RESUMEN
Transcription is extremely important for cellular processes but can be hindered by RNA polymerase II (RNAPII) pausing and stalling. Cockayne syndrome protein B (CSB) promotes the progression of paused RNAPII or initiates transcription-coupled nucleotide excision repair (TC-NER) to remove stalled RNAPII. However, the specific mechanism by which CSB initiates TC-NER upon damage remains unclear. In this study, we identified the indispensable role of the ARK2N-CK2 complex in the CSB-mediated initiation of TC-NER. The ARK2N-CK2 complex is recruited to damage sites through CSB and then phosphorylates CSB. Phosphorylation of CSB enhances its binding to stalled RNAPII, prolonging the association of CSB with chromatin and promoting CSA-mediated ubiquitination of stalled RNAPII. Consistent with this finding, Ark2n-/- mice exhibit a phenotype resembling Cockayne syndrome. These findings shed light on the pivotal role of the ARK2N-CK2 complex in governing the fate of RNAPII through CSB, bridging a critical gap necessary for initiating TC-NER.
Asunto(s)
Síndrome de Cockayne , ADN Helicasas , Enzimas Reparadoras del ADN , Reparación del ADN , Proteínas de Unión a Poli-ADP-Ribosa , ARN Polimerasa II , Enzimas Reparadoras del ADN/metabolismo , Enzimas Reparadoras del ADN/genética , ARN Polimerasa II/metabolismo , ARN Polimerasa II/genética , Proteínas de Unión a Poli-ADP-Ribosa/metabolismo , Proteínas de Unión a Poli-ADP-Ribosa/genética , Humanos , Animales , Ratones , ADN Helicasas/metabolismo , ADN Helicasas/genética , Síndrome de Cockayne/genética , Síndrome de Cockayne/metabolismo , Transcripción Genética , Fosforilación , Quinasa de la Caseína II/metabolismo , Quinasa de la Caseína II/genética , Ratones Noqueados , Daño del ADN , ATPasas Asociadas con Actividades Celulares Diversas/metabolismo , ATPasas Asociadas con Actividades Celulares Diversas/genética , Cromatina/metabolismo , Ubiquitinación , Reparación por EscisiónRESUMEN
Gliomas are the most common primary tumors of the central nervous system, with approximately half of patients presenting with the most aggressive form of glioblastoma. Although several molecular markers for glioma have been identified, they are not sufficient to predict the prognosis due to the extensive genetic heterogeneity within glioma. Our study reveals that the ratio of IMPDH1 to IMPDH2 expression levels serves as a molecular indicator for glioma treatment prognosis. Patients with a higher IMPDH1/IMPDH2 ratio exhibit a worse prognosis, while those with a lower ratio display a more favorable prognosis. We further demonstrate that IMPDH1 plays a crucial role in maintaining cellular GTP/GDP levels following DNA damage compared to IMPDH2. In the absence of IMPDH1, cells experience an imbalance in the GTP/GDP ratio, impairing DNA damage repair capabilities and rendering them more sensitive to TMZ. This study not only introduces a novel prognostic indicator for glioma clinical diagnosis but also offers innovative insights for precise and stratified glioma treatment.
Asunto(s)
Glioma , IMP Deshidrogenasa , Temozolomida , Humanos , Glioma/tratamiento farmacológico , Glioma/genética , Glioma/patología , Glioma/mortalidad , IMP Deshidrogenasa/genética , IMP Deshidrogenasa/metabolismo , Temozolomida/uso terapéutico , Temozolomida/farmacología , Pronóstico , Neoplasias Encefálicas/tratamiento farmacológico , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patología , Línea Celular Tumoral , Daño del ADN/efectos de los fármacos , Femenino , Masculino , Guanosina Trifosfato/metabolismoRESUMEN
Mutations in the BRCA2 tumor suppressor gene have been associated with an increased risk of developing prostate cancer. One of the paradoxes concerning BRCA2 is the fact that its inactivation affects genetic stability and is deleterious for cellular and organismal survival, while BRCA2-mutated cancer cells adapt to this detriment and malignantly proliferate. Therapeutic strategies for tumors arising from BRCA2 mutations may be discovered by understanding these adaptive mechanisms. In this study, we conducted forward genetic synthetic viability screenings in Caenorhabditis elegans brc-2 (Cebrc-2) mutants and found that Ceubxn-2 inactivation rescued the viability of Cebrc-2 mutants. Moreover, loss of NSFL1C, the mammalian ortholog of CeUBXN-2, suppressed the spindle assembly checkpoint (SAC) activation and promoted the survival of BRCA2-deficient cells. Mechanistically, NSFL1C recruited USP9X to inhibit the polyubiquitination of AURKB and reduce the removal of AURKB from the centromeres by VCP, which is essential for SAC activation. SAC inactivation is common in BRCA2-deficient prostate cancer patients, but PP2A inhibitors could reactivate the SAC and achieve BRCA2-deficient prostate tumor synthetic lethality. Our research reveals the survival adaptation mechanism of BRCA2-deficient prostate tumor cells and provides different angles for exploring synthetic lethal inhibitors in addition to targeting DNA damage repair pathways.
Asunto(s)
Neoplasias de la Próstata , Mutaciones Letales Sintéticas , Animales , Humanos , Masculino , Proteína BRCA2 , Caenorhabditis elegans/genética , Puntos de Control de la Fase M del Ciclo Celular/genética , Mamíferos/metabolismo , Mutación , Neoplasias de la Próstata/tratamiento farmacológico , Neoplasias de la Próstata/genética , Ubiquitina Tiolesterasa/genética , Proteína Fosfatasa 2/metabolismoRESUMEN
The replication-stress response is essential to ensure the faithful transmission of genetic information to daughter cells. Although several stress-resolution pathways have been identified to deal with replication stress, the precise regulatory mechanisms for replication fork stability are not fully understood. Our study identified Methyl-CpG Binding Domain 1 (MBD1) as essential for the maintaining genomic stability and protecting stalled replication forks in mammalian cells. Depletion of MBD1 increases DNA lesions and sensitivity to replication stress. Mechanistically, we found that loss of MBD1 leads to the dissociation of Poly(ADP-ribose) polymerase 1 (PARP1) from the replication fork, potentially accelerating fork progression and resulting in higher levels of transcription-replication conflicts (T-R conflicts). Using a proximity ligation assay combined with 5-ethynyl-2'-deoxyuridine, we revealed that the MBD1 and PARP1 proteins were recruited to stalled forks under hydroxyurea (HU) treatment. In addition, our study showed that the level of R-loops also increased in MBD1-delated cells. Without MBD1, stalled replication forks resulting from T-R conflicts were primarily degraded by the DNA2 nuclease. Our findings shed light on a new aspect of MBD1 in maintaining genome stability and providing insights into the mechanisms underlying replication stress response.
Asunto(s)
Daño del ADN , Replicación del ADN , Humanos , Animales , Inestabilidad Genómica , Mamíferos/metabolismo , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Factores de Transcripción , Poli(ADP-Ribosa) Polimerasa-1/genética , Poli(ADP-Ribosa) Polimerasa-1/metabolismoRESUMEN
BRCA1 expression is highly regulated to prevent genomic instability and tumorigenesis. Dysregulation of BRCA1 expression correlates closely with sporadic basal-like breast cancer and ovarian cancer. The most significant characteristic of BRCA1 regulation is periodic expression fluctuation throughout the cell cycle, which is important for the orderly progression of different DNA repair pathways throughout the various cell cycle phases and for further genomic stability. However, the underlying mechanism driving this phenomenon is poorly understood. Here, we demonstrate that RBM10-mediated RNA alternative splicing coupled to nonsense-mediated mRNA decay (AS-NMD), rather than transcription, determines the periodic fluctuations in G1/S-phase BRCA1 expression. Furthermore, AS-NMD broadly regulates the expression of period genes, such as DNA replication-related genes, in an uneconomical but more rapid manner. In summary, we identified an unexpected posttranscriptional mechanism distinct from canonical processes that mediates the rapid regulation of BRCA1 as well as other period gene expression during the G1/S-phase transition and provided insights into potential targets for cancer therapy.
Asunto(s)
Neoplasias de la Mama , Degradación de ARNm Mediada por Codón sin Sentido , Humanos , Femenino , Empalme Alternativo , Empalme del ARN , Neoplasias de la Mama/genética , Inestabilidad Genómica , Proteína BRCA1/genética , Proteínas de Unión al ARN/genéticaRESUMEN
Lysine crotonylation (Kcr) is a newly identified histone modification that is associated with active transcription in mammalian cells. Here we report that the chromodomain Y-like transcription corepressor CDYL negatively regulates histone Kcr by acting as a crotonyl-CoA hydratase to convert crotonyl-CoA to ß-hydroxybutyryl-CoA. We showed that the negative regulation of histone Kcr by CDYL is intrinsically linked to its transcription repression activity and functionally implemented in the reactivation of sex chromosome-linked genes in round spermatids and genome-wide histone replacement in elongating spermatids. Significantly, Cdyl transgenic mice manifest dysregulation of histone Kcr and reduction of male fertility with a decreased epididymal sperm count and sperm cell motility. Our study uncovers a biochemical pathway in the regulation of histone Kcr and implicates CDYL-regulated histone Kcr in spermatogenesis, adding to the understanding of the physiology of male reproduction and the mechanism of the spermatogenic failure in AZFc (Azoospermia Factor c)-deleted infertile men.