RESUMO
Proteins are manufactured by ribosomes-macromolecular complexes of protein and RNA molecules that are assembled within major nuclear compartments called nucleoli1,2. Existing models suggest that RNA polymerases I and III (Pol I and Pol III) are the only enzymes that directly mediate the expression of the ribosomal RNA (rRNA) components of ribosomes. Here we show, however, that RNA polymerase II (Pol II) inside human nucleoli operates near genes encoding rRNAs to drive their expression. Pol II, assisted by the neurodegeneration-associated enzyme senataxin, generates a shield comprising triplex nucleic acid structures known as R-loops at intergenic spacers flanking nucleolar rRNA genes. The shield prevents Pol I from producing sense intergenic noncoding RNAs (sincRNAs) that can disrupt nucleolar organization and rRNA expression. These disruptive sincRNAs can be unleashed by Pol II inhibition, senataxin loss, Ewing sarcoma or locus-associated R-loop repression through an experimental system involving the proteins RNaseH1, eGFP and dCas9 (which we refer to as 'red laser'). We reveal a nucleolar Pol-II-dependent mechanism that drives ribosome biogenesis, identify disease-associated disruption of nucleoli by noncoding RNAs, and establish locus-targeted R-loop modulation. Our findings revise theories of labour division between the major RNA polymerases, and identify nucleolar Pol II as a major factor in protein synthesis and nuclear organization, with potential implications for health and disease.
Assuntos
Nucléolo Celular/enzimologia , Nucléolo Celular/genética , DNA Ribossômico/genética , RNA Polimerase II/metabolismo , RNA não Traduzido/biossíntese , RNA não Traduzido/genética , Ribossomos/metabolismo , Proteína 9 Associada à CRISPR/genética , Proteína 9 Associada à CRISPR/metabolismo , Linhagem Celular Tumoral , Nucléolo Celular/fisiologia , DNA Helicases/metabolismo , DNA Intergênico/genética , Humanos , Enzimas Multifuncionais/metabolismo , Biossíntese de Proteínas , Estruturas R-Loop , RNA Helicases/metabolismo , RNA Polimerase I/antagonistas & inibidores , RNA Polimerase I/metabolismo , Ribonuclease H/metabolismo , Ribossomos/química , Ribossomos/genética , Sarcoma de Ewing/genética , Sarcoma de Ewing/patologiaRESUMO
Calorie restriction (CR) is a broadly effective environmental intervention that extends life by operating through numerous biological processes. Here, we discuss how non-coding RNA (ncRNA) molecules act as mediators and targets of lifespan-extending CR. We also highlight how these RNA molecules connect CR to its effects on genome stability, cell metabolism, programmed cell death, senescence, cancer, and neurodegeneration. We anticipate that an advanced understanding of the connections between CR and non-coding RNA will provide unique insights into aging mechanisms while pointing to novel approaches aimed at modulating aging and age-related diseases.
Assuntos
Envelhecimento/genética , Restrição Calórica , Instabilidade Genômica , RNA não Traduzido/genética , Animais , HumanosRESUMO
Dysfunction of key miRNA pathways regulating basic cellular processes is a common driver of many cancers. However, the biological roles and/or clinical applications of such pathways in Merkel cell carcinoma (MCC), a rare but lethal cutaneous neuroendocrine (NE) malignancy, have yet to be determined. Previous work has established that miR-375 is highly expressed in MCC tumors, but its biological role in MCC remains unknown. Herein, we show that elevated miR-375 expression is a specific feature of well-differentiated MCC cell lines that express NE markers. In contrast, miR-375 is strikingly down-regulated in highly aggressive, undifferentiated MCC cell lines. Enforced miR-375 expression in these cells induced NE differentiation, and opposed cancer cell viability, migration, invasion, and survival, pointing to tumor-suppressive roles for miR-375. Mechanistically, miR-375-driven phenotypes were caused by the direct post-transcriptional repression of multiple Notch pathway proteins (Notch2 and RBPJ) linked to cancer and regulation of cell fate. Thus, we detail a novel molecular axis linking tumor-suppressive miR-375 and Notch with NE differentiation and cancer cell behavior in MCC. Our findings identify miR-375 as a putative regulator of NE differentiation, provide insight into the cell of origin of MCC, and suggest that miR-375 silencing may promote aggressive cancer cell behavior through Notch disinhibition.