RESUMO
YAP and TAZ (YAP/TAZ), two major effectors of the Hippo signaling pathway, are frequently activated in human cancers. The activity of YAP/TAZ is strictly repressed upon phosphorylation by LATS1/2 tumor suppressors. However, it is unclear how LATS1/2 are precisely regulated by upstream factors such as Hippo kinases MST1/2. Here, we show that WWC proteins (WWC1/2/3) directly interact with LATS1/2 and SAV1, and SAV1, in turn, brings in MST1/2 to phosphorylate and activate LATS1/2. Hence, WWC1/2/3 play an organizer role in a signaling module that mediates LATS1/2 activation by MST1/2. Moreover, we have defined a minimum protein interaction interface on WWC1/2/3 that is sufficient to activate LATS1/2 in a robust and specific manner. The corresponding minigene, dubbed as SuperHippo, can effectively suppress tumorigenesis in multiple tumor models. Our study has uncovered a molecular mechanism underlying LATS1/2 regulation and provides a strategy for treating diverse malignancies related to Hippo pathway dysregulation.
Assuntos
Proteínas Serina-Treonina Quinases , Transdução de Sinais , Carcinogênese , Via de Sinalização Hippo , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Fosforilação , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais/fisiologia , Proteínas Supressoras de Tumor/metabolismoRESUMO
The Hippo pathway is a central regulator of organ size and tumorigenesis and is commonly depicted as a kinase cascade, with an increasing number of regulatory and adaptor proteins linked to its regulation over recent years. Here, we propose that two Hippo signaling modules, MST1/2-SAV1-WWC1-3 (HPO1) and MAP4K1-7-NF2 (HPO2), together regulate the activity of LATS1/2 kinases and YAP/TAZ transcriptional co-activators. In mouse livers, the genetic inactivation of either HPO1 or HPO2 module results in partial activation of YAP/TAZ, bile duct hyperplasia, and hepatocellular carcinoma (HCC). On the contrary, inactivation of both HPO1 and HPO2 modules results in full activation of YAP/TAZ, rapid development of intrahepatic cholangiocarcinoma (iCCA), and early lethality. Interestingly, HPO1 has a predominant role in regulating organ size. HPO1 inactivation causes a homogenous YAP/TAZ activation and cell proliferation across the whole liver, resulting in a proportional and rapid increase in liver size. Thus, this study has reconstructed the order of the Hippo signaling network and suggests that LATS1/2 and YAP/TAZ activities are finetuned by HPO1 and HPO2 modules to cause different cell fates, organ size changes, and tumorigenesis trajectories.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Camundongos , Animais , Via de Sinalização Hippo , Transdução de Sinais , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Carcinoma Hepatocelular/genética , Proteínas de Sinalização YAP , Neoplasias Hepáticas/genética , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Carcinogênese/genética , Transformação Celular Neoplásica , Fosfoproteínas/genética , Fosfoproteínas/metabolismoRESUMO
The scaffolding protein angiomotin (AMOT) is indispensable for vertebrate embryonic angiogenesis. Here, we report that AMOT undergoes cleavage in the presence of lysophosphatidic acid (LPA), a lipid growth factor also involved in angiogenesis. AMOT cleavage is mediated by aspartic protease DNA damage-inducible 1 homolog 2 (DDI2), and the process is tightly regulated by a signaling axis including neurofibromin 2 (NF2), tankyrase 1/2 (TNKS1/2), and RING finger protein 146 (RNF146), which induce AMOT membrane localization, poly ADP ribosylation, and ubiquitination, respectively. In both zebrafish and mice, the genetic inactivation of AMOT cleavage regulators leads to defective angiogenesis, and the phenotype is rescued by the overexpression of AMOT-CT, a C-terminal AMOT cleavage product. In either physiological or pathological angiogenesis, AMOT-CT is required for vascular expansion, whereas uncleavable AMOT represses this process. Thus, our work uncovers a signaling pathway that regulates angiogenesis by modulating a cleavage-dependent activation of AMOT.
Assuntos
Angiomotinas , Peixe-Zebra , Animais , Camundongos , Peixe-Zebra/metabolismo , Proteínas dos Microfilamentos/metabolismo , Peptídeo Hidrolases , Peptídeos e Proteínas de Sinalização Intercelular/genéticaRESUMO
Type I interferon (IFN)-induced Janus kinase (JAK)-signal transducer and activator of transcription (STAT) signaling drives the expression of IFN-stimulated genes (ISGs) to mediate antiviral response. The strength and duration of JAK-STAT signaling are tightly regulated to ensure effective antiviral defense while avoiding pathological inflammation and autoimmunity. Here, we report that cTAZ, an isoform of the Hippo pathway effector TAZ, is transcribed by an alternative promoter. Although majority of C-terminal sequences of TAZ is retained, cTAZ is not regulated by the Hippo signaling and does not mediate its growth-inhibitory functions. Instead, cTAZ negatively regulates JAK-STAT signaling by inhibiting STAT1/2 nuclear localization and ISG expression, and its expression is induced by type I IFN Thus, cTAZ functions as a modulator of JAK-STAT signaling and may play a role in fine-tuning cellular antiviral response.
Assuntos
Janus Quinases/metabolismo , Regiões Promotoras Genéticas , Fatores de Transcrição STAT/metabolismo , Transdução de Sinais , Transativadores/genética , Transcrição Gênica , Animais , Perfilação da Expressão Gênica , Via de Sinalização Hippo , Humanos , Camundongos , Modelos Biológicos , Fosforilação , Ligação Proteica , Multimerização Proteica , Proteínas Serina-Treonina Quinases/metabolismo , Transporte Proteico , Isoformas de RNA , Fatores de Transcrição STAT/química , Proteínas com Motivo de Ligação a PDZ com Coativador TranscricionalRESUMO
Yes-associated protein (YAP) is a key downstream effector of the highly conserved Hippo signaling pathway, which regulates organ size, regeneration and tumorigenesis. Known classically to function as a transcriptional co-activator, YAP interacts with TEA domain transcription factors (TEAD1-4) to induce expression of target genes. However, a number of genes are repressed upon YAP activation, suggesting a transcriptional repressor role of YAP. Here, we report that TP73 is a direct target gene of YAP, and its transcription is repressed by YAP in a TEAD-independent manner. On the other hand, WW domains of YAP are indispensable for the regulation of TP73 expression, which may recruit YAP to TP73 gene though interaction with ZEB1 and/or RUNX2, two transcriptional repressors. Moreover, YAP-mediated repression of TP73 promotes cancer cell survival in the presence of chemotherapeutic agents, suggesting YAP-TP73 signaling as a mechanism for cancer cell resistance to chemotherapies.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais , Fatores de Transcrição/metabolismo , Transcrição Gênica , Proteína Tumoral p73/genética , Proteínas Adaptadoras de Transdução de Sinal/química , Sequência de Bases , Linhagem Celular Tumoral , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Resistencia a Medicamentos Antineoplásicos , Via de Sinalização Hippo , Humanos , Domínios Proteicos , Proteínas Repressoras/metabolismo , Fatores de Transcrição/química , Proteína Tumoral p73/metabolismo , Proteínas de Sinalização YAP , Homeobox 1 de Ligação a E-box em Dedo de Zinco/metabolismoRESUMO
WWC1 regulates episodic learning and memory, and genetic nucleotide polymorphism of WWC1 is associated with neurodegenerative diseases such as Alzheimer's disease. However, the molecular mechanism through which WWC1 regulates neuronal function has not been fully elucidated. Here, we show that WWC1 and its paralogs (WWC2/3) bind directly to angiomotin (AMOT) family proteins (Motins), and recruit USP9X to deubiquitinate and stabilize Motins. Deletion of WWC genes in different cell types leads to reduced protein levels of Motins. In mice, neuron-specific deletion of Wwc1 and Wwc2 results in reduced expression of Motins and lower density of dendritic spines in the cortex and hippocampus, in association with impaired cognitive functions such as memory and learning. Interestingly, ectopic expression of AMOT partially rescues the neuronal phenotypes associated with Wwc1/2 deletion. Thus, WWC proteins modulate spinogenesis and cognition, at least in part, by regulating the protein stability of Motins.
Assuntos
Angiomotinas , Aprendizagem , Camundongos , Animais , Hipocampo/fisiologia , Neurônios , Proteínas dos Microfilamentos , CogniçãoRESUMO
Germline alterations of the NF2 gene cause neurofibromatosis type 2, a syndrome manifested with benign tumors, and Nf2 deletion in mice also results in slow tumorigenesis. As a regulator of the Hippo signaling pathway, NF2 induces LATS1/2 kinases and consequently represses YAP/TAZ. YAP/TAZ oncoproteins are also inhibited by motin family proteins (Motins). Here, we show that the Hippo signaling is fine-tuned by Motins in a NF2-dependent manner, in which NF2 recruits E3 ligase RNF146 to facilitate ubiquitination and subsequent degradation of Motins. In the absence of NF2, Motins robustly accumulate to restrict full activation of YAP/TAZ and prevent rapid tumorigenesis. Hence, NF2 deficiency not only activates YAP/TAZ by inhibiting LATS1/2 but also stabilizes Motins to keep YAP/TAZ activity in check. The upregulation of Motins upon NF2 deletion serves as a strategy for avoiding uncontrolled perturbation of the Hippo signaling and may contribute to the benign nature of most NF2-mutated tumors.
Assuntos
Carcinogênese/genética , Transformação Celular Neoplásica/genética , Genes da Neurofibromatose 2 , Via de Sinalização Hippo/fisiologia , Proteínas de Sinalização YAP/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Proteínas de Ciclo Celular/metabolismo , Via de Sinalização Hippo/genética , Humanos , Camundongos , Fosfoproteínas/metabolismo , Transativadores/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
Recombinant adeno-associated virus (rAAV) vectors selected from capsid libraries present enormous advantages in high selectivity of tissue tropism and their potential use in human gene therapy applications. For example, rAAV-LK03, was used in a gene therapy trial for hemophilia A (ClinicalTrials.gov: NCT03003533). However, high doses in patients resulted in severe adverse events and subsequent loss of factor VIII (FVIII) expression. Thus, additional strategies are needed to enhance the transduction efficiency of capsid library-derived rAAV vectors such that improved clinical efficacy can be achieved at low vector doses. In this study, we characterized two commonly used library-derived rAAV vectors, rAAV-DJ and rAAV-LK03. It was concluded that rAAV-DJ shared similar transport pathways (e.g., cell surface binding, endocytosis-dependent internalization, and cytoplasmic trafficking) with rAAV serotype 2, while rAAV-LK03 and rAAV serotype 3 shared similar transport pathways. We then performed site-directed mutagenesis of surface-exposed tyrosine (Y), serine (S), aspartic acid (D), and tryptophan (W) residues on rAAV-DJ and rAAV-LK03 capsids. Our results demonstrated that rAAV-DJ-S269T and rAAV-LK03-Y705+731F variants had significantly enhanced transduction efficiency compared to wild-type counterparts. Our studies suggest that the strategy of site-directed mutagenesis should be applicable to other non-natural AAV variants for their optimal use in human gene therapy.