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1.
J Virol ; 96(7): e0185721, 2022 04 13.
Artigo em Inglês | MEDLINE | ID: mdl-35285688

RESUMO

Human papillomaviruses (HPVs) are nonenveloped double-stranded DNA viruses that utilize heparan sulfate proteoglycans (HSPGs) as initial attachment factors prior to cell entry and infection. While extensively characterized, the selective interaction between HPV and HSPGs is generally studied using standard in vitro conditions, which fail to account for the effects that media additives, such as fetal bovine serum (FBS), can have on viral binding. As environmental conditions and growth factors associated with wound healing are thought to play a role in natural HPV infection, we sought to investigate the effects that serum or platelet extracts could have on the binding and infectivity of HPV. Here, we demonstrate that high concentrations of FBS and human serum greatly inhibit HPV16 binding, and that for FBS, this effect results from the obstruction of cell surface HSPGs by serum-derived heparin-binding proteins (HBPs). Surprisingly, we found that under these conditions, HPV particles utilize 6O-sulfated chondroitin sulfate proteoglycans (CSPGs) as initial binding receptors prior to infection. These findings were corroborated by small interfering RNA (siRNA)-mediated knockdown experiments, as well as through a cancer cell line screen, where we identified a strong association between viral binding in high serum and the expression of chondroitin sulfate biosynthesis genes. Furthermore, HPV binding in the presence of human platelet lysate also demonstrated an increased dependance on CSPGs, suggesting a possible role for these receptor proteoglycans in active wound healing environments. Overall, this work highlights the significant influence that serum/platelet factors can have on virus binding and identifies CSPGs as alternative cell attachment receptors for HPV. IMPORTANCE Heparan sulfate proteoglycans (HSPGs) have previously been identified as primary attachment factors for the initial binding of human papillomaviruses (HPVs) prior to infection. Here, we demonstrate that in vitro, HPV binding to HSPGs is strongly dependent on the surrounding experimental conditions, including the concentration of fetal bovine serum (FBS). We found that high concentrations of FBS can block HSPG-binding sites and cause a dependence on 6O-sulfated chondroitin sulfate proteoglycans (CSPGs) as alternative initial viral receptors. Further, we demonstrate that use of a human-derived alternative to FBS, human platelet lysate, also occludes HSPG-dependent binding, causing a shift toward CSPGs for viral attachment. As HPV infection of basal epithelial cells is thought to occur at sites of microtrauma with exposure to high serum levels and platelet factors, these unexpected findings highlight a possible role for CSPGs as important cellular receptors for the binding and infectivity of HPV in vivo.


Assuntos
Proteoglicanas de Sulfatos de Condroitina , Papillomavirus Humano 16 , Infecções por Papillomavirus , Linhagem Celular Tumoral , Proteoglicanas de Sulfatos de Condroitina/metabolismo , Proteoglicanas de Heparan Sulfato/metabolismo , Papillomavirus Humano 16/efeitos dos fármacos , Papillomavirus Humano 16/metabolismo , Humanos , Ligação Proteica , Soroalbumina Bovina/farmacologia
2.
J Cell Biol ; 218(10): 3212-3222, 2019 10 07.
Artigo em Inglês | MEDLINE | ID: mdl-31488583

RESUMO

Mutations associated with tumor development in certain tissues can be nontumorigenic in others, yet the mechanisms underlying these different outcomes remains poorly understood. To address this, we targeted an activating Hras mutation to hair follicle stem cells and discovered that Hras mutant cells outcompete wild-type neighbors yet are integrated into clinically normal skin hair follicles. In contrast, targeting the Hras mutation to the upper noncycling region of the skin epithelium leads to benign outgrowths. Follicular Hras mutant cells autonomously and nonautonomously enhance regeneration, which directs mutant cells into continuous tissue cycling to promote integration rather than aberrancy. This follicular tolerance is maintained under additional challenges that promote tumorigenesis in the epidermis, including aging, injury, and a secondary mutation. Thus, the hair follicle possesses a unique, enhanced capacity to integrate and contain Hras mutant cells within both homeostatic and perturbed tissue, demonstrating that in the skin, multiple, distinct mechanisms exist to suppress oncogenic growth.


Assuntos
Carcinogênese , Folículo Piloso/metabolismo , Neoplasias/metabolismo , Neoplasias/patologia , Regeneração , Proteínas ras/metabolismo , Animais , Camundongos , Camundongos Transgênicos
3.
Nat Commun ; 10(1): 3790, 2019 08 22.
Artigo em Inglês | MEDLINE | ID: mdl-31439867

RESUMO

Pediatric high-grade gliomas are among the deadliest of childhood cancers due to limited knowledge of early driving events in their gliomagenesis and the lack of effective therapies available. In this study, we investigate the oncogenic role of PPM1D, a protein phosphatase often found truncated in pediatric gliomas such as DIPG, and uncover a synthetic lethal interaction between PPM1D mutations and nicotinamide phosphoribosyltransferase (NAMPT) inhibition. Specifically, we show that mutant PPM1D drives hypermethylation of CpG islands throughout the genome and promotes epigenetic silencing of nicotinic acid phosphoribosyltransferase (NAPRT), a key gene involved in NAD biosynthesis. Notably, PPM1D mutant cells are shown to be sensitive to NAMPT inhibitors in vitro and in vivo, within both engineered isogenic astrocytes and primary patient-derived model systems, suggesting the possible application of NAMPT inhibitors for the treatment of pediatric gliomas. Overall, our results reveal a promising approach for the targeting of PPM1D mutant tumors, and define a critical link between oncogenic driver mutations and NAD metabolism, which can be exploited for tumor-specific cell killing.


Assuntos
Antineoplásicos/farmacologia , Neoplasias do Tronco Encefálico/genética , Glioma Pontino Intrínseco Difuso/genética , Nicotinamida Fosforribosiltransferase/genética , Proteína Fosfatase 2C/genética , Animais , Antineoplásicos/uso terapêutico , Neoplasias do Tronco Encefálico/tratamento farmacológico , Neoplasias do Tronco Encefálico/patologia , Linhagem Celular Tumoral , Criança , Citocinas/antagonistas & inibidores , Metilação de DNA , Glioma Pontino Intrínseco Difuso/tratamento farmacológico , Glioma Pontino Intrínseco Difuso/patologia , Repressão Epigenética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Camundongos , Nicotinamida Fosforribosiltransferase/antagonistas & inibidores , Nicotinamida Fosforribosiltransferase/metabolismo , Ponte/citologia , Ponte/patologia , Cultura Primária de Células , Proteína Fosfatase 2C/metabolismo , Mutações Sintéticas Letais , Ensaios Antitumorais Modelo de Xenoenxerto
4.
Sci Transl Med ; 9(375)2017 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-28148839

RESUMO

2-Hydroxyglutarate (2HG) exists as two enantiomers, (R)-2HG and (S)-2HG, and both are implicated in tumor progression via their inhibitory effects on α-ketoglutarate (αKG)-dependent dioxygenases. The former is an oncometabolite that is induced by the neomorphic activity conferred by isocitrate dehydrogenase 1 (IDH1) and IDH2 mutations, whereas the latter is produced under pathologic processes such as hypoxia. We report that IDH1/2 mutations induce a homologous recombination (HR) defect that renders tumor cells exquisitely sensitive to poly(adenosine 5'-diphosphate-ribose) polymerase (PARP) inhibitors. This "BRCAness" phenotype of IDH mutant cells can be completely reversed by treatment with small-molecule inhibitors of the mutant IDH1 enzyme, and conversely, it can be entirely recapitulated by treatment with either of the 2HG enantiomers in cells with intact IDH1/2 proteins. We demonstrate mutant IDH1-dependent PARP inhibitor sensitivity in a range of clinically relevant models, including primary patient-derived glioma cells in culture and genetically matched tumor xenografts in vivo. These findings provide the basis for a possible therapeutic strategy exploiting the biological consequences of mutant IDH, rather than attempting to block 2HG production, by targeting the 2HG-dependent HR deficiency with PARP inhibition. Furthermore, our results uncover an unexpected link between oncometabolites, altered DNA repair, and genetic instability.


Assuntos
Glioma/tratamento farmacológico , Glutaratos/farmacologia , Recombinação Homóloga , Isocitrato Desidrogenase/genética , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Animais , Linhagem Celular Tumoral , Quebras de DNA de Cadeia Dupla , Reparo do DNA , Feminino , Glioma/genética , Humanos , Isocitrato Desidrogenase/farmacologia , Camundongos Nus , Ensaios Antitumorais Modelo de Xenoenxerto
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