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1.
Oncogene ; 2020 Mar 10.
Artigo em Inglês | MEDLINE | ID: mdl-32157212

RESUMO

Lineage selective transcription factors (TFs) are important regulators of tumorigenesis, but their biological functions are often context dependent with undefined epigenetic mechanisms of action. In this study, we uncover a conditional role for the endodermal and pulmonary specifying TF GATA6 in lung adenocarcinoma (LUAD) progression. Impairing Gata6 in genetically engineered mouse models reduces the proliferation and increases the differentiation of Kras mutant LUAD tumors. These effects are influenced by the epithelial cell type that is targeted for transformation and genetic context of Kras-mediated tumor initiation. In LUAD cells derived from surfactant protein C expressing progenitors, we identify multiple genomic loci that are bound by GATA6. Moreover, suppression of Gata6 in these cells significantly alters chromatin accessibility, particularly at distal enhancer elements. Analogous to its paradoxical activity in lung development, GATA6 expression fluctuates during different stages of LUAD progression and can epigenetically control diverse transcriptional programs associated with bone morphogenetic protein signaling, alveolar specification, and tumor suppression. These findings reveal how GATA6 can modulate the chromatin landscape of lung cancer cells to control their proliferation and divergent lineage dependencies during tumor progression.

2.
Mol Cancer Res ; 17(12): 2343-2355, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31551255

RESUMO

The integrated stress response (ISR) is a conserved pathway that is activated by cells that are exposed to stress. In lung adenocarcinoma, activation of the ATF4 branch of the ISR by certain oncogenic mutations has been linked to the regulation of amino acid metabolism. In the present study, we provide evidence for ATF4 activation across multiple stages and molecular subtypes of human lung adenocarcinoma. In response to extracellular amino acid limitation, lung adenocarcinoma cells with diverse genotypes commonly induce ATF4 in an eIF2α-dependent manner, which can be blocked pharmacologically using an ISR inhibitor. Although suppressing eIF2α or ATF4 can trigger different biological consequences, adaptive cell-cycle progression and cell migration are particularly sensitive to inhibition of the ISR. These phenotypes require the ATF4 target gene asparagine synthetase (ASNS), which maintains protein translation independently of the mTOR/PI3K pathway. Moreover, NRF2 protein levels and oxidative stress can be modulated by the ISR downstream of ASNS. Finally, we demonstrate that ASNS controls the biosynthesis of select proteins, including the cell-cycle regulator cyclin B1, which are associated with poor lung adenocarcinoma patient outcome. Our findings uncover new regulatory layers of the ISR pathway and its control of proteostasis in lung cancer cells. IMPLICATIONS: We reveal novel regulatory mechanisms by which the ISR controls selective protein translation and is required for cell-cycle progression and migration of lung cancer cells.

3.
Medicine (Baltimore) ; 96(3): e5694, 2017 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-28099331

RESUMO

The general aim of this study was to evaluate the disease spectrum in patients presenting with a pure polymyositis (pPM) phenotype. Specific objectives were to characterize clinical features, autoantibodies (aAbs), and membrane attack complex (MAC) in muscle biopsies of patients with treatment-responsive, statin-exposed necrotizing autoimmune myositis (NAM). Patients from the Centre hospitalier de l'Université de Montréal autoimmune myositis (AIM) Cohort with a pPM phenotype, response to immunosuppression, and follow-up ≥3 years were included. Of 17 consecutive patients with pPM, 14 patients had a NAM, of whom 12 were previously exposed to atorvastatin (mean 38.8 months). These 12 patients were therefore suspected of atorvastatin-induced AIM (atorAIM) and selected for study. All had aAbs to 3-hydroxy-3-methylglutaryl coenzyme A reductase, and none had overlap aAbs, aAbs to signal recognition particle, or cancer. Three stages of myopathy were recognized: stage 1 (isolated serum creatine kinase [CK] elevation), stage 2 (CK elevation, normal strength, and abnormal electromyogram [EMG]), and stage 3 (CK elevation, proximal weakness, and abnormal EMG). At diagnosis, 10/12 (83%) patients had stage 3 myopathy (mean CK elevation: 7247 U/L). The presenting mode was stage 1 in 6 patients (50%) (mean CK elevation: 1540 U/L), all of whom progressed to stage 3 (mean delay: 37 months) despite atorvastatin discontinuation. MAC deposition was observed in all muscle biopsies (isolated sarcolemmal deposition on non-necrotic fibers, isolated granular deposition on endomysial capillaries, or mixed pattern). Oral corticosteroids alone failed to normalize CKs and induce remission. Ten patients (83%) received intravenous immune globulin (IVIG) as part of an induction regimen. Of 10 patients with ≥1 year remission on stable maintenance therapy, IVIG was needed in 50%, either with methotrexate (MTX) monotherapy or combination immunosuppression. In the remaining patients, MTX monotherapy or combination therapy maintained remission without IVIG. AtorAIM emerged as the dominant entity in patients with a pPM phenotype and treatment-responsive myopathy. Isolated CK elevation was the mode of presentation of atorAIM. The new onset of isolated CK elevation on atorvastatin and persistent CK elevation on statin discontinuation should raise early suspicion for atorAIM. Statin-induced AIM should be included in the differential diagnosis of asymptomatic hyperCKemia. Three patterns of MAC deposition, while nonpathognomonic, were pathological clues to atorAIM. AtorAIM was uniformly corticosteroid resistant but responsive to IVIG as induction and maintenance therapy.


Assuntos
Atorvastatina/efeitos adversos , Doenças Autoimunes/induzido quimicamente , Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Inibidores de Hidroximetilglutaril-CoA Redutases/efeitos adversos , Polimiosite/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Autoanticorpos/sangue , Doenças Autoimunes/tratamento farmacológico , Doenças Autoimunes/metabolismo , Doenças Autoimunes/patologia , Feminino , Glucocorticoides/uso terapêutico , Humanos , Inibidores de Hidroximetilglutaril-CoA Redutases/imunologia , Quimioterapia de Indução , Estudos Longitudinais , Masculino , Metilprednisolona/uso terapêutico , Pessoa de Meia-Idade , Músculo Esquelético/patologia , Polimiosite/tratamento farmacológico , Polimiosite/metabolismo , Polimiosite/patologia
4.
Rheumatology (Oxford) ; 56(4): 581-588, 2017 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-28013205

RESUMO

Objectives: The aim was to establish the prevalence and severity of faecal incontinence (FI) in SSc, its association with other intestinal manifestations and potential predictors of FI, and its impact on quality of life. Methods: A multicentre, cross-sectional study of 298 SSc subjects followed in the Canadian Scleroderma Research Group cohort was performed using validated questionnaires: Jorge-Wexner score (an FI severity scale), Bristol stool scale (a visual scale of stool consistency) and FI Quality-of-Life scale. Constipation was defined by the Rome III criteria. Associations between the Jorge-Wexner score and other clinical variables were determined using multivariate regression analyses. Results: Eighty-one (27.2%) subjects had FI, which was mild in 37 (12.4%) and moderate to severe in 44 (14.8%). Most patients had well-formed stools, 111 (38.8%) reported constipation and 38 (13.4%) had been previously treated for small intestinal bacterial overgrowth (SIBO). Variables independently associated with FI were: loose vs well-formed stools [odds ratio (OR) = 7.01, 95% CI: 2.09, 23.51)], constipation (OR = 3.64, 95% CI: 1.61, 8.27, P = 0.002), history of SIBO (OR = 2.97, 95% CI: 1.06, 8.27) and urinary incontinence (OR = 2.45, 95% CI: 1.14, 5.27). Quality of life measured with the FI Quality-of-Life scale was inversely correlated with FI severity (correlation coefficients between -0.602 and -0.702, P < 0.001). Conclusion: FI was common and often severe in SSc. Loose stools, SIBO, constipation and urinary incontinence were strongly associated with FI. Other than targeting anorectal dysfunction, concomitant treatment of clinical correlates could lead to improvement in FI and quality of life in SSc.


Assuntos
Incontinência Fecal/etiologia , Escleroderma Sistêmico/complicações , Adaptação Psicológica , Antibacterianos/uso terapêutico , Infecções Bacterianas/tratamento farmacológico , Canadá , Constipação Intestinal/tratamento farmacológico , Constipação Intestinal/etiologia , Estudos Transversais , Depressão/etiologia , Emoções , Incontinência Fecal/tratamento farmacológico , Feminino , Fármacos Gastrointestinais/uso terapêutico , Humanos , Enteropatias/tratamento farmacológico , Intestino Delgado , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Qualidade de Vida , Autoimagem , Incontinência Urinária/etiologia
5.
Cell Rep ; 10(8): 1288-96, 2015 Mar 03.
Artigo em Inglês | MEDLINE | ID: mdl-25732820

RESUMO

Recent evidence supports the presence of an L-glutamyl methyltransferase(s) in eukaryotic cells, but this enzyme class has been defined only in certain prokaryotic species. Here, we characterize the human C6orf211 gene product as "acidic residue methyltransferase-1" (Armt1), an enzyme that specifically targets proliferating cell nuclear antigen (PCNA) in breast cancer cells, predominately methylating glutamate side chains. Armt1 homologs share structural similarities with the SAM-dependent methyltransferases, and negative regulation of activity by automethylation indicates a means for cellular control. Notably, shRNA-based knockdown of Armt1 expression in two breast cancer cell lines altered survival in response to genotoxic stress. Increased sensitivity to UV, adriamycin, and MMS was observed in SK-Br-3 cells, while in contrast, increased resistance to these agents was observed in MCF7 cells. Together, these results lay the foundation for defining the mechanism by which this post-translational modification operates in the DNA damage response (DDR).


Assuntos
Reparo do DNA , Antígeno Nuclear de Célula em Proliferação/metabolismo , Proteína O-Metiltransferase/metabolismo , Sequência de Aminoácidos , Animais , Linhagem Celular Tumoral , Dano ao DNA , Humanos , Células MCF-7 , Metilação , Dados de Sequência Molecular , Antígeno Nuclear de Célula em Proliferação/química , Proteína O-Metiltransferase/antagonistas & inibidores , Proteína O-Metiltransferase/genética , Estrutura Terciária de Proteína , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Alinhamento de Sequência , Especificidade por Substrato
6.
EMBO Mol Med ; 5(2): 235-49, 2013 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-23165896

RESUMO

Immunoglobulins, antigens and complement can assemble to form immune complexes (IC). ICs can be detrimental as they propagate inflammation in autoimmune diseases. Like ICs, submicron extracellular vesicles termed microparticles (MP) are present in the synovial fluid from patients affected with autoimmune arthritis. We examined MPs in rheumatoid arthritis (RA) using high sensitivity flow cytometry and electron microscopy. We find that the MPs in RA synovial fluid are highly heterogeneous in size. The observed larger MPs were in fact MP-containing ICs (mpICs) and account for the majority of the detectable ICs. These mpICs frequently express the integrin CD41, consistent with platelet origin. Despite expression of the Fc receptor FcγRIIa by platelet-derived MPs, we find that the mpICs form independently of this receptor. Rather, mpICs display autoantigens vimentin and fibrinogen, and recognition of these targets by anti-citrullinated peptide antibodies contributes to the production of mpICs. Functionally, platelet mpICs are highly pro-inflammatory, eliciting leukotriene production by neutrophils. Taken together, our data suggest a unique role for platelet MPs as autoantigen-expressing elements capable of perpetuating formation of inflammatory ICs.


Assuntos
Complexo Antígeno-Anticorpo/química , Complexo Antígeno-Anticorpo/imunologia , Artrite Reumatoide/imunologia , Autoantígenos/imunologia , Autoantígenos/química , Humanos , Tamanho da Partícula , Líquido Sinovial/química , Líquido Sinovial/imunologia
7.
J Mol Biol ; 346(4): 1163-72, 2005 Mar 04.
Artigo em Inglês | MEDLINE | ID: mdl-15701524

RESUMO

The human protein p54nrb and its mouse homolog NonO have been implicated in a variety of nuclear processes including transcription, pre-mRNA processing, nuclear retention of edited RNA and DNA relaxation. We have identified p54nrb as an antigen of the phosphodependent monoclonal antibodies CC-3 and MPM-2 and shown that this protein is phosphorylated on multiple sites during mitosis. The use of the cyclin-dependent protein kinase inhibitor roscovitine and immunodepletion studies with an anti-cyclin B1 antibody established that Cdk1 was responsible for the phosphorylation of the carboxy-terminal extremity of p54nrb whereas a different kinase appeared to be involved in the generation of CC-3 epitope(s) in the amino-terminal moiety of the protein. Like many CC-3 and MPM-2 antigens, we show that p54nrb is a target of the peptidylprolyl isomerase Pin1, suggesting that it may be regulated by phosphorylation-dependent conformational changes as many other nuclear proteins upon entry into mitosis. In addition, site-directed mutagenesis indicated that the interaction of Pin1 with p54nrb was mediated by three threonine residues located in the proline-rich carboxy-terminal extremity of the protein. Our results also showed that Pin1 binding was favored when at least two of the three threonine residues were phosphorylated, suggesting a regulation mechanism based on multisite phosphorylation.


Assuntos
Mitose , Proteínas Associadas à Matriz Nuclear/metabolismo , Proteínas Nucleares/metabolismo , Peptidilprolil Isomerase/metabolismo , Proteínas de Ligação a RNA/metabolismo , Animais , Proteína Quinase CDC2/metabolismo , Epitopos/imunologia , Células HeLa , Humanos , Camundongos , Peptidilprolil Isomerase de Interação com NIMA , Proteínas Associadas à Matriz Nuclear/genética , Proteínas Nucleares/genética , Fosforilação , Ligação Proteica , Proteínas Serina-Treonina Quinases/imunologia , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas de Ligação a RNA/genética , Especificidade por Substrato
8.
BMC Cell Biol ; 5: 22, 2004 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-15171797

RESUMO

BACKGROUND: The peptidyl-prolyl isomerase Pin1 recently revealed itself as a new player in the regulation of protein function by phosphorylation. Pin1 isomerizes the peptide bond of specific phosphorylated serine or threonine residues preceding proline in several proteins involved in various cellular events including mitosis, transcription, differentiation and DNA damage response. Many Pin1 substrates are antigens of the phosphodependent monoclonal antibody MPM-2, which reacts with a subset of proteins phosphorylated at the G2/M transition. RESULTS: As MPM-2 is not a general marker of mitotic phosphoproteins, and as most mitotic substrates are phosphorylated more than once, we used a different phosphodependent antibody, mAb CC-3, to identify additional mitotic phosphoproteins and eventual Pin1 substrates by combining affinity purification, MALDI-TOF mass spectrometry and immunoblotting. Most CC-3-reactive phosphoproteins appeared to be known or novel MPM-2 antigens and included the RNA-binding protein p54nrb/nmt55, the spliceosomal protein SAP155, the Ki-67 antigen, MAP-1B, DNA topoisomerases II alpha and beta, the elongation factor hSpt5 and the largest subunit of RNA polymerase II. The CC-3 mitotic antigens were also shown to be Pin1 targets. The fine CC-3- and MPM-2-epitope mapping of the RNA polymerase II carboxy-terminal domain confirmed that the epitopes were different and could be generated in vitro by distinct kinases. Finally, the post-mitotic dephosphorylation of both CC-3 and MPM-2 antigens was prevented when cellular Pin1 activity was blocked by the selective inhibitor juglone. CONCLUSION: These observations indicate that the mitotic phosphoproteins associated with Pin1 are phosphorylated on multiple sites, suggesting combinatorial regulation of substrate recognition and isomerization.


Assuntos
Anticorpos Monoclonais/imunologia , Mitose , Peptidilprolil Isomerase/metabolismo , Fosfoproteínas/imunologia , Fosfoproteínas/metabolismo , Antígenos/química , Antígenos/imunologia , Antígenos/metabolismo , Inibidores Enzimáticos/farmacologia , Mapeamento de Epitopos , Células HeLa , Humanos , Peptidilprolil Isomerase de Interação com NIMA , Naftoquinonas/farmacologia , Peptidilprolil Isomerase/antagonistas & inibidores , Fosfoproteínas/química , Fosforilação , Estrutura Terciária de Proteína , RNA Polimerase II/química , RNA Polimerase II/imunologia
9.
Med Sci (Paris) ; 19(12): 1251-8, 2003 Dec.
Artigo em Francês | MEDLINE | ID: mdl-14691750

RESUMO

Peptidyl-prolyl isomerases (PPIases) are chaperone enzymes which alter the peptide bond between a given amino acid and a proline, changing it from the cis to the trans conformation and vice versa. This modification can cause dramatic structural modifications which can affect the properties of targeted proteins. The ubiquitous PPIase Pin1, conserved from yeast to human, has been shown to be necessary for entry into mitosis. The yeast homologue, Ess1, is essential for cell survival. Pin1 possesses a WW domain which specifically recognizes pSer-Pro and pThr-Pro motifs in which the first amino acid is phosphorylated. Pin1 binds to many proteins implicated in cell cycle regulation (e.g. p53, Myt1, Wee1, and Cdc25C). Pin1 also targets tau, a protein forming part of hte neuronal cytoskeleton which is hyper-phosphorylated in patients suffering from Alzheimer's disease (AD). Pin1 could, therefore, be involved in the pathogenesis of Ad. Furthermore, Pin1 also binds two proteins involved in transcription: Rpb1, the largest subunit of RNA polymerase II and Spt5, a regulator of the elongation of transcription. Both theses proteins possess domains rich in S/T-P motifs which can be targeted by Pin1 when phosphorylated. Recent studies show that Pin1 modulates the dephosphorylation of some proteins by allowing trans-specific phosphatases to recognize their target after isomerization. This unexpected role might allow protein regulation via peptidyl-prolyl isomerase activity.


Assuntos
Doença de Alzheimer/fisiopatologia , Sobrevivência Celular , Peptidilprolil Isomerase/farmacologia , Ciclo Celular/fisiologia , Humanos , Peptidilprolil Isomerase de Interação com NIMA , Peptidilprolil Isomerase/metabolismo , Fosforilação , Ligação Proteica , Transcrição Genética , Proteínas tau/metabolismo
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