RESUMEN
Cyclins are central engines of cell cycle progression in conjunction with cyclin-dependent kinases (CDKs). Among the different cyclins controlling cell cycle progression, cyclin F does not partner with a CDK, but instead forms via its F-box domain an SCF (Skp1-Cul1-F-box)-type E3 ubiquitin ligase module. Although various substrates of cyclin F have been identified, the vulnerabilities of cells lacking cyclin F are not known. Thus, we assessed viability of cells lacking cyclin F upon challenging them with more than 180 different kinase inhibitors. The screen revealed a striking synthetic lethality between Chk1 inhibition and cyclin F loss. Chk1 inhibition in cells lacking cyclin F leads to DNA replication catastrophe. Replication catastrophe depends on accumulation of the transcription factor E2F1 in cyclin F-depleted cells. We find that SCF-cyclin F controls E2F1 ubiquitylation and degradation during the G2/M phase of the cell cycle and upon challenging cells with Chk1 inhibitors. Thus, Cyclin F restricts E2F1 activity during the cell cycle and upon checkpoint inhibition to prevent DNA replication stress. Our findings pave the way for patient selection in the clinical use of checkpoint inhibitors.
Asunto(s)
Quinasa 1 Reguladora del Ciclo Celular (Checkpoint 1)/antagonistas & inhibidores , Ciclinas/metabolismo , Factor de Transcripción E2F1/metabolismo , Inhibidores de Proteínas Quinasas/farmacología , Proteolisis , Proteínas Ligasas SKP Cullina F-box/metabolismo , Mutaciones Letales Sintéticas , Ciclo Celular/efectos de los fármacos , Quinasa 1 Reguladora del Ciclo Celular (Checkpoint 1)/genética , Ciclinas/genética , Replicación del ADN , Factor de Transcripción E2F1/genética , Células HeLa , Humanos , Fosforilación , Unión Proteica , Proteínas Ligasas SKP Cullina F-box/genética , UbiquitinaciónRESUMEN
BACKGROUND: Extraneural metastases (ENM) rarely occur in medulloblastoma (MBL) patients and only few cases of subcutaneous localizations have been described. ENM indicate an aggressive disease associated with a worse prognosis. The characterization of metastatic tumours might be useful to understand their pathogenesis and to identify the most appropriate therapeutic strategies. CASE PRESENTATION: We present the case of a child with Large Cell Anaplastic (LC/A) MBL, who developed multiple subcutaneous metastases in the scalp area after a ventriculo-peritoneal shunting procedure. The disease rapidly progressed and the child died despite chemotherapy and primary tumour surgical debulking.We molecularly classified the tumour as a group 3 MBL; in addition, we derived stem-like cells (SLC) from a metastatic lesion. Primary tumour, metastases and SLC were further analysed, particularly focusing on features linked to the cutaneous dissemination. Indeed, molecules involved in angiogenesis, cell invasion and epidermal growth factor signalling resulted highly expressed. CONCLUSIONS: The present report describes a very rare case of subcutaneous metastatic MBL. The tumour, metastases and SLC have been clinically, pathologically and molecularly characterized. Our case is an example of multidisciplinary approach aiming to characterize MBL aggressive behaviour.
Asunto(s)
Meduloblastoma/genética , Metástasis de la Neoplasia/genética , Células Madre Neoplásicas/patología , Preescolar , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , Meduloblastoma/diagnóstico , Meduloblastoma/patología , Metástasis de la Neoplasia/patología , PronósticoRESUMEN
Orderly progressions of events in the cell division cycle are necessary to ensure the replication of DNA and cell division. Checkpoint systems allow the accurate execution of each cell-cycle phase. The precise regulation of the levels of cyclin proteins is fundamental to coordinate cell division with checkpoints, avoiding genome instability. Cyclin F has important functions in regulating the cell cycle during the G2 checkpoint; however, the mechanisms underlying the regulation of cyclin F are poorly understood. Here, we observe that cyclin F is regulated by proteolysis through ß-TrCP. ß-TrCP recognizes cyclin F through a non-canonical degron site (TSGXXS) after its phosphorylation by casein kinase II. The degradation of cyclin F mediated by ß-TrCP occurs at the G2/M transition. This event is required to promote mitotic progression and favors the activation of a transcriptional program required for mitosis.
Asunto(s)
Quinasa de la Caseína II/metabolismo , Ciclinas/metabolismo , Mitosis , Proteolisis , Proteínas con Repetición de beta-Transducina/metabolismo , Ciclinas/química , Células HEK293 , Células HeLa , HumanosRESUMEN
Suppressor of Fused (SuFu), a tumour suppressor mutated in medulloblastoma, is a central player of Hh signalling, a pathway crucial for development and deregulated in cancer. Although the control of Gli transcription factors by SuFu is critical in Hh signalling, our understanding of the mechanism regulating this key event remains limited. Here, we show that the Itch/ß-arrestin2 complex binds SuFu and induces its Lys63-linked polyubiquitylation without affecting its stability. This process increases the association of SuFu with Gli3, promoting the conversion of Gli3 into a repressor, which keeps Hh signalling off. Activation of Hh signalling antagonises the Itch-dependent polyubiquitylation of SuFu. Notably, different SuFu mutations occurring in medulloblastoma patients are insensitive to Itch activity, thus leading to deregulated Hh signalling and enhancing medulloblastoma cell growth. Our findings uncover mechanisms controlling the tumour suppressive functions of SuFu and reveal that their alterations are implicated in medulloblastoma tumorigenesis.
Asunto(s)
Proteínas Hedgehog/metabolismo , Meduloblastoma/metabolismo , Proteínas Represoras/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Arrestina beta 2/metabolismo , Secuencias de Aminoácidos , Animales , Carcinogénesis , Femenino , Proteínas Hedgehog/genética , Humanos , Meduloblastoma/enzimología , Meduloblastoma/genética , Meduloblastoma/patología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos NOD , Ratones Noqueados , Ratones SCID , Proteínas Represoras/química , Proteínas Represoras/genética , Transducción de Señal , Ubiquitina-Proteína Ligasas/genética , Ubiquitinación , Arrestina beta 2/genéticaRESUMEN
2-Substituted 3-aroylquinolin-4(1H)-ones, prepared through a palladium-catalyzed carbonylative cyclization of N-(2-iodoaryl)enaminones, proved to inhibit efficiently the Hedgehog pathway through direct antagonism of the wild-type and drug-resistant form of the Smoothened receptor. Notably, these compounds repressed the Hh-dependent growth events and the proliferation of tumor cells with aberrant activation of the Hh pathway, which plays a crucial role in development and tumorigenesis.