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BACKGROUND AND AIMS: Antimicrobial proteins of the regenerating family member 3 alpha (REG3A) family provide a first line of protection against infections and transformed cells. Their expression is inducible by inflammation, which makes their role in cancer biology less clear since an immune-inflammatory context may preexist or coexist with cancer, as occurs in HCC. The aim of this study is to clarify the role of REG3A in liver carcinogenesis and to determine whether its carbohydrate-binding functions are involved. APPROACH AND RESULTS: This study provides evidence for a suppressive role of REG3A in HCC by reducing O -GlcNAcylation in 2 mouse models of HCC, in vitro cell studies, and clinical samples. REG3A expression in hepatocytes significantly reduced global O -GlcNAcylation and O -GlcNAcylation of c-MYC in preneoplastic and tumor livers and markedly inhibited HCC development in REG3A-c-MYC double transgenic mice and mice exposed to diethylnitrosamine. REG3A modified O -GlcNAcylation without altering the expression or activity of O-linked N-acetylglucosaminyltransferase, O-linked N-acetylglucosaminyl hydrolase, or glutamine fructose-6-phosphate amidotransferase. Reduced O -GlcNAcylation was consistent with decreased levels of UDP-GlcNAc in precancerous and cancerous livers. This effect was linked to the ability of REG3A to bind glucose and glucose-6 phosphate, suggested by a REG3A mutant unable to bind glucose and glucose-6 phosphate and alter O -GlcNAcylation. Importantly, patients with cirrhosis with high hepatic REG3A expression had lower levels of O -GlcNAcylation and longer cancer-free survival than REG3A-negative cirrhotic livers. CONCLUSIONS: REG3A helps fight liver cancer by reducing O -GlcNAcylation. This study suggests a new paradigm for the regulation of O -GlcNAc signaling in cancer-related pathways through interactions with the carbohydrate-binding function of REG3A.
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Innate immune mediators of pathogen clearance, including the secreted C-type lectins REG3 of the antimicrobial peptide (AMP) family, are known to be involved in the regulation of tissue repair and homeostasis. Their role in metabolic homeostasis remains unknown. Here we show that an increase in human REG3A improves glucose and lipid homeostasis in nutritional and genetic mouse models of obesity and type 2 diabetes. Mice overexpressing REG3A in the liver show improved glucose homeostasis, which is reflected in better insulin sensitivity in normal weight and obese states. Delivery of recombinant REG3A protein to leptin-deficient ob/ob mice or wild-type mice on a high-fat diet also improves glucose homeostasis. This is accompanied by reduced oxidative protein damage, increased AMPK phosphorylation and insulin-stimulated glucose uptake in skeletal muscle tissue. Oxidative damage in differentiated C2C12 myotubes is greatly attenuated by REG3A, as is the increase in gp130-mediated AMPK activation. In contrast, Akt-mediated insulin action, which is impaired by oxidative stress, is not restored by REG3A. These data highlight the importance of REG3A in controlling oxidative protein damage involved in energy and metabolic pathways during obesity and diabetes, and provide additional insight into the dual function of host-immune defense and metabolic regulation for AMP.
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Antiinfecciosos , Diabetes Mellitus Experimental , Diabetes Mellitus Tipo 2 , Resistencia a la Insulina , Ratones , Humanos , Animales , Ratones Obesos , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Experimental/metabolismo , Proteínas Quinasas Activadas por AMP/metabolismo , Glucosa/metabolismo , Obesidad/genética , Insulina/farmacología , Homeostasis , Antiinfecciosos/farmacologíaRESUMEN
The bypass of DNA lesions that block replicative polymerases during DNA replication relies on DNA damage tolerance pathways. The error-prone translesion synthesis (TLS) pathway depends on specialized DNA polymerases that incorporate nucleotides in front of base lesions, potentially inducing mutagenesis. Two error-free pathways can bypass the lesions: the template switching pathway, which uses the sister chromatid as a template, and the homologous recombination pathway (HR), which also can use the homologous chromosome as template. The balance between error-prone and error-free pathways controls the mutagenesis level. Therefore, it is crucial to precisely characterize factors that influence the pathway choice to better understand genetic stability at replication forks. In yeast, the complex formed by the Rad51 paralogs Rad55 and Rad57 promotes HR and template-switching at stalled replication forks. At DNA double-strand breaks (DSBs), this complex promotes Rad51 filament formation and stability, notably by counteracting the Srs2 anti-recombinase. To explore the role of the Rad55-Rad57 complex in error-free pathways, we monitored the genetic interactions between Rad55-Rad57, the translesion polymerases Polζ or Polη, and Srs2 following UV radiation that induces mostly single-strand DNA gaps. We found that the Rad55-Rad57 complex was involved in three ways. First, it protects Rad51 filaments from Srs2, as it does at DSBs. Second, it promotes Rad51 filament stability independently of Srs2. Finally, we observed that UV-induced HR is almost abolished in Rad55-Rad57 deficient cells, and is partially restored upon Polζ or Polη depletion. Hence, we propose that the Rad55-Rad57 complex is essential to promote Rad51 filament stability on single-strand DNA gaps, notably to counteract the error-prone TLS polymerases and mutagenesis.
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Proteínas de Saccharomyces cerevisiae , Adenosina Trifosfatasas/genética , ADN/metabolismo , Daño del ADN/genética , ADN Helicasas/genética , Reparación del ADN/genética , Enzimas Reparadoras del ADN/genética , ADN de Cadena Simple/genética , ADN de Cadena Simple/metabolismo , Proteínas de Unión al ADN/genética , ADN Polimerasa Dirigida por ADN/genética , ADN Polimerasa Dirigida por ADN/metabolismo , Recombinasa Rad51/genética , Recombinasa Rad51/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Rayos UltravioletaRESUMEN
Deciphering the molecular architecture of coat coloration for a better understanding of the biological mechanisms underlying pigmentation still remains a challenge. We took advantage of a rabbit French experimental population in which both a pattern and a gradient of coloration from white to brown segregated within the himalayan phenotype. The whole experimental design was genotyped using the high density Affymetrix® AxiomOrcun™ SNP Array and phenotyped into 6 different groups ordered from the lighter to the darker. Genome-wide association analyses pinpointed an oligogenic determinism, under recessive and additive inheritance, involving genes already known in melanogenesis (ASIP, KIT, MC1R, TYR), and likely processed pseudogenes linked to ribosomal function, RPS20 and RPS14. We also identified (i) gene-gene interactions through ASIP:MC1R affecting light cream/beige phenotypes while KIT:RPS responsible of dark chocolate/brown colors and (ii) a genome-wide epistatic network involving several others coloration genes such as POT1 or HPS5. Finally, we determined the recessive inheritance of the English spotting phenotype likely involving a copy number variation affecting at least the end of the coding sequence of the KIT gene. Our analyses of coloration as a continuous trait allowed us to go beyond much of the established knowledge through the detection of additional genes and gene-gene interactions that may contribute to the molecular architecture of the coloration phenotype.
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Variaciones en el Número de Copia de ADN , Estudio de Asociación del Genoma Completo , Animales , Conejos , Proteína de Señalización Agouti/genética , Pigmentación/genética , Fenotipo , ExtremidadesRESUMEN
BACKGROUND: Liver reduction is the main curative treatment for primary liver cancer, but its use remains limited as liver regeneration requires a minimum of 30% functional parenchyma. AIM: To study the dynamics of the liver regeneration process and consequent behavior of cell cycle regulators in rats after extended hepatectomy (90%) and postoperative glucose infusions. METHODS: Post-hepatectomy liver failure was triggered in 84 Wistar rats by reducing their liver mass by 90%. The animals received a post-operative glucose infusion and were randomly assigned to two groups: One to investigate the survival rate and the other for biochemical analyses. Animals that underwent laparotomy or 70% hepatectomy were used as controls. Blood and liver samples were collected on postoperative days 1 to 7. Liver morphology, function, and regeneration were studied with histology, immunohistochemistry, and western blotting. RESULTS: Postoperative mortality after major resection reached 20% and 55% in the first 24 h and 48 h, respectively, with an overall total of 70% 7 d after surgery. No apparent signs of apoptotic cell death were detected in the extended hepatectomy rat livers, but hepatocytes displaying a clear cytoplasm and an accumulation of hyaline material testified to changes affecting their functional activities. Liver regeneration started properly, as early events initiating cell proliferation occurred within the first 3 h, and the G1 to S transition was detected in less than 12 h. However, a rise in p27 (Kip1) followed by p21 (Waf1/Cip1) cell cycle inhibitor levels led to a delayed S phase progression and mitosis. Overall, liver regeneration in rats with a 90% hepatectomy was delayed by 24 h and associated with a delayed onset and lower peak magnitude of hepatocellular deoxyribonucleic acid synthesis. CONCLUSION: This work highlights the critical importance of the cyclin/cyclin-dependent kinase inhibitors of the Cip/Kip family in regulating the liver regeneration timeline following extended hepatectomy.
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In Saccharomyces cerevisiae, dicentric chromosomes stemming from telomere fusions preferentially break at the fusion. This process restores a normal karyotype and protects chromosomes from the detrimental consequences of accidental fusions. Here, we address the molecular basis of this rescue pathway. We observe that tandem arrays tightly bound by the telomere factor Rap1 or a heterologous high-affinity DNA binding factor are sufficient to establish breakage hotspots, mimicking telomere fusions within dicentrics. We also show that condensins generate forces sufficient to rapidly refold dicentrics prior to breakage by cytokinesis and are essential to the preferential breakage at telomere fusions. Thus, the rescue of fused telomeres results from a condensin- and Rap1-driven chromosome folding that favors fusion entrapment where abscission takes place. Because a close spacing between the DNA-bound Rap1 molecules is essential to this process, Rap1 may act by stalling condensins.