and ER stress, leads to the regulation of mRNA stability and protein synthesis through posttranscriptional mechanisms. Dysregulation of PARP activity in these processes can promote disease states. Collectively, these results highlight the importance of PARP family members and ADPRylation in gene regulation, mRNA processing, and protein abundance. Future studies in these areas will yield new insights into the fundamental mechanisms and a broader utility for PARP-targeted therapeutic agents.
Assuntos
ADP-Ribosilação/fisiologia , Expressão Gênica/fisiologia , Poli(ADP-Ribose) Polimerases/metabolismo , Biossíntese de Proteínas/fisiologia , Proteostase/fisiologia , Animais , Humanos , Processamento de Proteína Pós-Traducional , RNA/metabolismoRESUMO
The nucleolus is the site of ribosome biogenesis, one of the most resource-intensive processes in eukaryotic cells. Accordingly, nucleolar morphology and activity are highly responsive to growth signaling and nucleolar insults which are collectively included in the actively evolving concept of nucleolar stress. Importantly, nucleolar alterations are a prominent feature of multiple human pathologies, including cancer and neurodegeneration, as well as being associated with aging. The past decades have seen numerous attempts to isolate compounds targeting different facets of nucleolar activity. We provide an overview of therapeutic opportunities for targeting nucleoli in different pathologies and currently available therapies.
Assuntos
Neoplasias , Ribossomos , Humanos , Nucléolo Celular/patologia , Neoplasias/tratamento farmacológico , Neoplasias/patologia , EnvelhecimentoRESUMO
Oncogenic RAS signaling reprograms gene expression through both transcriptional and post-transcriptional mechanisms. While transcriptional regulation downstream of RAS is relatively well characterized, how RAS post-transcriptionally modulates gene expression to promote malignancy remains largely unclear. Using quantitative RNA interactome capture analysis, we here reveal that oncogenic RAS signaling reshapes the RNA-bound proteomic landscape of pancreatic cancer cells, with a network of nuclear proteins centered around nucleolin displaying enhanced RNA-binding activity. We show that nucleolin is phosphorylated downstream of RAS, which increases its binding to pre-ribosomal RNA (rRNA), boosts rRNA production, and promotes ribosome biogenesis. This nucleolin-dependent enhancement of ribosome biogenesis is crucial for RAS-induced pancreatic cancer cell proliferation and can be targeted therapeutically to inhibit tumor growth. Our results reveal that oncogenic RAS signaling drives ribosome biogenesis by regulating the RNA-binding activity of nucleolin and highlight a crucial role for this mechanism in RAS-mediated tumorigenesis.