RESUMO
Clear cell renal cell carcinoma (ccRCC) is the most common kidney cancer in the adult population. Late diagnosis, resistance to therapeutics and recurrence of metastatic lesions account for the highest mortality rate among kidney cancer patients. Identifying novel biomarkers for early cancer detection and elucidating the mechanisms underlying ccRCC will provide clues to treat this aggressive malignant tumor. Here, we report that the ubiquitin ligase praja2 forms a complex with-and ubiquitylates the AP2 adapter complex, contributing to receptor endocytosis and clearance. In human RCC tissues and cells, downregulation of praja2 by oncogenic miRNAs (oncomiRs) and the proteasome markedly impairs endocytosis and clearance of the epidermal growth factor receptor (EGFR), and amplifies downstream mitogenic and proliferative signaling. Restoring praja2 levels in RCC cells downregulates EGFR, rewires cancer cell metabolism and ultimately inhibits tumor cell growth and metastasis. Accordingly, genetic ablation of praja2 in mice upregulates RTKs (i.e. EGFR and VEGFR) and induces epithelial and vascular alterations in the kidney tissue.In summary, our findings identify a regulatory loop between oncomiRs and the ubiquitin proteasome system that finely controls RTKs endocytosis and clearance, positively impacting mitogenic signaling and kidney cancer growth.
Assuntos
Carcinoma de Células Renais , Neoplasias Renais , Adulto , Animais , Humanos , Camundongos , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/patologia , Regulação para Baixo , Endocitose , Receptores ErbB/genética , Receptores ErbB/metabolismo , Neoplasias Renais/genética , Neoplasias Renais/patologia , Complexo de Endopeptidases do Proteassoma/metabolismo , Receptores Proteína Tirosina Quinases/genética , Ubiquitina/metabolismoRESUMO
Bardet-Biedl syndrome (BBS) is a ciliopathy characterized by retinal degeneration, obesity, renal abnormalities, postaxial polydactyly, and developmental defects. Genes mutated in BBS encode for components and regulators of the BBSome, an octameric complex that controls the trafficking of cargos and receptors within the primary cilium. Although both structure and function of the BBSome have been extensively studied, the impact of ubiquitin signaling on BBSome is largely unknown. We identify the E3 ubiquitin ligase PJA2 as a novel resident of the ciliary compartment and regulator of the BBSome. Upon GPCR-cAMP stimulation, PJA2 ubiquitylates BBSome subunits. We demonstrate that ubiquitylation of BBS1 at lysine 143 increases the stability of the BBSome and promotes its binding to BBS3, an Arf-like GTPase protein controlling the targeting of the BBSome to the ciliary membrane. Downregulation of PJA2 or expression of a ubiquitylation-defective BBS1 mutant (BBS1K143R ) affects the trafficking of G-protein-coupled receptors (GPCRs) and Shh-dependent gene transcription. Expression of BBS1K143R in vivo impairs cilium formation, embryonic development, and photoreceptors' morphogenesis, thus recapitulating the BBS phenotype in the medaka fish model.
Assuntos
Síndrome de Bardet-Biedl , Cílios , Animais , Cílios/metabolismo , Transporte Proteico , Transdução de Sinais , Síndrome de Bardet-Biedl/genética , Receptores Acoplados a Proteínas G/genética , UbiquitinaçãoRESUMO
Cellular homeostasis is tightly connected to the broad variety of mitochondrial functions. To stay healthy, cells need a constant supply of nutrients, energy production and antioxidants defenses, undergoing programmed death when a serious, irreversible damage occurs. The key element of a functional integration of all these processes is the correct crosstalk between cell signaling and mitochondrial activities. Once this crosstalk is interrupted, the cell is not able to communicate its needs to mitochondria, resulting in oxidative stress and development of pathological conditions. Conversely, dysfunctional mitochondria may affect cell viability, even in the presence of nutrients supply and energy production, indicating the existence of feed-back control mechanisms between mitochondria and other cellular compartments. The ubiquitin proteasome system (UPS) is a multi-step biochemical pathway that, through the conjugation of ubiquitin moieties to specific protein substrates, controls cellular proteostasis and signaling, removing damaged or aged proteins that might otherwise accumulate and affect cell viability. In response to specific needs or changed extracellular microenvironment, the UPS modulates the turnover of mitochondrial proteins, thus influencing the organelle shape, dynamics and function. Alterations of the dynamic and reciprocal regulation between mitochondria and UPS underpin genetic and proliferative disorders. This review focuses on the mitochondrial metabolism and activities supervised by UPS and examines how deregulation of this control mechanism results in proliferative disorders and cancer.
Assuntos
Neoplasias , Ubiquitina , Humanos , Idoso , Ubiquitina/metabolismo , Mitocôndrias/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Transdução de Sinais , Proteínas Mitocondriais/metabolismo , Neoplasias/metabolismo , Microambiente TumoralRESUMO
Glioblastoma multiforme (GBM) is the most frequent and aggressive form of primary brain tumor in the adult population; its high recurrence rate and resistance to current therapeutics urgently demand a better therapy. Regulation of protein stability by the ubiquitin proteasome system (UPS) represents an important control mechanism of cell growth. UPS deregulation is mechanistically linked to the development and progression of a variety of human cancers, including GBM. Thus, the UPS represents a potentially valuable target for GBM treatment. Using an integrated approach that includes proteomics, transcriptomics and metabolic profiling, we identify praja2, a RING E3 ubiquitin ligase, as the key component of a signaling network that regulates GBM cell growth and metabolism. Praja2 is preferentially expressed in primary GBM lesions expressing the wild-type isocitrate dehydrogenase 1 gene (IDH1). Mechanistically, we found that praja2 ubiquitylates and degrades the kinase suppressor of Ras 2 (KSR2). As a consequence, praja2 restrains the activity of downstream AMP-dependent protein kinase in GBM cells and attenuates the oxidative metabolism. Delivery in the brain of siRNA targeting praja2 by transferrin-targeted self-assembling nanoparticles (SANPs) prevented KSR2 degradation and inhibited GBM growth, reducing the size of the tumor and prolonging the survival rate of treated mice. These data identify praja2 as an essential regulator of cancer cell metabolism, and as a potential therapeutic target to suppress GBM growth.
Assuntos
Neoplasias Encefálicas , Glioblastoma , Adulto , Animais , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Glioblastoma/metabolismo , Humanos , Camundongos , Complexo de Endopeptidases do Proteassoma/metabolismo , Transdução de Sinais , UbiquitinaRESUMO
Primary cilia are microtubule-based, non-motile sensory organelles present in most types of growth-arrested eukaryotic cells. They are transduction hubs that receive and transmit external signals to the cells in order to control growth, differentiation and development. Mutations of genes involved in the formation, maintenance or disassembly of ciliary structures cause a wide array of developmental genetic disorders, also known as ciliopathies. The primary cilium is formed during G1 in the cell cycle and disassembles at the G2/M transition. Following the completion of the cell division, the cilium reassembles in G1. This cycle is finely regulated at multiple levels. The ubiquitin-proteasome system (UPS) and the autophagy machinery, two main protein degradative systems in cells, play a fundamental role in cilium dynamics. Evidence indicate that UPS, autophagy and signaling pathways may act in synergy to control the ciliary homeostasis. However, the mechanisms involved and the links between these regulatory systems and cilium biogenesis, dynamics and signaling are not well defined yet. Here, we discuss the reciprocal regulation of signaling pathways and proteolytic machineries in the control of the assembly and disassembly of the primary cilium, and the impact of the derangement of these regulatory networks in human ciliopathies.
RESUMO
The primary cilium is a microtubule-based sensory organelle that dynamically links signalling pathways to cell differentiation, growth, and development. Genetic defects of primary cilia are responsible for genetic disorders known as ciliopathies. Orofacial digital type I syndrome (OFDI) is an X-linked congenital ciliopathy caused by mutations in the OFD1 gene and characterized by malformations of the face, oral cavity, digits and, in the majority of cases, polycystic kidney disease. OFD1 plays a key role in cilium biogenesis. However, the impact of signalling pathways and the role of the ubiquitin-proteasome system (UPS) in the control of OFD1 stability remain unknown. Here, we identify a novel complex assembled at centrosomes by TBC1D31, including the E3 ubiquitin ligase praja2, protein kinase A (PKA), and OFD1. We show that TBC1D31 is essential for ciliogenesis. Mechanistically, upon G-protein-coupled receptor (GPCR)-cAMP stimulation, PKA phosphorylates OFD1 at ser735, thus promoting OFD1 proteolysis through the praja2-UPS circuitry. This pathway is essential for ciliogenesis. In addition, a non-phosphorylatable OFD1 mutant dramatically affects cilium morphology and dynamics. Consistent with a role of the TBC1D31/praja2/OFD1 axis in ciliogenesis, alteration of this molecular network impairs ciliogenesis in vivo in Medaka fish, resulting in developmental defects. Our findings reveal a multifunctional transduction unit at the centrosome that links GPCR signalling to ubiquitylation and proteolysis of the ciliopathy protein OFD1, with important implications on cilium biology and development. Derangement of this control mechanism may underpin human genetic disorders.