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1.
BMC Cancer ; 24(1): 1129, 2024 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-39256694

RESUMO

BACKGROUND: Breast cancer metastasis remains the leading cause of cancer-related deaths in women worldwide. Infiltration of tumor-associated macrophages (TAMs) in the tumor stroma is known to be correlated with reduced overall survival. The inhibitors of TAMs are sought after for reprogramming the tumor microenvironment. Signal transducer and activator of transcription 3 (STAT3) is well known to contribute in pro-tumoral properties of TAMs. 2-Methoxyestradiol (2ME2), a potent anticancer and antiangiogenic agent, has been in clinical trials for treatment of breast cancer. Here, we investigated the potential of 2ME2 in modulating the pro-tumoral effects of TAMs in breast cancer. METHODS: THP-1-derived macrophages were polarized to macrophages with or without 2ME2. The effect of 2ME2 on macrophage surface markers and anti-inflammatory genes was determined by Western blotting, flow cytometry, immunofluorescence, qRT‒PCR. The concentration of cytokines secreted by cells was monitored by ELISA. The effect of M2 macrophages on malignant properties of breast cancer cells was determined using colony formation, wound healing, transwell, and gelatin zymography assays. An orthotopic model of breast cancer was used to determine the effect of 2ME2 on macrophage polarization and metastasis in vivo. RESULTS: First, our study found that polarization of monocytes to alternatively activated M2 macrophages is associated with the reorganization of the microtubule cytoskeleton. At lower concentrations, 2ME2 treatment depolymerized microtubules and reduced the expression of CD206 and CD163, suggesting that it inhibits the polarization of macrophages to M2 phenotype. However, the M1 polarization was not significantly affected at these concentrations. Importantly, 2ME2 inhibited the expression of several anti-inflammatory cytokines and growth factors, including CCL18, TGF-ß, IL-10, FNT, arginase, CXCL12, MMP9, and VEGF-A, and hindered the metastasis-promoting effects of M2 macrophages. Concurrently, 2ME2 treatment reduced the expression of CD163 in tumors and inhibited lung metastasis in the orthotopic breast cancer model. Mechanistically, 2ME2 treatment reduced the phosphorylation and nuclear translocation of STAT3, an effect which was abrogated by colivelin. CONCLUSIONS: Our study presents novel findings on mechanism of 2ME2 from the perspective of its effects on the polarization of the TAMs via the STAT3 signaling in breast cancer. Altogether, the data supports further clinical investigation of 2ME2 and its derivatives as therapeutic agents to modulate the tumor microenvironment and immune response in breast carcinoma.


Assuntos
2-Metoxiestradiol , Neoplasias da Mama , Fator de Transcrição STAT3 , Macrófagos Associados a Tumor , 2-Metoxiestradiol/farmacologia , Humanos , Feminino , Fator de Transcrição STAT3/metabolismo , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Neoplasias da Mama/metabolismo , Camundongos , Animais , Macrófagos Associados a Tumor/efeitos dos fármacos , Macrófagos Associados a Tumor/metabolismo , Macrófagos Associados a Tumor/imunologia , Microambiente Tumoral/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Macrófagos/imunologia , Linhagem Celular Tumoral , Ativação de Macrófagos/efeitos dos fármacos , Células THP-1 , Ensaios Antitumorais Modelo de Xenoenxerto , Citocinas/metabolismo
2.
Chem Commun (Camb) ; 60(64): 8419-8422, 2024 Aug 06.
Artigo em Inglês | MEDLINE | ID: mdl-39028297

RESUMO

Light and pH dual-responsive ion transporters offer better applicability for cancer due to higher tunability and low cytotoxicity. Herein, we demonstrate the development of pH-responsive ß-carboline-based ionophores and photocleavable-linker appended ß-carboline-based proionophores to facilitate the controlled transport of Cl- across membranes, leading to apoptotic and autophagic cancer cell death.


Assuntos
Carbolinas , Luz , Carbolinas/química , Carbolinas/farmacologia , Humanos , Concentração de Íons de Hidrogênio , Apoptose/efeitos dos fármacos , Antineoplásicos/farmacologia , Antineoplásicos/química , Transporte de Íons/efeitos dos fármacos , Linhagem Celular Tumoral , Estrutura Molecular , Ionóforos/química , Ionóforos/farmacologia , Ensaios de Seleção de Medicamentos Antitumorais
3.
Biochim Biophys Acta Gene Regul Mech ; 1867(3): 195044, 2024 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-38763317

RESUMO

Maintenance of genome integrity is a precise but tedious and complex job for the cell. Several post-translational modifications (PTMs) play vital roles in maintaining the genome integrity. Although ubiquitination is one of the most crucial PTMs, which regulates the localization and stability of the nonhistone proteins in various cellular and developmental processes, ubiquitination of the histones is a pivotal epigenetic event critically regulating chromatin architecture. In addition to genome integrity, importance of ubiquitination of core histones (H2A, H2A, H3, and H4) and linker histone (H1) have been reported in several cellular processes. However, the complex interplay of histone ubiquitination and other PTMs, as well as the intricate chromatin architecture and dynamics, pose a significant challenge to unravel how histone ubiquitination safeguards genome stability. Therefore, further studies are needed to elucidate the interactions between histone ubiquitination and other PTMs, and their role in preserving genome integrity. Here, we review all types of histone ubiquitinations known till date in maintaining genomic integrity during transcription, replication, cell cycle, and DNA damage response processes. In addition, we have also discussed the role of histone ubiquitination in regulating other histone PTMs emphasizing methylation and acetylation as well as their potential implications in chromatin architecture. Further, we have also discussed the involvement of deubiquitination enzymes (DUBs) in controlling histone ubiquitination in modulating cellular processes.


Assuntos
Cromatina , Instabilidade Genômica , Histonas , Processamento de Proteína Pós-Traducional , Ubiquitinação , Histonas/metabolismo , Humanos , Cromatina/metabolismo , Animais , Dano ao DNA , Epigênese Genética
4.
Cell Death Dis ; 14(9): 623, 2023 09 22.
Artigo em Inglês | MEDLINE | ID: mdl-37736741

RESUMO

Oncogene Moesin plays critical role in initiation, progression, and metastasis of multiple cancers. It exerts oncogenic activity due to its high-level expression as well as posttranslational modification in cancer. However, factors responsible for its high-level expression remain elusive. In this study, we identified positive as well as negative regulators of Moesin. Our study reveals that Moesin is a cellular target of F-box protein FBXW2. We showed that FBXW2 suppresses breast cancer progression through directing proteasomal degradation of Moesin. In contrast, AKT kinase plays an important role in oncogenic function of Moesin by protecting it from FBXW2-mediated proteasomal degradation. Mechanistically, AKT phosphorylates Moesin at Thr-558 and thereby prevents its degradation by FBXW2 via weakening the association between FBXW2 and Moesin. Further, accumulated Moesin prevents FBXW2-mediated degradation of oncogene SKP2, showing that Moesin functions as an upstream regulator of oncogene SKP2. In turn, SKP2 stabilizes Moesin by directing its non-degradable form of polyubiquitination and therefore AKT-Moesin-SKP2 oncogenic axis plays crucial role in breast cancer progression. Collectively, our study reveals that FBXW2 functions as a tumor suppressor in breast cancer by restricting AKT-Moesin-SKP2 axis. Thus, AKT-Moesin-SKP2 axis may be explored for the development of therapeutics for cancer treatment.


Assuntos
Neoplasias da Mama , Proteínas F-Box , Proteínas Proto-Oncogênicas c-akt , Humanos , Transformação Celular Neoplásica , Proteínas F-Box/genética , Proteínas dos Microfilamentos , Oncogenes , Neoplasias da Mama/genética , Neoplasias da Mama/patologia
5.
J Med Chem ; 66(16): 11078-11093, 2023 08 24.
Artigo em Inglês | MEDLINE | ID: mdl-37466499

RESUMO

The increasing resistance of bacteria to commercially available antibiotics threatens patient safety in healthcare settings. Perturbation of ion homeostasis has emerged as a potential therapeutic strategy to fight against antibacterial resistance and other channelopathies. This study reports the development of 8-aminoquinoline (QN) derivatives and their transmembrane Zn2+ transport activities. Our findings showed that a potent QN-based Zn2+ transporter exhibits promising antibacterial properties against Gram-positive bacteria with reduced hemolytic activity and cytotoxicity to mammalian cells. Furthermore, this combination showed excellent in vivo efficacy against Staphylococcus aureus. Interestingly, this combination prevented bacterial resistance and restored susceptibility of gentamicin and methicillin-resistant S. aureus to commercially available ß-lactam and other antibiotics that had lost their activity against the drug-resistant bacterial strain. Our findings suggest that the transmembrane transport of Zn2+ by QN derivatives could be a promising strategy to combat bacterial infections and restore the activity of other antibiotics.


Assuntos
Staphylococcus aureus Resistente à Meticilina , Quinolinas , Infecções Estafilocócicas , Animais , Humanos , Zinco , Ionóforos/uso terapêutico , Tioureia/farmacologia , Tioureia/uso terapêutico , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Infecções Estafilocócicas/tratamento farmacológico , Quinolinas/farmacologia , Quinolinas/uso terapêutico , Testes de Sensibilidade Microbiana , Mamíferos
6.
Immunology ; 170(1): 60-82, 2023 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-37185810

RESUMO

Nutritional availability during fasting and refeeding affects the temporal redistribution of lymphoid and myeloid immune cells among the circulating and tissue-resident pools. Conversely, nutritional imbalance and impaired glucose metabolism are associated with chronic inflammation, aberrant immunity and anomalous leukocyte trafficking. Despite being exposed to periodic alterations in blood insulin levels upon fasting and feeding, studies exploring the physiological effects of these hormonal changes on quiescent immune cell function and trafficking are scanty. Here, we report that oral glucose load in mice and healthy men enhances the adherence of circulating peripheral blood mononuclear cells (PBMCs) and lymphocytes to fibronectin. Adherence to fibronectin is also observed upon regular intake of breakfast following overnight fasting in healthy subjects. This glucose load-induced phenomenon is abrogated in streptozotocin-injected mice that lack insulin. Intra-vital microscopy in mice demonstrated that oral glucose feeding enhances the homing of PBMCs to injured blood vessels in vivo. Furthermore, employing flow cytometry, Western blotting and adhesion assays for PBMCs and Jurkat-T cells, we elucidate that insulin enhances fibronectin adherence of quiescent lymphocytes through non-canonical signalling involving insulin-like growth factor-1 receptor (IGF-1R) autophosphorylation, phospholipase C gamma-1 (PLCγ-1) Tyr783 phosphorylation and inside-out activation of ß-integrins respectively. Our findings uncover the physiological relevance of post-prandial insulin spikes in regulating the adherence and trafficking of circulating quiescent T-cells through fibronectin-integrin interaction.

7.
Inorg Chem ; 62(18): 7003-7013, 2023 May 08.
Artigo em Inglês | MEDLINE | ID: mdl-37097171

RESUMO

Herein, we present an unprecedented formation of a heterodinuclear complex [{(ppy)2IrIII}(µ-phpy){RuII(tpy)}](ClO4)2 {[1](ClO4)2} using terpyridyl/phenylpyridine as ancillary ligands and asymmetric phpy as a bridging ligand. The asymmetric binding mode (N∧N-∩-N∧N∧C-) of the phpy ligand in {[1](ClO4)2} is confirmed by 1H, 13C, 1H-1H correlated spectroscopy (COSY), high-resolution mass spectrum (HRMS), single-crystal X-ray crystallography techniques, and solution conductivity measurements. Theoretical investigation suggests that the highest occupied molecular orbital (HOMO) and the least unoccupied molecular orbital (LUMO) of [1]2+ are located on iridium/ppy and phpy, respectively. The complex displays a broad low energy charge transfer (CT) band within 450-575 nm. The time-dependent density functional theory (TDDFT) analysis suggests this as a mixture of metal-to-ligand charge transfer (MLCT) and ligand-to-ligand charge transfer (LLCT), where both ruthenium, iridium, and ligands are involved. Complex {[1](ClO4)2} exhibits RuIIIrIII/RuIIIIrIII- and RuIIIIrIII/RuIIIIrIV-based oxidative couples at 0.83 and 1.39 V, respectively. The complex shows anticancer activity and selectivity toward human breast cancer cells (IC50; MCF-7: 9.3 ± 1.2 µM, and MDA-MB-231: 8.6 ± 1.2 µM) over normal breast cells (MCF 10A: IC50 ≈ 21 ± 1.3 µM). The Western blot analysis and fluorescence microscopy images suggest that combined apoptosis and autophagy are responsible for cancer cell death.


Assuntos
Compostos Organometálicos , Humanos , Estrutura Molecular , Compostos Organometálicos/química , Ligantes , Irídio/farmacologia , Irídio/química , Análise Espectral
8.
Sci Adv ; 9(12): eade1851, 2023 03 22.
Artigo em Inglês | MEDLINE | ID: mdl-36947610

RESUMO

Sensing of pathogens by ubiquitination is a critical arm of cellular immunity. However, universal ubiquitination targets on microbes remain unidentified. Here, using in vitro, ex vivo, and in vivo studies, we identify the first protein-based ubiquitination substrates on phylogenetically diverse bacteria by unveiling a strategy that uses recognition of degron-like motifs. Such motifs form a new class of intra-cytosolic pathogen-associated molecular patterns (PAMPs). Their incorporation enabled recognition of nonubiquitin targets by host ubiquitin ligases. We find that SCFFBW7 E3 ligase, supported by the regulatory kinase, glycogen synthase kinase 3ß, is crucial for effective pathogen detection and clearance. This provides a mechanistic explanation for enhanced risk of infections in patients with chronic lymphocytic leukemia bearing mutations in F-box and WD repeat domain containing 7 protein. We conclude that exploitation of this generic pathogen sensing strategy allows conservation of host resources and boosts antimicrobial immunity.


Assuntos
Proteínas F-Box , Humanos , Proteína 7 com Repetições F-Box-WD/genética , Proteína 7 com Repetições F-Box-WD/metabolismo , Proteínas F-Box/genética , Proteínas F-Box/metabolismo , Proteínas de Ciclo Celular/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Proteínas de Membrana/metabolismo , Fosforilação , Ubiquitinação , Bactérias/metabolismo
9.
Chemistry ; 29(10): e202202887, 2023 Feb 16.
Artigo em Inglês | MEDLINE | ID: mdl-36399427

RESUMO

Artificial channels capable of facilitating the transport of Cl- ions across cell membranes while being nontoxic to the cells are rare. Such synthetic ion channels can mimic the functions of membrane transport proteins and, therefore, have the potential to treat channelopathies by replacing defective ion channels. Here we report isophthalic acid-based structurally simple molecules 1 a and 2 a, which self-assemble to render supramolecular nanochannels that allow selective transport of Cl- ions. As evident from the single-crystal X-ray diffraction analysis, the self-assembly is governed by intermolecular hydrogen bonding and π-π stacking interactions. The MD simulation studies for both 1 a and 2 a confirmed the formation of stable Cl- channel assembly in the lipid membrane and Cl- transport through them. The MQAE assay showed the efficacy of the compounds in delivering Cl- ions into cells, and the MTT assays proved that the compounds are nontoxic to cells even at a concentration of 100 µM.


Assuntos
Canais de Cloreto , Ácidos Ftálicos , Canais Iônicos/química , Células Epiteliais
10.
Sci Rep ; 12(1): 17070, 2022 10 12.
Artigo em Inglês | MEDLINE | ID: mdl-36224238

RESUMO

Prostate cancer (PCa) progresses from a hormone-sensitive, androgen-dependent to a hormone-refractory, androgen-independent metastatic phenotype. Among the many genes implicated, ANXA2, a calcium-dependent phospholipid binding protein, has been found to have a critical role in the progression of PCa into more invasive metastatic phenotype. However, the molecular mechanisms underlying the absence of ANXA2 in early PCa and its recurrence in advanced stage are yet unknown. Moreover, recent studies have observed the deregulation of microRNAs (miRNAs) are involved in the development and progression of PCa. In this study, we found the down-regulation of miR-936 in metastatic PCa wherein its target ANXA2 was overexpressed. Subsequently, it has been shown that the downregulation of miRNA biogenesis by siRNA treatment in ANXA2-null LNCaP cells could induce the expression of ANXA2, indicating the miRNA mediated regulation of ANXA2 expression. Additionally, we demonstrate that miR-936 regulates ANXA2 expression by direct interaction at coding as well as 3'UTR region of ANXA2 mRNA by luciferase reporter assay. Furthermore, the overexpression of miR-936 suppresses the cell proliferation, cell cycle progression, cell migration, and invasion abilities of metastatic PCa PC-3 cells in vitro and tumor forming ability in vivo. These results indicate that miR-936 have tumor suppressor properties by regulating the over expression of ANXA2 in hormone-independent metastatic PCa. Moreover, our results suggest that this tumor suppressor miR-936 could be developed as a targeted therapeutic molecule for metastatic PCa control and to improve the prognosis in PCa patients.


Assuntos
MicroRNAs , Neoplasias da Próstata , Regiões 3' não Traduzidas , Androgênios , Cálcio/metabolismo , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , MicroRNAs/genética , MicroRNAs/metabolismo , Fenótipo , Fosfolipídeos , Neoplasias da Próstata/patologia , RNA Interferente Pequeno
11.
Subcell Biochem ; 100: 427-472, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36301502

RESUMO

Cancer begins due to uncontrolled cell division. Cancer cells are insensitive to the signals that control normal cell proliferation. This uncontrolled cell division is due to the accumulation of abnormalities in different factors associated with the cell division, including different cyclins, cell cycle checkpoint inhibitors, and cellular signaling. Cellular signaling pathways are aberrantly activated in cancer mainly due to epigenetic regulation and post-translational regulation. In this chapter, the role of epigenetic regulation in aberrant activation of PI3K/AKT, Ras, Wnt, Hedgehog, Notch, JAK/STAT, and mTOR signaling pathways in cancer progression is discussed. The role of epigenetic regulators in controlling the upstream regulatory proteins and downstream effector proteins responsible for abnormal cellular signaling-mediated cancer progression is covered in this chapter. Similarly, the role of signaling pathways in controlling epigenetic gene regulation-mediated cancer progression is also discussed. We have tried to ascertain the current status of potential epigenetic drugs targeting several epigenetic regulators to prevent different cancers.


Assuntos
Epigênese Genética , Neoplasias , Transdução de Sinais , Humanos , Neoplasias/metabolismo , Regulação Neoplásica da Expressão Gênica
12.
Biochim Biophys Acta Rev Cancer ; 1877(4): 188753, 2022 07.
Artigo em Inglês | MEDLINE | ID: mdl-35752404

RESUMO

Cancer metastasis is the primary cause of morbidity and mortality in cancer as it remains the most complicated, devastating, and enigmatic aspect of cancer. Several decades of extensive research have identified several key players closely associated with metastasis. Among these players, cytoskeletal linker Ezrin (the founding member of the ERM (Ezrin-Radixin-Moesin) family) was identified as a critical promoter of metastasis in pediatric cancers in the early 21st century. Ezrin was discovered 40 years ago as a aminor component of intestinal epithelial microvillus core protein, which is enriched in actin-containing cell surface structures. It controls gastric acid secretion and plays diverse physiological roles including maintaining cell polarity, regulating cell adhesion, cell motility and morphogenesis. Extensive research for more than two decades evinces that Ezrin is frequently dysregulated in several human cancers. Overexpression, altered subcellular localization and/or aberrant activation of Ezrin are closely associated with higher metastatic incidence and patient mortality, thereby justifying Ezrin as a valuable prognostic biomarker in cancer. Ezrin plays multifaceted role in multiple aspects of cancer, with its significant contribution in the complex metastatic cascade, through reorganizing the cytoskeleton and deregulating various cellular signaling pathways. Current preclinical studies using genetic and/or pharmacological approaches reveal that inactivation of Ezrin results in significant inhibition of Ezrin-mediated tumor growth and metastasis as well as increase in the sensitivity of cancer cells to various chemotherapeutic drugs. In this review, we discuss the recent advances illuminating the molecular mechanisms responsible for Ezrin dysregulation in cancer and its pleiotropic role in cancer progression and metastasis. We also highlight its potential as a prognostic biomarker and therapeutic target in various cancers. More importantly, we put forward some potential questions, which we strongly believe, will stimulate both basic and translational research to better understand Ezrin-mediated malignancy, ultimately leading to the development of Ezrin-targeted cancer therapy for the betterment of human life.


Assuntos
Neoplasias , Actinas , Biomarcadores/metabolismo , Criança , Proteínas do Citoesqueleto , Citoesqueleto/metabolismo , Humanos , Neoplasias/tratamento farmacológico , Neoplasias/genética , Neoplasias/metabolismo
13.
Int J Biochem Cell Biol ; 147: 106228, 2022 06.
Artigo em Inglês | MEDLINE | ID: mdl-35598880

RESUMO

F-box proteins form SCF (Cullin1, SKP1 and F-box-protein) ubiquitin ligase complexes to ubiquitinate cellular proteins. They play key role in several biological processes, including cell cycle progression, cellular signaling, stress response and cell death pathways. Therefore, deregulation of F-box proteins is closely associated with cancer progression. However, the role of most of the F-box proteins, including FBXO41, in cancer progression remains elusive. Here, we unravel the role of FBXO41 in cancer progression. We show that FBXO41 suppresses cancer cell proliferation and tumor growth by inducing autophagic cell death through an alternative pathway. Results revealed that FBXO41-mediated autophagic cell death induction is dependent on accumulation of cell cycle checkpoint protein p21. We found that FBXO41 increases the expression levels of p21 at the post-translational level by promoting the proteasomal degradation of SKP2, an oncogenic F-box protein. Mechanistically, FBXO41 along with p21 disrupts the inhibitory BCL2 (anti-apoptotic protein)-Beclin1 (autophagy initiating factor) complex of autophagy induction to release Beclin1, thereby inducing autophagy. Overall, the present study establishes a new FBXO41-SKP2-p21 axis for induction of autophagic cell death to prevent cancer growth, which could be explored to develop promising cancer therapeutics.


Assuntos
Morte Celular Autofágica , Fenômenos Biológicos , Neoplasias da Mama , Proteínas F-Box/metabolismo , Proteínas Quinases Associadas a Fase S/metabolismo , Proteína Beclina-1/metabolismo , Neoplasias da Mama/genética , Proteínas de Ciclo Celular/genética , Proteínas Culina/genética , Proteínas F-Box/genética , Feminino , Humanos , Oncogenes , Proteínas Quinases Associadas a Fase S/genética , Proteínas Ligases SKP Culina F-Box/genética , Proteínas Ligases SKP Culina F-Box/metabolismo
14.
Toxicol Appl Pharmacol ; 441: 115973, 2022 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-35278439

RESUMO

Arsenic trioxide (ATO), a potent anti-neoplastic drug, is known to prevent cancer cell growth through induction of autophagic cell death. However, importance of cellular factors in ATO-mediated autophagic cell death is poorly understood. In this study, using biochemical and immunofluorescence techniques, we show that F-box protein FBXO41 plays a critical role in anti-proliferative activity of ATO. Our study reveals the importance of FBXO41 in induction of autophagic death of cancer cells by ATO. Further, we show that the autophagic cell death induced by FBXO41 is distinct and independent of apoptosis and necrosis, showing that FBXO41 may play vital role in inducing autophagic death of apoptosis resistant cancer cells. Overall, our study elucidates the importance of FBXO41 in ATO induced autophagic cell death to prevent cancer progression, which could be explored to develop promising cancer therapeutic strategy.


Assuntos
Antineoplásicos , Arsenicais , Morte Celular Autofágica , Proteínas F-Box , Neoplasias , Antineoplásicos/farmacologia , Apoptose , Trióxido de Arsênio/farmacologia , Arsenicais/farmacologia , Linhagem Celular Tumoral , Humanos , Neoplasias/tratamento farmacológico , Óxidos/farmacologia
15.
Int J Cancer ; 150(9): 1512-1524, 2022 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-34706096

RESUMO

FBXO31, a member of F-box protein family, has been shown to play an important role in preventing tumorigenesis by preserving genomic stability during cell proliferation as well as upon genotoxic stress. Inactivation of FBXO31 due to loss of heterozygosity is associated with various cancers, including ovarian cancer, one of the deadliest forms of gynecological cancers. However, the role and regulation of FBXO31 in ovarian cancer remained elusive. Here, using biochemical and molecular biology techniques, we show that c-Myc suppresses the mRNA levels of FBXO31 in ovarian cancer. Chromatin immunoprecipitation experiment showed that c-Myc is recruited to the promoter region of FBXO31 and prevents FBXO31 mRNA synthesis. In contrast, FBXO31 maintains the c-Myc expression at an optimum through proteasome pathway. FBXO31 interacts with and facilitates the polyubiquitination of c-Myc through the SCF complex and thereby inhibits ovarian cancer growth both in vitro and in vivo. Moreover, FBXO31-mediated proteasomal degradation of c-Myc is unique. Unlike other negative regulators, FBXO31 recognizes c-Myc in phosphorylation independent manner to direct its degradation. Further, expression levels analysis revealed that c-Myc and FBXO31 share a converse correlation of expression in ovarian cancer cell lines and patient samples. We observed an increase in the expression levels of c-Myc with a concomitant decrease in the levels of FBXO31 in higher grades of ovarian cancer patient samples. In conclusion, our study demonstrated that oncogene c-Myc impairs the tumor-suppressive functions of FBXO31 to promote ovarian cancer progression, and therefore c-Myc-FBXO31 axis can be explored to develop better cancer therapy.


Assuntos
Proteínas F-Box , Neoplasias Ovarianas , Proteínas Supressoras de Tumor , Carcinogênese/genética , Linhagem Celular Tumoral , Proteínas F-Box/genética , Retroalimentação , Feminino , Humanos , Neoplasias Ovarianas/genética , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , RNA Mensageiro , Proteínas Supressoras de Tumor/genética
16.
DNA Repair (Amst) ; 109: 103261, 2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-34920250

RESUMO

The maintenance of genomic integrity is of utmost importance for the organisms to survive and to accurately inherit traits to their progenies. Any kind of DNA damage either due to defect in DNA duplication and/ or uncontrolled cell division or intracellular insults or environment radiation can result in gene mutation, chromosomal aberration and ultimately genomic instability, which may cause several diseases including cancers. Therefore, cells have evolved machineries for the surveillance of genomic integrity. Enormous exciting studies in the past indicate that ubiquitination (a posttranslational modification of proteins) plays a crucial role in maintaining the genomic integrity by diverse ways. In fact, various E3 ubiquitin ligases catalyse ubiquitination of key proteins to control their central role during cell cycle, DNA damage response (DDR) and DNA repair. Some E3 ligases promote genomic instability while others prevent it, deregulation of both of which leads to several malignancies. In this review, we consolidate the recent findings wherein the role of ubiquitination in conferring genome integrity is highlighted. We also discuss the latest discoveries on the mechanisms utilized by various E3 ligases to preserve genomic stability, with a focus on their actions during cell cycle progression and different types of DNA damage response as well as repair pathways.


Assuntos
Reparo do DNA , Instabilidade Genômica , Ubiquitina-Proteína Ligases/metabolismo , Animais , Ciclo Celular , Dano ao DNA , Humanos
17.
Int J Biol Macromol ; 190: 233-243, 2021 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-34478796

RESUMO

F-box protein FBXW8 is known to interact with scaffolding protein Cullin1 and Cullin7 to form SCF (SKP1, Cullin and F-box protein) complex. However, detail understanding about the importance of both Cullins for SCF-FBXW8 complex formation as well as its ubiquitin ligase activity remains elusive. Here, we show that, through in vitro and in vivo studies, Cullin1 and Cullin7 increase each other's binding to FBXW8 synergistically. Interestingly, absence of either Cullin results in abrogation of binding of other Cullin to FBXW8. Binding of SKP1 to FBXW8 also increases in the presence of both the Cullins. Thus, SKP1, Cullin1 and Cullin7 are essential to form Cullin1-SKP1-FBXW8-Cullin7 functional ubiquitin ligase complex. Further, using computational, mutational and biochemical analysis, we found that Cullin1 binds to N-terminus of FBXW8 through SKP1 while Cullin7 associates with C-terminus of FBXW8 to form Cullin1-SKP1-FBXW8-Cullin7 functional complex in a cooperative manner. Results showed that Cullin1-SKP1-FBXW8-Cullin7 complex plays a key role in maintaining the basal level expression of ß-TrCP1. Moreover, Cullin1-SKP1-FBXW8-Cullin7 complex promotes cell migration by activating ß-catenin via directing proteasomal degradation of ß-TrCP1. Overall, our study reveals the intriguing molecular mechanism of assembly of SKP1, Cullin1, Cullin7 and FBXW8 to form Cullin1-SKP1-FBXW8-Cullin7 functional complex that control the function of ß-TrCP1.


Assuntos
Proteínas Culina/metabolismo , Proteínas F-Box/metabolismo , Complexos Multiproteicos/metabolismo , Proteínas Quinases Associadas a Fase S/metabolismo , Proteínas Contendo Repetições de beta-Transducina/metabolismo , Movimento Celular , Proteínas Culina/química , Proteínas F-Box/química , Humanos , Células MCF-7 , Ligação Proteica , Estabilidade Proteica , Proteólise , Proteínas Quinases Associadas a Fase S/química , Especificidade por Substrato , beta Catenina/metabolismo , Proteínas Contendo Repetições de beta-Transducina/química
18.
J Biol Chem ; 297(4): 101253, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34587475

RESUMO

Apoptosis is a programmed cell death that efficiently removes damaged cells to maintain tissue homeostasis. Defect in apoptotic machinery can lead to tumor development, progression, and resistance to chemotherapy. PUMA (p53 upregulated modulator of apoptosis) and BAX (BCL2-associated X protein) are among the most well-known inducers of apoptosis. It has been reported that expression levels of BAX and PUMA are controlled at the posttranslational level by phosphorylation. However, the posttranslational regulation of these proapoptotic proteins remains largely unexplored. In this study, using biochemical, molecular biology, flow cytometric, and immunohistochemistry techniques, we show that PUMA and BAX are the direct target of the F-box protein FBXL20, which restricts their cellular levels. FBXL20 directs the proteasomal degradation of PUMA and BAX in a protein kinase AKT1-dependent manner to promote cancer cell proliferation and tumor growth. Interestingly, inactivation of AKT1 results in activation of another protein kinase GSK3α/ß, which facilitates the proteasomal degradation of FBXL20 by another F-box protein, FBXO31. Thus, a switch between two signaling kinases AKT1 and GSK3α/ß modulates the functional activity of these proapoptotic regulators, thereby determining cell survival or death. RNAi-mediated ablation of FBXL20 results in increased levels of PUMA as well as BAX, which further enhances the sensitivity of cancer cells to chemotherapeutic drugs. We showed that high level expression of FBXL20 in cancer cells reduces therapeutic drug-induced apoptosis and promotes chemoresistance. Overall, this study highlights the importance of targeting FBXL20 in cancers in conjunction with chemotherapy and may represent a promising anticancer strategy to overcome chemoresistance.


Assuntos
Proteínas Reguladoras de Apoptose/metabolismo , Apoptose , Neoplasias da Mama/metabolismo , Proteínas F-Box/metabolismo , Proteólise , Proteínas Proto-Oncogênicas/metabolismo , Proteína X Associada a bcl-2/metabolismo , Proteínas Reguladoras de Apoptose/genética , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Resistencia a Medicamentos Antineoplásicos/genética , Proteínas F-Box/genética , Feminino , Células HEK293 , Humanos , Células MCF-7 , Proteínas Proto-Oncogênicas/genética , Proteína X Associada a bcl-2/genética
19.
FEBS J ; 288(18): 5474-5497, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-33742524

RESUMO

Sequential alteration in the expression levels of cell cycle regulatory proteins is crucial for faithful cell cycle progression to maintain the cellular homeostasis. F-box protein ß-TrCP1 is known to control the expression levels of several important cell cycle regulatory proteins. However, how the function of ß-TrCP1 is regulated in spatiotemporal manner during cell cycle progression remains elusive. Here, we show that expression levels of ß-TrCP1 oscillate during cell cycle progression with a minimum level at the G1 and S phases of cell cycle. Using biochemical, flow cytometry, and immunofluorescence techniques, we found that oscillation of ß-TrCP1 expression is controlled by another F-box protein FBXW8. FBXW8 directs the proteasomal degradation of ß-TrCP1 in MAPK pathway-dependent manner. Interestingly, we found that the attenuation of ß-TrCP1 by FBXW8 is important for Cdc25A-mediated cell cycle transition from G1 phase to S phase as well as DNA damage-free progression of S phase. Overall, our study reveals the intriguing molecular mechanism and significance of maintenance of ß-TrCP1 levels during cell cycle progression by FBXW8-mediated proteasomal degradation.


Assuntos
Proteínas de Ciclo Celular/genética , Proteínas F-Box/genética , Proteínas Contendo Repetições de beta-Transducina/genética , Fosfatases cdc25/genética , Divisão Celular/genética , Dano ao DNA/genética , Citometria de Fluxo , Fase G1/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Células HEK293 , Humanos , Células MCF-7 , Complexo de Endopeptidases do Proteassoma/genética , Proteólise , Fase S/genética , Ubiquitina/genética
20.
J Biol Chem ; 296: 100511, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33676897

RESUMO

F-box proteins ß-TrCP1 and ß-TrCP2 are paralogs present in the human genome. They control several cellular processes including cell cycle and DNA damage signaling. Moreover, it is reported that they facilitate DNA damage-induced accumulation of p53 by directing proteasomal degradation of MDM2, a protein that promotes p53 degradation. However, the individual roles of ß-TrCP1 and ß-TrCP2 in the genotoxic stress-induced activation of cell cycle checkpoints and DNA damage repair remain largely unknown. Here, using biochemical, molecular biology, flow cytometric, and immunofluorescence techniques, we show that ß-TrCP1 and ß-TrCP2 communicate during genotoxic stress. We found that expression levels of ß-TrCP1 are significantly increased while levels of ß-TrCP2 are markedly decreased upon induction of genotoxic stress. Further, our results revealed that DNA damage-induced activation of ATM kinase plays an important role in maintaining the reciprocal expression levels of ß-TrCP1 and ß-TrCP2 via the phosphorylation of ß-TrCP1 at Ser158. Phosphorylated ß-TrCP1 potently promotes the proteasomal degradation of ß-TrCP2 and MDM2, resulting in the activation of p53. Additionally, ß-TrCP1 impedes MDM2 accumulation via abrogation of its lysine 63-linked polyubiquitination by ß-TrCP2. Thus, ß-TrCP1 helps to arrest cells at the G2/M phase of the cell cycle and promotes DNA repair upon DNA damage through attenuation of ß-TrCP2. Collectively, our findings elucidate an intriguing posttranslational regulatory mechanism of these two paralogs under genotoxic stress and revealed ß-TrCP1 as a key player in maintaining the genome integrity through the attenuation of ß-TrCP2 levels in response to genotoxic stress.


Assuntos
Pontos de Checagem do Ciclo Celular , Dano ao DNA , Reparo do DNA , Proteólise , Ubiquitinação , Proteínas Contendo Repetições de beta-Transducina/metabolismo , Sobrevivência Celular , Humanos , Fosforilação , Transdução de Sinais , Proteínas Contendo Repetições de beta-Transducina/genética
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