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1.
Mol Psychiatry ; 26(11): 6411-6426, 2021 11.
Artigo em Inglês | MEDLINE | ID: mdl-34002021

RESUMO

Several psychiatric, neurologic and neurodegenerative disorders present increased brain ventricles volume, being hydrocephalus the disease with the major manifestation of ventriculomegaly caused by the accumulation of high amounts of cerebrospinal fluid (CSF). The molecules and pathomechanisms underlying cerebral ventricular enlargement are widely unknown. Kinase D interacting substrate of 220 kDa (KIDINS220) gene has been recently associated with schizophrenia and with a novel syndrome characterized by spastic paraplegia, intellectual disability, nystagmus and obesity (SINO syndrome), diseases frequently occurring with ventriculomegaly. Here we show that Kidins220, a transmembrane protein effector of various key neuronal signalling pathways, is a critical regulator of CSF homeostasis. We observe that both KIDINS220 and the water channel aquaporin-4 (AQP4) are markedly downregulated at the ventricular ependymal lining of idiopathic normal pressure hydrocephalus (iNPH) patients. We also find that Kidins220 deficient mice develop ventriculomegaly accompanied by water dyshomeostasis and loss of AQP4 in the brain ventricular ependymal layer and astrocytes. Kidins220 is a known cargo of the SNX27-retromer, a complex that redirects endocytosed plasma membrane proteins (cargos) back to the cell surface, thus avoiding their targeting to lysosomes for degradation. Mechanistically, we show that AQP4 is a novel cargo of the SNX27-retromer and that Kidins220 deficiency promotes a striking and unexpected downregulation of the SNX27-retromer that results in AQP4 lysosomal degradation. Accordingly, SNX27 silencing decreases AQP4 levels in wild-type astrocytes whereas SNX27 overexpression restores AQP4 content in Kidins220 deficient astrocytes. Together our data suggest that the KIDINS220-SNX27-retromer-AQP4 pathway is involved in human ventriculomegaly and open novel therapeutic perspectives.


Assuntos
Hidrocefalia , Animais , Aquaporina 4/genética , Aquaporina 4/metabolismo , Epêndima/metabolismo , Humanos , Hidrocefalia/genética , Hidrocefalia/metabolismo , Lisossomos/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Nexinas de Classificação/genética
2.
Haematologica ; 105(3): 730-740, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31221787

RESUMO

Metastases, the major cause of death from cancer, require cells' acquisition of the ability to migrate and involve multiple steps, including local tumor cell invasion and basement membrane penetration. Certain lymphoid tumors are highly metastatic, but the mechanisms of invasion by lymphoma cells are poorly understood. We recently showed that CDCA7, a protein induced by MYC, is overexpressed in lymphoid tumors and that its knockdown decreases lymphoid tumor growth without inhibiting the proliferation of normal cells. Here we show that CDCA7 is critical for invasion and migration of lymphoma cells. Indeed, CDCA7 knockdown in lymphoma cells limited tumor cell invasion in matrigel-coated transwell plates and tumor invasion of neighboring tissues in a mouse xenograft model and in a zebrafish model of cell invasion. CDCA7 silencing markedly inhibited lymphoma cell migration on fibronectin without modifying cell adhesion to this protein. Instead, CDCA7 knockdown markedly disrupted the precise dynamic reorganization of actomyosin and tubulin cytoskeletons required for efficient migration. In particular, CDCA7 silencing impaired tubulin and actomyosin cytoskeleton polarization, increased filamentous actin formation, and induced myosin activation. Of note, inhibitors of actin polymerization, myosin II, or ROCK reestablished the migration capacity of CDCA7-silenced lymphoma cells. Given the critical role of CDCA7 in lymphoma-genesis and invasion, therapies aimed at inhibiting its expression or activity might provide significant control of lymphoma growth, invasion, and metastatic dissemination.


Assuntos
Linfoma , Peixe-Zebra , Animais , Carcinogênese , Linhagem Celular Tumoral , Movimento Celular , Citoesqueleto , Linfoma/genética , Camundongos , Invasividade Neoplásica
3.
Nucleic Acids Res ; 45(17): 9960-9975, 2017 Sep 29.
Artigo em Inglês | MEDLINE | ID: mdl-28973440

RESUMO

Most E2F-binding sites repress transcription through the recruitment of Retinoblastoma (RB) family members until the end of the G1 cell-cycle phase. Although the MYB promoter contains an E2F-binding site, its transcription is activated shortly after the exit from quiescence, before RB family members inactivation, by unknown mechanisms. We had previously uncovered a nuclear factor distinct from E2F, Myb-sp, whose DNA-binding site overlapped the E2F element and had hypothesized that this factor might overcome the transcriptional repression of MYB by E2F-RB family members. We have purified Myb-sp and discovered that Myc-associated zinc finger proteins (MAZ) are major components. We show that various MAZ isoforms are present in Myb-sp and activate transcription via the MYB-E2F element. Moreover, while forced RB or p130 expression repressed the activity of a luciferase reporter driven by the MYB-E2F element, co-expression of MAZ proteins not only reverted repression, but also activated transcription. Finally, we show that MAZ binds the MYB promoter in vivo, that its binding site is critical for MYB transactivation, and that MAZ knockdown inhibits MYB expression during the exit from quiescence. Together, these data indicate that MAZ is essential to bypass MYB promoter repression by RB family members and to induce MYB expression.


Assuntos
Proteínas de Ligação a DNA/genética , Fatores de Transcrição E2F/genética , Fase G1/genética , Regulação da Expressão Gênica , Proteínas Oncogênicas v-myb/genética , Regiões Promotoras Genéticas , Fatores de Transcrição/genética , Sítios de Ligação , Linhagem Celular Tumoral , Proteína Substrato Associada a Crk/genética , Proteína Substrato Associada a Crk/metabolismo , Proteínas de Ligação a DNA/antagonistas & inibidores , Proteínas de Ligação a DNA/metabolismo , Fatores de Transcrição E2F/metabolismo , Genes Reporter , Células HEK293 , Humanos , Células Jurkat , Luciferases/genética , Luciferases/metabolismo , Linfócitos/citologia , Linfócitos/metabolismo , Proteínas Oncogênicas v-myb/metabolismo , Osteoblastos/citologia , Osteoblastos/metabolismo , Ligação Proteica , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Proteína do Retinoblastoma/genética , Proteína do Retinoblastoma/metabolismo , Fatores de Transcrição/antagonistas & inibidores , Fatores de Transcrição/metabolismo , Transcrição Gênica
4.
Haematologica ; 103(10): 1669-1678, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-29880607

RESUMO

Tumor formation involves the acquisition of numerous capacities along the progression from a normal cell into a malignant cell, including limitless proliferation (immortalization) and anchorage-independent growth, a capacity that correlates extremely well with tumorigenesis. Great efforts have been made to uncover genes involved in tumor formation, but most genes identified participate in processes related to cell proliferation. Accordingly, therapies targeting these genes also affect the proliferation of normal cells. To identify potential targets for therapeutic intervention more specific to tumor cells, we looked for genes implicated in the acquisition of anchorage-independent growth and in vivo tumorigenesis capacity. A transcriptomic analysis identified CDCA7 as a candidate gene. Indeed, CDCA7 protein was upregulated in Burkitt's lymphoma cell lines and human tumor biopsy specimens relative to control cell lines and tissues, respectively. CDCA7 levels were also markedly elevated in numerous T and B-lymphoid tumor cell lines. While CDCA7 was not required for anchorage-dependent growth of normal fibroblasts or non-malignant lymphocytes, it was essential but not sufficient for anchorage-independent growth of lymphoid tumor cells and for lymphomagenesis. These data suggest that therapies aimed at inhibiting CDCA7 expression or function might significantly decrease the growth of lymphoid tumors.


Assuntos
Linfoma de Burkitt/metabolismo , Carcinogênese/metabolismo , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Proteínas de Neoplasias/biossíntese , Proteínas Nucleares/biossíntese , Regulação para Cima , Linfoma de Burkitt/genética , Linfoma de Burkitt/patologia , Carcinogênese/genética , Carcinogênese/patologia , Feminino , Células HCT116 , Células HeLa , Humanos , Células Jurkat , Células K562 , Masculino , Proteínas de Neoplasias/genética , Proteínas Nucleares/genética , Células U937
5.
Pharmacol Res ; 133: 236-249, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-29309904

RESUMO

Cyclooxygenase-2 (COX-2) derived-prostanoids participate in the altered vascular function and mechanical properties in cardiovascular diseases. We investigated whether regulator of calcineurin 1 (Rcan1) participates in vascular contractility and stiffness through the regulation of COX-2. For this, wild type (Rcan1+/+) and Rcan1-deficient (Rcan1-/-) mice untreated or treated with the COX-2 inhibitor rofecoxib were used. Vascular function and structure were analysed by myography. COX-2 and phospo-p65 expression were studied by western blotting and immunohistochemistry and TXA2 production by ELISA. We found that Rcan1 deficiency increases COX-2 and IL-6 expression and NF-κB activation in arteries and vascular smooth muscle cells (VSMC). Adenoviral-mediated re-expression of Rcan1.4 in Rcan1-/- VSMC normalized COX-2 expression. Phenylephrine-induced vasoconstrictor responses were greater in aorta from Rcan1-/- compared to Rcan1+/+ mice. This increased response were diminished by etoricoxib, furegrelate, SQ 29548, cyclosporine A and parthenolide, inhibitors of COX-2, TXA2 synthase, TP receptors, calcineurin and NF-κB, respectively. Endothelial removal and NOS inhibition increased phenylephrine responses only in Rcan1+/+ mice. TXA2 levels were greater in Rcan1-/- mice. In small mesenteric arteries, vascular function and structure were similar in both groups of mice; however, vessels from Rcan1-/- mice displayed an increase in vascular stiffness that was diminished by rofecoxib. In conclusion, our results suggest that Rcan1 might act as endogenous negative modulator of COX-2 expression and activity by inhibiting calcineurin and NF-kB pathways to maintain normal contractility and vascular stiffness in aorta and small mesenteric arteries, respectively. Our results uncover a new role for Rcan1 in vascular contractility and mechanical properties.


Assuntos
Aorta Torácica/fisiologia , Ciclo-Oxigenase 2/fisiologia , Peptídeos e Proteínas de Sinalização Intracelular/fisiologia , Artérias Mesentéricas/fisiologia , Proteínas Musculares/fisiologia , Músculo Liso Vascular/fisiologia , Animais , Proteínas de Ligação ao Cálcio , Células Cultivadas , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Contração Muscular , Músculo Liso Vascular/citologia , Miócitos de Músculo Liso/fisiologia
7.
Hum Mol Genet ; 22(3): 466-82, 2013 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-23118350

RESUMO

Failures in neurotrophic support and signalling play key roles in Alzheimer's disease (AD) pathogenesis. We previously demonstrated that downregulation of the neurotrophin effector Kinase D interacting substrate (Kidins220) by excitotoxicity and cerebral ischaemia contributed to neuronal death. This downregulation, triggered through overactivation of N-methyl-D-aspartate receptors (NMDARs), involved proteolysis of Kidins220 by calpain and transcriptional inhibition. As excitotoxicity is at the basis of AD aetiology, we hypothesized that Kidins220 might also be downregulated in this disease. Unexpectedly, Kidins220 is augmented in necropsies from AD patients where it accumulates with hyperphosphorylated tau. This increase correlates with enhanced Kidins220 resistance to calpain processing but no higher gene transcription. Using AD brain necropsies, glycogen synthase kinase 3-ß (GSK3ß)-transgenic mice and cell models of AD-related neurodegeneration, we show that GSK3ß phosphorylation decreases Kidins220 susceptibility to calpain proteolysis, while protein phosphatase 1 (PP1) action has the opposite effect. As altered activities of GSK3ß and phosphatases are involved in tau aggregation and constitute hallmarks in AD, a GSK3ß/PP1 imbalance may also contribute to Kidins220 decreased clearance, accumulation and hampered neurotrophin signalling from early stages of the disease pathogenesis. These results encourage searches for mutations in Kidins220 gene and their possible associations to dementias. Finally, our data support a model where the effects of excitotoxicity drastically differ when occurring in cerebral ischaemia versus progressively sustained toxicity along AD progression. The striking differences in Kidins220 stability resulting from chronic versus acute brain damage may also have important implications for the therapeutic intervention of neurodegenerative disorders.


Assuntos
Doença de Alzheimer/metabolismo , Calpaína/metabolismo , Quinase 3 da Glicogênio Sintase/metabolismo , Proteínas de Membrana/genética , Proteínas do Tecido Nervoso/genética , Proteína Fosfatase 1/metabolismo , Proteínas tau/metabolismo , Idoso , Idoso de 80 Anos ou mais , Doença de Alzheimer/genética , Doença de Alzheimer/patologia , Animais , Isquemia Encefálica/genética , Isquemia Encefálica/patologia , Calpaína/genética , Morte Celular , Células Cultivadas , Modelos Animais de Doenças , Regulação para Baixo , Feminino , Quinase 3 da Glicogênio Sintase/genética , Glicogênio Sintase Quinase 3 beta , Células HEK293 , Humanos , Masculino , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Transgênicos , Fatores de Crescimento Neural/genética , Fatores de Crescimento Neural/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Doenças Neurodegenerativas/induzido quimicamente , Doenças Neurodegenerativas/patologia , Neurônios/citologia , Neurônios/patologia , Ácido Okadáico/efeitos adversos , Fosforilação , Proteína Fosfatase 1/antagonistas & inibidores , Proteína Fosfatase 1/genética , Proteólise , Ratos , Ratos Wistar , Receptores de N-Metil-D-Aspartato/genética , Receptores de N-Metil-D-Aspartato/metabolismo , Transdução de Sinais , Proteínas tau/genética
8.
EMBO Mol Med ; 16(1): 132-157, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-38177536

RESUMO

Thoracic aortic aneurysm and dissection (TAAD) is a life-threatening condition associated with Marfan syndrome (MFS), a disease caused by fibrillin-1 gene mutations. While various conditions causing TAAD exhibit aortic accumulation of the proteoglycans versican (Vcan) and aggrecan (Acan), it is unclear whether these ECM proteins are involved in aortic disease. Here, we find that Vcan, but not Acan, accumulated in Fbn1C1041G/+ aortas, a mouse model of MFS. Vcan haploinsufficiency protected MFS mice against aortic dilation, and its silencing reverted aortic disease by reducing Nos2 protein expression. Our results suggest that Acan is not an essential contributor to MFS aortopathy. We further demonstrate that Vcan triggers Akt activation and that pharmacological Akt pathway inhibition rapidly regresses aortic dilation and Nos2 expression in MFS mice. Analysis of aortic tissue from MFS human patients revealed accumulation of VCAN and elevated pAKT-S473 staining. Together, these findings reveal that Vcan plays a causative role in MFS aortic disease in vivo by inducing Nos2 via Akt activation and identify Akt signaling pathway components as candidate therapeutic targets.


Assuntos
Aneurisma da Aorta Torácica , Doenças da Aorta , Dissecção Aórtica , Azidas , Desoxiglucose , Síndrome de Marfan , Animais , Humanos , Camundongos , Aneurisma da Aorta Torácica/complicações , Aneurisma da Aorta Torácica/genética , Aneurisma da Aorta Torácica/metabolismo , Doenças da Aorta/complicações , Desoxiglucose/análogos & derivados , Síndrome de Marfan/complicações , Síndrome de Marfan/genética , Síndrome de Marfan/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Versicanas/metabolismo
9.
J Immunol ; 186(12): 6726-36, 2011 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-21572027

RESUMO

c-Myc, a member of the Myc family of transcription factors, is involved in numerous biological functions including the regulation of cell proliferation, differentiation, and apoptosis in various cell types. Of all of its functions, the role of c-Myc in cell differentiation is one of the least understood. We addressed the role of c-Myc in B lymphocyte differentiation. We found that c-Myc is essential from early stages of B lymphocyte differentiation in vivo and regulates this process by providing B cell identity via direct transcriptional regulation of the ebf-1 gene. Our data show that c-Myc influences early B lymphocyte differentiation by promoting activation of B cell identity genes, thus linking this transcription factor to the EBF-1/Pax-5 pathway.


Assuntos
Linfócitos B/citologia , Diferenciação Celular/imunologia , Regulação da Expressão Gênica/imunologia , Proteínas Proto-Oncogênicas c-myc/imunologia , Animais , Camundongos , Fator de Transcrição PAX5/metabolismo , Proto-Oncogene Mas , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Transativadores/genética , Transativadores/metabolismo , Fatores de Transcrição/imunologia , Transcrição Gênica
10.
Cell Death Dis ; 14(8): 500, 2023 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-37542079

RESUMO

In the adult mammalian brain, neural stem cells (NSCs) located in highly restricted niches sustain the generation of new neurons that integrate into existing circuits. A reduction in adult neurogenesis is linked to ageing and neurodegeneration, whereas dysregulation of proliferation and survival of NSCs have been hypothesized to be at the origin of glioma. Thus, unravelling the molecular underpinnings of the regulated activation that NSCs must undergo to proliferate and generate new progeny is of considerable relevance. Current research has identified cues promoting or restraining NSCs activation. Yet, whether NSCs depend on external signals to survive or if intrinsic factors establish a threshold for sustaining their viability remains elusive, even if this knowledge could involve potential for devising novel therapeutic strategies. Kidins220 (Kinase D-interacting substrate of 220 kDa) is an essential effector of crucial pathways for neuronal survival and differentiation. It is dramatically altered in cancer and in neurological and neurodegenerative disorders, emerging as a regulatory molecule with important functions in human disease. Herein, we discover severe neurogenic deficits and hippocampal-based spatial memory defects accompanied by increased neuroblast death and high loss of newly formed neurons in Kidins220 deficient mice. Mechanistically, we demonstrate that Kidins220-dependent activation of AKT in response to EGF restraints GSK3 activity preventing NSCs apoptosis. We also show that NSCs with Kidins220 can survive with lower concentrations of EGF than the ones lacking this molecule. Hence, Kidins220 levels set a molecular threshold for survival in response to mitogens, allowing adult NSCs growth and expansion. Our study identifies Kidins220 as a key player for sensing the availability of growth factors to sustain adult neurogenesis, uncovering a molecular link that may help paving the way towards neurorepair.


Assuntos
Células-Tronco Adultas , Células-Tronco Neurais , Adulto , Animais , Humanos , Camundongos , Células-Tronco Adultas/metabolismo , Fator de Crescimento Epidérmico/metabolismo , Quinase 3 da Glicogênio Sintase/metabolismo , Hipocampo/metabolismo , Mamíferos , Células-Tronco Neurais/metabolismo , Neurogênese/fisiologia , Neurônios/metabolismo
11.
Eur J Immunol ; 41(4): 1035-46, 2011 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-21381019

RESUMO

Kinase D interacting substrate of 220 kDa (Kidins220), also known as ankyrin repeat-rich membrane spanning (ARMS), is a protein that is mainly expressed in brain and neural cells where its function is only starting to be characterized. Here, we show that Kidins220/ARMS is also expressed in T lymphocytes where it is highly concentrated at the uropod of polarized T cells. In this cellular model, Kidins220/ARMS colocalizes with typical uropod T-cell molecules and coimmunoprecipitates with ICAM-3. Furthermore, Kidins220/ARMS associates with raft domains at the uropod and coimmunoprecipitates with caveolin-1, a molecule we show here to be also expressed in T cells. Importantly, induction of morphological polarization in primary T lymphocytes and Jurkat cells enhances Kidins220/ARMS colocalization with ICAM-3. Conversely, disruption of cell polarity provokes Kidins220/ARMS redistribution from the uropod to other cellular regions and drastically impairs its association with ICAM-3 in a protein kinase C-dependent manner. Finally, Kidins220/ARMS knockdown in human polarized T-cell lines promotes both basal and stromal cell-derived factor-1α-induced directed migration, identifying a novel function for this molecule. Altogether, our findings show that Kidins220/ARMS is a novel component of the uropod involved in the regulation of T-cell motility, an essential process for the immune response.


Assuntos
Antígenos CD/imunologia , Moléculas de Adesão Celular/imunologia , Movimento Celular , Proteínas de Membrana/imunologia , Proteínas do Tecido Nervoso/imunologia , Linfócitos T/citologia , Linfócitos T/imunologia , Animais , Caveolina 1/metabolismo , Polaridade Celular , Células Cultivadas , Regulação da Expressão Gênica , Humanos , Proteínas de Membrana/genética , Proteínas do Tecido Nervoso/genética , Ligação Proteica , Ratos
12.
Br J Pharmacol ; 179(7): 1287-1303, 2022 04.
Artigo em Inglês | MEDLINE | ID: mdl-34599830

RESUMO

Recent studies have shown that NO is a central mediator in diseases associated with thoracic aortic aneurysm, such as Marfan syndrome. The progressive dilation of the aorta in thoracic aortic aneurysm ultimately leads to aortic dissection. Unfortunately, current medical treatments have neither halt aortic enlargement nor prevented rupture, leaving surgical repair as the only effective treatment. There is therefore a pressing need for effective therapies to delay or even avoid the need for surgical repair in thoracic aortic aneurysm patients. Here, we summarize the mechanisms through which NO signalling dysregulation causes thoracic aortic aneurysm, particularly in Marfan syndrome. We discuss recent advances based on the identification of new Marfan syndrome mediators related to pathway overactivation that represent potential disease biomarkers. Likewise, we propose iNOS, sGC and PRKG1, whose pharmacological inhibition reverses aortopathy in Marfan syndrome mice, as targets for therapeutic intervention in thoracic aortic aneurysm and are candidates for clinical trials.


Assuntos
Aneurisma da Aorta Torácica , Aneurisma Aórtico , Dissecção Aórtica , Síndrome de Marfan , Dissecção Aórtica/complicações , Dissecção Aórtica/cirurgia , Animais , Aorta , Aneurisma Aórtico/complicações , Aneurisma da Aorta Torácica/etiologia , Aneurisma da Aorta Torácica/prevenção & controle , Aneurisma da Aorta Torácica/cirurgia , Proteína Quinase Dependente de GMP Cíclico Tipo I , Humanos , Síndrome de Marfan/complicações , Síndrome de Marfan/tratamento farmacológico , Síndrome de Marfan/cirurgia , Camundongos
13.
Nat Commun ; 12(1): 2628, 2021 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-33976159

RESUMO

Thoracic aortic aneurysm, as occurs in Marfan syndrome, is generally asymptomatic until dissection or rupture, requiring surgical intervention as the only available treatment. Here, we show that nitric oxide (NO) signaling dysregulates actin cytoskeleton dynamics in Marfan Syndrome smooth muscle cells and that NO-donors induce Marfan-like aortopathy in wild-type mice, indicating that a marked increase in NO suffices to induce aortopathy. Levels of nitrated proteins are higher in plasma from Marfan patients and mice and in aortic tissue from Marfan mice than in control samples, indicating elevated circulating and tissue NO. Soluble guanylate cyclase and cGMP-dependent protein kinase are both activated in Marfan patients and mice and in wild-type mice treated with NO-donors, as shown by increased plasma cGMP and pVASP-S239 staining in aortic tissue. Marfan aortopathy in mice is reverted by pharmacological inhibition of soluble guanylate cyclase and cGMP-dependent protein kinase and lentiviral-mediated Prkg1 silencing. These findings identify potential biomarkers for monitoring Marfan Syndrome in patients and urge evaluation of cGMP-dependent protein kinase and soluble guanylate cyclase as therapeutic targets.


Assuntos
Aneurisma da Aorta Torácica/patologia , Proteína Quinase Dependente de GMP Cíclico Tipo I/metabolismo , Síndrome de Marfan/complicações , Guanilil Ciclase Solúvel/metabolismo , Animais , Aorta/citologia , Aorta/diagnóstico por imagem , Aorta/efeitos dos fármacos , Aorta/patologia , Aneurisma da Aorta Torácica/diagnóstico , Aneurisma da Aorta Torácica/etiologia , Aneurisma da Aorta Torácica/prevenção & controle , Biomarcadores/sangue , Biomarcadores/metabolismo , Carbazóis/administração & dosagem , GMP Cíclico/sangue , GMP Cíclico/metabolismo , Modelos Animais de Doenças , Feminino , Fibrilina-1/genética , Técnicas de Silenciamento de Genes , Humanos , Masculino , Síndrome de Marfan/sangue , Síndrome de Marfan/genética , Síndrome de Marfan/patologia , Camundongos , Músculo Liso Vascular/citologia , Mutação , Miócitos de Músculo Liso , Óxido Nítrico/metabolismo , Doadores de Óxido Nítrico/administração & dosagem , Cultura Primária de Células , Guanilil Ciclase Solúvel/antagonistas & inibidores , Ultrassonografia
14.
Brain Pathol ; 30(1): 120-136, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31264746

RESUMO

Huntington's disease (HD) is an inherited progressive neurodegenerative disease characterized by brain atrophy particularly in the striatum that produces motor impairment, and cognitive and psychiatric disturbances. Multiple pathogenic mechanisms have been proposed including dysfunctions in neurotrophic support and calpain-overactivation, among others. Kinase D-interacting substrate of 220 kDa (Kidins220), also known as ankyrin repeat-rich membrane spanning (ARMS), is an essential mediator of neurotrophin signaling. In adult brain, Kidins220 presents two main isoforms that differ in their carboxy-terminal length and critical protein-protein interaction domains. These variants are generated through alternative terminal exon splicing of the conventional exon 32 (Kidins220-C32) and the recently identified exon 33 (Kidins220-C33). The lack of domains encoded by exon 32 involved in key neuronal functions, including those controlling neurotrophin pathways, pointed to Kidins220-C33 as a form detrimental for neurons. However, the functional role of Kidins220-C33 in neurodegeneration or other pathologies, including HD, has not been explored. In the present work, we discover an unexpected selective downregulation of Kidins220-C33, in the striatum of HD patients, as well as in the R6/1 HD mouse model starting at early symptomatic stages. These changes are C33-specific as Kidins220-C32 variant remains unchanged. We also find the early decrease in Kidins220-C33 levels takes place in neurons, suggesting an unanticipated neuroprotective role for this isoform. Finally, using ex vivo assays and primary neurons, we demonstrate that Kidins220-C33 is downregulated by mechanisms that depend on the activation of the protease calpain. Altogether, these results strongly suggest that calpain-mediated Kidins220-C33 proteolysis modulates onset and/or progression of HD.


Assuntos
Doença de Huntington/genética , Doença de Huntington/metabolismo , Proteínas de Membrana/genética , Proteínas do Tecido Nervoso/genética , Adulto , Idoso , Processamento Alternativo , Animais , Corpo Estriado/metabolismo , Modelos Animais de Doenças , Éxons/genética , Feminino , Hipocampo/metabolismo , Humanos , Doença de Huntington/patologia , Masculino , Proteínas de Membrana/metabolismo , Camundongos , Pessoa de Meia-Idade , Proteínas do Tecido Nervoso/metabolismo , Doenças Neurodegenerativas/metabolismo , Doenças Neurodegenerativas/patologia , Neurônios/metabolismo , Neurônios/patologia , Ligação Proteica , Isoformas de Proteínas/genética , Transdução de Sinais
15.
Carcinogenesis ; 30(3): 440-8, 2009 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-19126644

RESUMO

Although C-MYC is overexpressed in a number of tumors, the mechanisms governing its expression in normal or tumor cells are not completely understood. Recruitment of the Retinoblastoma protein family members to gene promoters by E2F factors has a dominant negative effect on their activity during the G(0) and G(1) phases of the cell cycle. Despite the presence of an E2F-binding site on the C-MYC promoter, it escapes the repressive effect of E2F-Retinoblastoma complexes through unknown mechanisms during exit from quiescence. We hypothesized that occupancy of E2F elements by factors distinct from E2F might account for this escape. To test this hypothesis, we investigated whether the E2F element in the C-MYC promoter is regulated differently than E2F elements in promoters that are activated at the G(1)-S transition. Employing gel shift analysis, the E2F element from the C-MYC promoter was found to form a unique non-E2F complex, referred to as E2F C-MYC Specific (EMYCS), which is not observed with E2F elements from several other promoters. The DNA contact residues for EMYCS are distinct but overlapping with residues required for binding of E2F proteins. Finally, the approximate estimated molecular weight of the DNA-binding component of EMCYS is 105 kDa. Functional studies indicate that EMYCS has transcriptional transactivation capacity and suggest that it is required to activate the C-MYC promoter during exit from quiescence.


Assuntos
Ciclo Celular/fisiologia , Proteínas de Ligação a DNA/metabolismo , Proteínas Proto-Oncogênicas c-myc/genética , Fatores de Transcrição/metabolismo , Animais , Sítios de Ligação , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/genética , Fatores de Transcrição E2F/genética , Fatores de Transcrição E2F/metabolismo , Regulação da Expressão Gênica , Humanos , Camundongos , Regiões Promotoras Genéticas , Sequências Reguladoras de Ácido Nucleico , Fatores de Transcrição/genética , Ativação Transcricional
16.
J Clin Invest ; 115(9): 2351-62, 2005 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-16127461

RESUMO

The myelodysplastic/myeloproliferative diseases (MDS/MPDs) are a heterogeneous group of myeloid neoplasms that share characteristics with chronic myeloproliferative diseases and myelodysplastic syndromes. The broad spectrum of clinical manifestations makes MDS/MPDs extremely difficult to diagnose and treat, with a median survival time of 1-5 years. No single gene defect has been firmly associated with MDS/MPDs, and no animal models have been developed for these diseases. The association of deletions on chromosome 20q with myeloid malignancies suggests the presence of unidentified tumor suppressor genes in this region. Here we show that the recently identified death inducer-obliterator (Dido) gene gives rise to at least 3 polypeptides (Dido1, Dido2, and Dido3) through alternative splicing, and we map the human gene to the long arm of chromosome 20. We found that targeting of murine Dido caused a transplantable disease whose symptoms and signs suggested MDS/MPDs. Furthermore, 100% of human MDS/MPD patients analyzed showed Dido expression abnormalities, which we also found in other myeloid but not lymphoid neoplasms or in healthy donors. Our findings suggest that Dido might be one of the tumor suppressor genes at chromosome 20q and that the Dido-targeted mouse may be a suitable model for studying MDS/MPD diseases and testing new approaches to their diagnosis and treatment.


Assuntos
Proteínas de Ligação a DNA , Regulação da Expressão Gênica , Neoplasias Hematológicas , Síndromes Mielodisplásicas , Transtornos Mieloproliferativos , Isoformas de Proteínas , Fatores de Transcrição , Processamento Alternativo , Sequência de Aminoácidos , Animais , Medula Óssea/metabolismo , Medula Óssea/patologia , Linhagem Celular , Cromossomos Humanos Par 20 , Clonagem Molecular , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Modelos Animais de Doenças , Marcação de Genes , Neoplasias Hematológicas/genética , Neoplasias Hematológicas/fisiopatologia , Humanos , Camundongos , Dados de Sequência Molecular , Síndromes Mielodisplásicas/genética , Síndromes Mielodisplásicas/fisiopatologia , Transtornos Mieloproliferativos/genética , Transtornos Mieloproliferativos/fisiopatologia , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Alinhamento de Sequência , Baço/metabolismo , Baço/patologia , Taxa de Sobrevida , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
17.
Trends Mol Med ; 24(10): 825-837, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-30213701

RESUMO

Lentiviral vectors (LVs) transduce quiescent cells and provide stable integration to maintain transgene expression. Several approaches have been adopted to optimize LV safety profiles. Similarly, LV targeting has been tailored through strategies including the modification of envelope components, the use of specific regulatory elements, and the selection of appropriate administration routes. Models of aortic disease based on a single injection of pleiotropic LVs have been developed that efficiently transduce the three aorta layers in wild type mice. This approach allows the dissection of pathways involved in aortic aneurysm formation and the identification of targets for gene therapy in aortic diseases. LVs provide a fast, efficient, and affordable alternative to genetically modified mice to study disease mechanisms and develop therapeutic tools.


Assuntos
Proteína ADAMTS1/genética , Aneurisma Aórtico/terapia , Terapia Genética/métodos , Vetores Genéticos/química , Lentivirus/genética , Proteína ADAMTS1/antagonistas & inibidores , Proteína ADAMTS1/imunologia , Animais , Aneurisma Aórtico/genética , Aneurisma Aórtico/imunologia , Aneurisma Aórtico/patologia , Ensaios Clínicos como Assunto , Modelos Animais de Doenças , Regulação da Expressão Gênica , Vetores Genéticos/imunologia , Humanos , Imunidade Inata , Lentivirus/imunologia , Camundongos , Segurança do Paciente , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Transdução Genética/métodos , Transgenes
18.
Nat Commun ; 9(1): 4795, 2018 11 15.
Artigo em Inglês | MEDLINE | ID: mdl-30442942

RESUMO

Aortic intramural hematoma (IMH) can evolve toward reabsorption, dissection or aneurysm. Hypertension is the most common predisposing factor in IMH and aneurysm patients, and the hypertensive mediator angiotensin-II induces both in mice. We have previously shown that constitutive deletion of Rcan1 isoforms prevents Angiotensin II-induced aneurysm in mice. Here we generate mice conditionally lacking each isoform or all isoforms in vascular smooth muscle cells, endothelial cells, or ubiquitously, to determine the contribution to aneurysm development of Rcan1 isoforms in vascular cells. Surprisingly, conditional Rcan1 deletion in either vascular cell-type induces a hypercontractile phenotype and aortic medial layer disorganization, predisposing to hypertension-mediated aortic rupture, IMH, and aneurysm. These processes are blocked by ROCK inhibition. We find that Rcan1 associates with GSK-3ß, whose inhibition decreases myosin activation. Our results identify potential therapeutic targets for intervention in IMH and aneurysm and call for caution when interpreting phenotypes of constitutively and inducibly deficient mice.


Assuntos
Dissecção Aórtica/genética , Ruptura Aórtica/genética , Glicogênio Sintase Quinase 3 beta/genética , Hematoma/genética , Hipertensão/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteínas Musculares/genética , Quinases Associadas a rho/genética , Dissecção Aórtica/metabolismo , Dissecção Aórtica/patologia , Dissecção Aórtica/prevenção & controle , Animais , Anti-Hipertensivos/farmacologia , Aorta/efeitos dos fármacos , Aorta/metabolismo , Aorta/patologia , Ruptura Aórtica/metabolismo , Ruptura Aórtica/patologia , Ruptura Aórtica/prevenção & controle , Proteínas de Ligação ao Cálcio , Modelos Animais de Doenças , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Deleção de Genes , Regulação da Expressão Gênica , Predisposição Genética para Doença , Glicogênio Sintase Quinase 3 beta/antagonistas & inibidores , Glicogênio Sintase Quinase 3 beta/metabolismo , Hematoma/metabolismo , Hematoma/patologia , Hematoma/prevenção & controle , Humanos , Hipertensão/tratamento farmacológico , Hipertensão/metabolismo , Hipertensão/patologia , Peptídeos e Proteínas de Sinalização Intracelular/deficiência , Masculino , Camundongos , Camundongos Knockout , Proteínas Musculares/deficiência , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/patologia , Cultura Primária de Células , Isoformas de Proteínas/deficiência , Isoformas de Proteínas/genética , Inibidores de Proteínas Quinases/farmacologia , Quinases Associadas a rho/antagonistas & inibidores , Quinases Associadas a rho/metabolismo
19.
Nat Commun ; 9(1): 473, 2018 01 30.
Artigo em Inglês | MEDLINE | ID: mdl-29382840

RESUMO

The original version of this Article contained an error in the spelling of the author Álvaro Sebastián-Serrano, which was incorrectly given as Álvaro Sebastián Serrano. This has now been corrected in both the PDF and HTML versions of the Article.

20.
Mol Cell Biol ; 24(5): 2181-9, 2004 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-14966295

RESUMO

Cyclin G2 is an unconventional cyclin highly expressed in postmitotic cells. Unlike classical cyclins that promote cell cycle progression, cyclin G2 blocks cell cycle entry. Here we studied the mechanisms that regulate cyclin G2 mRNA expression during the cell cycle. Analysis of synchronized NIH 3T3 cell cultures showed elevated cyclin G2 mRNA expression levels at G(0), with a considerable reduction as cells enter cell cycle. Downregulation of cyclin G2 mRNA levels requires activation of phosphoinositide 3-kinase, suggesting that this enzyme controls cyclin G2 mRNA expression. Because the phosphoinositide 3-kinase pathway inhibits the FoxO family of forkhead transcription factors, we examined the involvement of these factors in the regulation of cyclin G2 expression. We show that active forms of the forkhead transcription factor FoxO3a (FKHRL1) increase cyclin G2 mRNA levels. Cyclin G2 has forkhead consensus motifs in its promoter, which are transactivated by constitutive active FoxO3a forms. Finally, interference with forkhead-mediated transcription by overexpression of an inactive form decreases cyclin G2 mRNA expression levels. These results show that FoxO genes regulate cyclin G2 expression, illustrating a new role for phosphoinositide 3-kinase and FoxO transcription factors in the control of cell cycle entry.


Assuntos
Ciclo Celular/fisiologia , Ciclinas/genética , Proteínas de Ligação a DNA/metabolismo , Regulação da Expressão Gênica , Fosfatidilinositol 3-Quinases/metabolismo , RNA Mensageiro/metabolismo , Fatores de Transcrição/metabolismo , Animais , Sequência de Bases , Proteínas de Ciclo Celular , Ciclina G1 , Ciclina G2 , Ciclinas/metabolismo , Proteínas de Ligação a DNA/genética , Regulação para Baixo , Fibroblastos/citologia , Fibroblastos/metabolismo , Proteína Forkhead Box O3 , Fatores de Transcrição Forkhead , Camundongos , Dados de Sequência Molecular , Células NIH 3T3 , Regiões Promotoras Genéticas , Transdução de Sinais/fisiologia , Fatores de Transcrição/genética , Ativação Transcricional
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