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1.
Mol Cell ; 84(2): 277-292.e9, 2024 Jan 18.
Artigo em Inglês | MEDLINE | ID: mdl-38183983

RESUMO

iRhoms are pseudoprotease members of the rhomboid-like superfamily and are cardinal regulators of inflammatory and growth factor signaling; they function primarily by recognizing transmembrane domains of their clients. Here, we report a mechanistically distinct nuclear function of iRhoms, showing that both human and mouse iRhom2 are non-canonical substrates of signal peptidase complex (SPC), the protease that removes signal peptides from secreted proteins. Cleavage of iRhom2 generates an N-terminal fragment that enters the nucleus and modifies the transcriptome, in part by binding C-terminal binding proteins (CtBPs). The biological significance of nuclear iRhom2 is indicated by elevated levels in skin biopsies of patients with psoriasis, tylosis with oesophageal cancer (TOC), and non-epidermolytic palmoplantar keratoderma (NEPPK); increased iRhom2 cleavage in a keratinocyte model of psoriasis; and nuclear iRhom2 promoting proliferation of keratinocytes. Overall, this work identifies an unexpected SPC-dependent ER-to-nucleus signaling pathway and demonstrates that iRhoms can mediate nuclear signaling.


Assuntos
Psoríase , Transdução de Sinais , Animais , Humanos , Camundongos , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Psoríase/genética , Serina Endopeptidases/genética , Serina Endopeptidases/metabolismo
2.
EMBO J ; 39(17): e104415, 2020 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-32715522

RESUMO

Animals have evolved multiple mechanisms to protect themselves from the cumulative effects of age-related cellular damage. Here, we reveal an unexpected link between the TNF (tumour necrosis factor) inflammatory pathway, triggered by the metalloprotease ADAM17/TACE, and a lipid droplet (LD)-mediated mechanism of protecting retinal cells from age-related degeneration. Loss of ADAM17, TNF and the TNF receptor Grindelwald in pigmented glial cells of the Drosophila retina leads to age-related degeneration of both glia and neurons, preceded by an abnormal accumulation of glial LDs. We show that the glial LDs initially buffer the cells against damage caused by glial and neuronally generated reactive oxygen species (ROS), but that in later life the LDs dissipate, leading to the release of toxic peroxidated lipids. Finally, we demonstrate the existence of a conserved pathway in human iPS-derived microglia-like cells, which are central players in neurodegeneration. Overall, we have discovered a pathway mediated by TNF signalling acting not as a trigger of inflammation, but as a cytoprotective factor in the retina.


Assuntos
Proteína ADAM17/metabolismo , Proteínas de Drosophila/metabolismo , Gotículas Lipídicas/metabolismo , Proteínas de Membrana/metabolismo , Neuroglia/metabolismo , Retina/metabolismo , Transdução de Sinais , Fator de Necrose Tumoral alfa/metabolismo , Proteína ADAM17/genética , Animais , Proteínas de Drosophila/genética , Drosophila melanogaster , Humanos , Gotículas Lipídicas/patologia , Proteínas de Membrana/genética , Neuroglia/patologia , Espécies Reativas de Oxigênio/metabolismo , Retina/patologia , Fator de Necrose Tumoral alfa/genética
3.
Nat Cell Biol ; 9(6): 698-705, 2007 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-17496887

RESUMO

The cause or consequence of overexpression of p73 (refs 1, 2), the structural and functional homologue of the tumour-suppressor gene product p53 (refs 3, 4), in human cancers is poorly understood. Here, we report a role for p73 in supporting cellular growth through the upregulation of AP-1 transcriptional activity. p73 suppresses growth when overexpressed alone, but synergises with the proto-oncogene c-Jun to promote cellular survival. Conversely, silencing of p73 expression compromises cellular proliferation. Molecular analysis revealed that expression of the AP-1 target-gene product cyclinD1 (ref. 5) is reduced concomitant with p73, but not p53, silencing. Moreover, cyclinD1 was induced by p73 expression in a c-Jun-dependent manner, and was required for p73-mediated cell survival. Furthermore, c-Jun-dependent AP-1 transcriptional activity was augmented by p73 and, consistently, induction of endogenous AP-1 target genes was compromised in the absence of p73. Chromatin immunoprecipitation and electrophoretic mobility shift analysis indicated that p73 enhanced the binding of phosphorylated c-Jun and Fra-1, another AP-1 family member, to AP-1 consensus DNA sequences, by regulating c-Jun phosphorylation and Fra-1 expression. Collectively, our data demonstrates a novel and unexpected role of p73 in augmenting AP-1 transcriptional activity through which it supports cellular growth.


Assuntos
Proliferação de Células , Transformação Celular Neoplásica/metabolismo , Proteínas de Ligação a DNA/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Proto-Oncogênicas c-jun/metabolismo , Fator de Transcrição AP-1/metabolismo , Ativação Transcricional/genética , Proteínas Supressoras de Tumor/metabolismo , Animais , Células COS , Linhagem Celular Tumoral , Sobrevivência Celular/genética , Transformação Celular Neoplásica/genética , Células Cultivadas , Chlorocebus aethiops , Ciclina D1/genética , Proteínas de Ligação a DNA/genética , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Camundongos , Camundongos Knockout , Células NIH 3T3 , Proteínas Nucleares/genética , Fosforilação , Ligação Proteica/genética , Proto-Oncogene Mas , Proteínas Proto-Oncogênicas c-fos/genética , Proteínas Proto-Oncogênicas c-fos/metabolismo , Proteínas Proto-Oncogênicas c-jun/genética , Fator de Transcrição AP-1/genética , Proteína Tumoral p73 , Proteínas Supressoras de Tumor/genética , Regulação para Cima/genética
4.
Proc Natl Acad Sci U S A ; 107(11): 4902-7, 2010 Mar 16.
Artigo em Inglês | MEDLINE | ID: mdl-20185758

RESUMO

p73, the structural and functional homologue of p53, exists as two major forms: the transactivation-proficient, proapoptotic TAp73 or the transactivation-deficient, antiapoptotic DNp73. Expectedly, expression of both these major forms has to be coordinated precisely to achieve the desired cellular outcome. Genotoxic insults resulting in cell death lead to the stabilization of TAp73, mainly through posttranslational modifications, and the concomitant degradation of DNp73, through poorly understood mechanisms. We have therefore investigated the possible mechanisms of stress-induced DNp73 degradation and show here that c-Jun, the AP-1 family member activated by stress signals and involved in stabilizing TAp73, promotes DNp73 degradation. Genotoxic stress-mediated DNp73 degradation was found to occur in a c-Jun-dependent manner through a ubiquitin-independent but proteasome-dependent mechanism. Absence or down-regulation of c-Jun expression abrogated the reduction of DNp73 levels upon stress insults, whereas overexpression of c-Jun led to its degradation. c-Jun controlled DNp73 degradation through the nonclassical, polyamine-induced antizyme (Az) pathway by regulating the latter's processing during stress response. Consistently, expression of c-Jun or Az, or addition of polyamines, promoted DNp73 degradation, whereas silencing Az expression or inhibiting Az activity in cells exposed to stress reduced c-Jun-dependent DNp73 degradation. Moreover, Az was able to bind to DNp73. These data together demonstrate the existence of a c-Jun-dependent mechanism regulating the abundance of the antiapoptotic DNp73 in response to genotoxic stress.


Assuntos
Apoptose , Dano ao DNA , Proteínas de Ligação a DNA/metabolismo , Proteínas Nucleares/metabolismo , Processamento de Proteína Pós-Traducional , Proteínas/metabolismo , Proteínas Proto-Oncogênicas c-jun/metabolismo , Transdução de Sinais , Proteínas Supressoras de Tumor/metabolismo , Animais , Linhagem Celular , Humanos , Camundongos , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteína Tumoral p73 , Ubiquitina/metabolismo , Ubiquitinação
5.
Nat Commun ; 13(1): 1257, 2022 03 10.
Artigo em Inglês | MEDLINE | ID: mdl-35273168

RESUMO

The folding capacity of membrane and secretory proteins in the endoplasmic reticulum (ER) can be challenged by physiological and pathological perturbations, causing ER stress. If unresolved, this leads to cell death. We report a role for iRhom pseudoproteases in controlling apoptosis due to persistent ER stress. Loss of iRhoms causes cells to be resistant to ER stress-induced apoptosis. iRhom1 and iRhom2 interact with IP3 receptors, critical mediators of intracellular Ca2+ signalling, and regulate ER stress-induced transport of Ca2+ into mitochondria, a primary trigger of mitochondrial membrane depolarisation and cell death. iRhoms also bind to the anti-apoptotic regulator BCL-2, attenuating the inhibitory interaction between BCL-2 and IP3 receptors, which promotes ER Ca2+ release. The discovery of the participation of iRhoms in the control of ER stress-induced cell death further extends their potential pathological significance to include diseases dependent on protein misfolding and aggregation.


Assuntos
Estresse do Retículo Endoplasmático , Proteínas Proto-Oncogênicas c-bcl-2 , Apoptose , Proteínas de Transporte/metabolismo , Retículo Endoplasmático/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo
6.
Open Biol ; 9(3): 190003, 2019 03 29.
Artigo em Inglês | MEDLINE | ID: mdl-30890028

RESUMO

iRhom proteins are catalytically inactive relatives of rhomboid intramembrane proteases. There is a rapidly growing body of evidence that these pseudoenzymes have a central function in regulating inflammatory and growth factor signalling and consequent roles in many diseases. iRhom pseudoproteases have evolved new domains from their proteolytic ancestors, which are integral to their modular regulation and functions. Although we cannot yet conclude the full extent of their molecular and cellular mechanisms, there is a clearly emerging theme that they regulate the stability and trafficking of other membrane proteins. In the best understood case, iRhoms act as regulatory cofactors of the ADAM17 protease, controlling its function of shedding cytokines and growth factors. It seems likely that as the involvement of iRhoms in human diseases is increasingly recognized, they will become the focus of pharmaceutical interest, and here we discuss what is known about their molecular mechanisms and relevance in known pathologies.


Assuntos
Regulação da Expressão Gênica , Predisposição Genética para Doença/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , Nefropatias/genética , Mutação , Neoplasias Gástricas/genética , Proteína ADAM17/genética , Proteína ADAM17/metabolismo , Animais , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/classificação , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Filogenia , Transporte Proteico/genética
7.
Artigo em Inglês | MEDLINE | ID: mdl-30057793

RESUMO

TAp73, the homologue of the tumour suppressor p53, has dual roles in tumourigenesis: both as a tumour suppressor and as a promoter of tumour growth. We have recently shown that hypoxia, a condition prevalent in tumours, results in the stabilisation of TAp73 through a mechanism involving HIF-1α-mediated repression of the E3 ligase Siah1. Elevated TAp73 in turn regulates the angiogenic transcriptional programme, exemplified by vegf-A activation, thereby promoting angiogenesis and tumour growth. To further understand hypoxia-mediated TAp73 regulation, we have focused on the Adenosine monophosphate (AMP)-dependent protein kinase (AMPK) signalling pathway induced by hypoxia. We show that hypoxia-mediated AMPK activation is required for efficient TAp73 stabilisation, through multiple means by using AMPK-deficient cells or inhibiting its activity and expression. Conversely, direct AMPK activation using its activator AICAR is also sufficient to induce TAp73 stabilisation but this is independent of putative AMPK phosphorylation sites on TAp73, HIF-1α activation, and transcriptional repression of Siah1. Furthermore, while vegf-A up-regulation upon hypoxia requires AMPK, direct activation of AMPK by AICAR does not activate vegf-A. Consistently, supernatant from cells exposed to hypoxia, but not AICAR, was able to induce tube formation in HUVECs. These data therefore highlight that the processes of TAp73 stabilisation and transcriptional activation of angiogenic target genes by AMPK activation can be decoupled. Collectively, these results suggest that the context of AMPK activation determines the effect on TAp73, and proposes a model in which hypoxia-induced TAp73 stabilisation occurs by parallel pathways converging to mediate its transactivation potential.

9.
Cell Cycle ; 14(22): 3533-9, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26267146

RESUMO

P73, the homolog of p53, exists in 2 major forms: either as a pro-apoptotic TAp73 or an amino-terminally truncated DNp73, the latter lacking the first transactivation domain. While TAp73s tumor suppressive functions have been established, DNp73 is an anti-apoptotic protein conferring chemoresistance and is associated with poor survival. However, both forms are variably overexpressed in many human cancers. In this context, we have recently demonstrated that TAp73 is stabilized by hypoxia, a tumor-relevant condition that is associated with cell survival, via HIF-1α-mediated suppression of Siah1 E3 ligase that degrades TAp73. Consequently, hypoxic signals lead to TAp73-mediated activation of several angiogenic genes and blood vessel formation, thereby supporting tumorigenesis. We show here that, similar to TAp73, DNp73 is stabilized by hypoxia in a HIF-1α-dependent manner, which otherwise is degraded by Siah1. Moreover, DNp73 is capable of inducing the expression of Vegf-A, the prototypic angiogenic gene, and loss of DNp73 expression results in reduction in tumor vasculature and size. These data therefore indicate a common mode of regulation for both p73 forms by hypoxia, resulting in the promotion of angiogenesis and tumor growth, highlighting common functionality of these antagonistic proteins under specific physiological contexts.


Assuntos
Proteínas de Ligação a DNA/genética , Regulação Neoplásica da Expressão Gênica , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Hipóxia/genética , Proteínas Nucleares/genética , Proteínas Supressoras de Tumor/genética , Ubiquitina-Proteína Ligases/genética , Fator A de Crescimento do Endotélio Vascular/genética , Proteínas E1A de Adenovirus/genética , Proteínas E1A de Adenovirus/metabolismo , Animais , Hipóxia Celular , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/metabolismo , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Feminino , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Células HEK293 , Humanos , Hipóxia/metabolismo , Hipóxia/patologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Camundongos , Camundongos SCID , Proteínas Nucleares/metabolismo , Oxigênio/farmacologia , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Estabilidade Proteica , Proteólise , Transdução de Sinais , Transformação Genética , Proteína Tumoral p73 , Proteínas Supressoras de Tumor/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto , Proteínas ras/genética , Proteínas ras/metabolismo
10.
Nat Cell Biol ; 17(4): 511-23, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25774835

RESUMO

The functional significance of the overexpression of unmutated TAp73, a homologue of the tumour suppressor p53, in multiple human cancers is unclear, but raises the possibility of unidentified roles in promoting tumorigenesis. We show here that TAp73 is stabilized by hypoxia, a condition highly prevalent in tumours, through HIF-1α-mediated repression of the ubiquitin ligase Siah1, which targets TAp73 for degradation. Consequently, TAp73-deficient tumours are less vascular and reduced in size, and conversely, TAp73 overexpression leads to increased vasculature. Moreover, we show that TAp73 is a critical regulator of the angiogenic transcriptome and is sufficient to directly activate the expression of several angiogenic genes.  Finally, expression of TAp73 positively correlates with these angiogenic genes in several human tumours, and the angiogenic gene signature is sufficient to segregate the TAp73(Hi)- from TAp73(Low)-expressing tumours. These data demonstrate a pro-angiogenic role for TAp73 in supporting tumorigenesis, providing a rationale for its overexpression in cancers.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Neoplasias/irrigação sanguínea , Neovascularização Patológica/genética , Proteínas Nucleares/antagonistas & inibidores , Proteínas Nucleares/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Ubiquitina-Proteína Ligases/antagonistas & inibidores , Animais , Hipóxia Celular , Linhagem Celular Tumoral , Transformação Celular Neoplásica/genética , Proteínas de Ligação a DNA/biossíntese , Proteínas de Ligação a DNA/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Células HCT116 , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/biossíntese , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos SCID , Proteínas Nucleares/biossíntese , Proteínas Nucleares/genética , Ligação Proteica , Proteína Tumoral p73 , Proteínas Supressoras de Tumor/biossíntese , Proteínas Supressoras de Tumor/genética , Ubiquitina-Proteína Ligases/genética , Ubiquitinação , Fator A de Crescimento do Endotélio Vascular/biossíntese
11.
Neoplasia ; 6(5): 546-57, 2004.
Artigo em Inglês | MEDLINE | ID: mdl-15548364

RESUMO

Although p73 is a structural and functional homologue of the tumor-suppressor gene p53, it is not mutated in many human cancers as p53. Besides, p73 was shown to be activated by only a subset of signals that activate p53, such as gamma-irradiation and cisplatin, but not by other common genotoxic stress-inducing agents such as ultraviolet (UV) irradiation, although many of these signals are also capable of inducing p53-independent cell death. Using a p73-specific antibody, we confirmed that c-Abl is required for cisplatin-induced p73 upregulation, and further demonstrate that the p73 protein is upregulated by UV irradiation and other stress stimuli including sorbitol, hydrogen peroxide, nocodazol, and taxol. These stress signals upregulate both p73 mRNA and increases the stability of p73, indicating that p73 is regulated transcriptionally and posttranslationally. Cells stably expressing the dominant-negative p73 inhibitor protein (p73DD) and p73(-/-) fibroblasts are more resistant than control cells to apoptosis induced by these stress signals, suggesting that p73 contributes to apoptosis induction. Together, the data demonstrate that several stress signals can signal to p73 in vivo, which raises the possibility of eradicating cancers with an unmutated p73 gene by activating them with stress-inducing agents or their mimetics.


Assuntos
Proteínas de Ligação a DNA/biossíntese , Proteínas Nucleares/biossíntese , Regulação para Cima , Animais , Antibióticos Antineoplásicos/farmacologia , Apoptose/genética , Células COS , Extratos Celulares/imunologia , Chlorocebus aethiops , Cisplatino/farmacologia , Proteínas de Ligação a DNA/análise , Proteínas de Ligação a DNA/genética , Doxorrubicina/farmacologia , Expressão Gênica/efeitos dos fármacos , Expressão Gênica/efeitos da radiação , Genes Supressores de Tumor , Humanos , Proteínas Nucleares/análise , Proteínas Nucleares/genética , Paclitaxel/farmacologia , Proteínas Proto-Oncogênicas c-abl/análise , Proteínas Proto-Oncogênicas c-abl/metabolismo , RNA Mensageiro/análise , RNA Mensageiro/metabolismo , Proteína Tumoral p73 , Proteína Supressora de Tumor p53/metabolismo , Proteínas Supressoras de Tumor , Raios Ultravioleta
12.
J Biol Chem ; 280(31): 28203-14, 2005 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-15919663

RESUMO

The tumor suppressor p53 regulates its own stability by transcriptionally activating Mdm2, Pirh2, and COP1, which target p53 for degradation. However, whether such a negative feedback mechanism exists to regulate the stability of p73, the structural and functional homologue of p53, is unclear. Unlike p53, p73 is not mutated in cancers, but its expression is significantly elevated. Thus, we have investigated the regulation of p73 turnover. Our data suggest the existence of a negative feedback mechanism for p73 degradation. p73 mutants with compromised transactivation activity are generally more stable than the full-length TAp73 form. TAp73 appears to promote its own turnover as well as that of other p73 forms, including the DeltaNp73 that lacks the amino-terminal transactivation domain, in a transactivation-dependent manner. This degradation-inducing property of TAp73 was inhibited only by p73 mutants that also inhibit the transactivation activity TAp73 but not by mutant p53, highlighting the specificity in the regulation of p73 stability. Moreover, regions in the amino and carboxyl termini of p73 confer both stabilizing and destabilizing effects on the protein, independent of its transactivation ability. Finally, we have identified the regions between amino acids 56 and 248 of p73 as being the region required for p73-mediated and for ubiquitin-mediated degradation. Taken together, the data suggest that p73 turnover is tightly regulated in a transactivation-dependent and -independent manner, resulting in the controlled expression of the various p73 forms.


Assuntos
Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Sequência de Bases , Linhagem Celular Tumoral , Clonagem Molecular , Cicloeximida/farmacologia , Primers do DNA , Proteínas de Ligação a DNA/efeitos dos fármacos , Genes Reporter , Genes Supressores de Tumor/efeitos dos fármacos , Homeostase , Humanos , Cinética , Luciferases/genética , Luciferases/metabolismo , Neoplasias Pulmonares , Proteínas Nucleares/efeitos dos fármacos , Deleção de Sequência , Ativação Transcricional , Proteína Tumoral p73 , Proteínas Supressoras de Tumor
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