RESUMEN
Chitin is the main component of insect exoskeleton and midgut peritrophic membrane. Insect molting is the result of the balance and coordination of chitin synthesis and degradation in chitin metabolism under the action of hormones. In this study, a 678 bp dsRNA fragment was designed and synthesized according to the known CHI (Chitinase) sequence of Spodoptera frugiperda. It was injected into the larvae to observe the molting and development of S. frugiperda. At the same time, the activities of trehalase and chitinase, the contents of trehalose, chitin and other substances were detected, and the expression of related genes in the chitin synthesis pathway was determined. The results showed that CHI gene was highly expressed at the end of each instar, prepupa and pupal stage before molting; At 12 and 24 h after dsRNA injection of CHI gene of S. frugiperda, the expression of CHI gene decreased significantly, and the chitinase activity decreased significantly from 12 to 48 h. The expression of chitin synthase (CHSB) gene decreased significantly, and the chitin content increased significantly. Some larvae could not molt normally and complete development, leading to certain mortality. Secondly, after RNAi of CHI gene, the content of glucose and glycogen increased first and then decreased, while the content of trehalose decreased significantly or showed a downward trend. The activities of the two types of trehalase and the expression levels of trehalase genes decreased first and then increased, especially the trehalase activities increased significantly at 48 h after dsCHI injection. And trehalose-6-phosphate synthase (TPS), glutamine: fructose-6-phosphate amidotransferase (GFAT), UDP-N-acetylglucosamine pyrophosphorylases (UAP), hexokinase (HK), glucose-6-phosphate isomerase (G6PI) and phosphoacetylglucosamine mutase (PAGM) all decreased significantly at 24 h, and then increased or significantly increased at 48 h. These results indicated that when the expression of chitinase gene of S. frugiperda was inhibited, it affected the degradation of chitin in the old epidermis and the formation of new epidermis, and the content of chitin increased, which led to the failure of larvae to molt normally. Moreover, the chitin synthesis pathway and trehalose metabolism were also regulated. The relevant results provide a theoretical basis for screening target genes and developing green insecticides to control pests by using the chitin metabolism pathway.
RESUMEN
Thoracic aortic aneurysm (TAA) has been associated with mutations affecting members of the TGF-ß signaling pathway, or components and regulators of the vascular smooth muscle cell (VSMC) actomyosin cytoskeleton. Although both clinical groups present similar phenotypes, the existence of potential common mechanisms of pathogenesis remain obscure. Here we show that mutations affecting TGF-ß signaling and VSMC cytoskeleton both lead to the formation of a ternary complex comprising the histone deacetylase HDAC9, the chromatin-remodeling enzyme BRG1, and the long noncoding RNA MALAT1. The HDAC9-MALAT1-BRG1 complex binds chromatin and represses contractile protein gene expression in association with gain of histone H3-lysine 27 trimethylation modifications. Disruption of Malat1 or Hdac9 restores contractile protein expression, improves aortic mural architecture, and inhibits experimental aneurysm growth. Thus, we highlight a shared epigenetic pathway responsible for VSMC dysfunction in both forms of TAA, with potential therapeutic implication for other known HDAC9-associated vascular diseases.
Asunto(s)
Aneurisma de la Aorta Torácica/genética , ADN Helicasas/genética , Histona Desacetilasas/genética , Músculo Liso Vascular/patología , Proteínas Nucleares/genética , ARN Largo no Codificante/genética , Proteínas Represoras/genética , Factores de Transcripción/genética , Factor de Crecimiento Transformador beta/metabolismo , Actomiosina/genética , Actomiosina/metabolismo , Animales , Aorta/patología , Aneurisma de la Aorta Torácica/patología , Línea Celular , Núcleo Celular/metabolismo , Cromatina/metabolismo , ADN Helicasas/metabolismo , Metilación de ADN , Modelos Animales de Enfermedad , Femenino , Técnica del Anticuerpo Fluorescente , Histona Desacetilasas/metabolismo , Histonas/metabolismo , Humanos , Masculino , Ratones , Ratones Noqueados , Músculo Liso Vascular/citología , Mutación , Miocitos del Músculo Liso , Proteínas Nucleares/metabolismo , Fenotipo , Cultivo Primario de Células , Interferencia de ARN , ARN Largo no Codificante/metabolismo , ARN Interferente Pequeño/metabolismo , Proteínas Represoras/metabolismo , Transducción de Señal/genética , Factores de Transcripción/metabolismoRESUMEN
Thoracic aortic aneurysms (TAs) occur in reproducible patterns, but etiologic factors determining the anatomic distribution of these aneurysms are not well understood. This study sought to gain insight into etiologic differences and clinical outcomes associated with repetitive anatomic distributions of TAs. From 3,247 patients registered in an institutional Thoracic Aortic Center database from July 1992 to August 2013, we identified 844 patients with full aortic dimensional imaging by computerized axial tomography or magnetic resonance imaging scan (mean age 62.8 ± 14 years, 37% women, median follow-up 40 months) with TA diameter >4.0 cm and without evidence of previous aortic dissection. Patient demographic and imaging data were analyzed in 3 groups: isolated ascending thoracic aortic aneurysms (AAs; n = 628), isolated descending TAs (DTAs; n = 130), and combined AA and DTA (mixed thoracic aortic aneurysm, MTA; n = 86). Patients with DTA had more hypertension (82% vs 59%, p <0.001) and a higher burden of atherosclerosis (88% vs 9%, p <0.001) than AA. Conversely, patients with isolated AA were younger (59.5 ± 13.5 vs 71.0 ± 11.8 years, p <0.001) and contained almost every case of overt, genetically triggered TA. Patients with isolated DTA were demographically indistinguishable from patients with MTA. In follow-up, patients with DTA/MTA experienced more aortic events (aortic dissection/rupture) and had higher mortality than patients with isolated AA. In multivariate analysis, aneurysm size (odds ratio 1.1, 95% CI 1.07 to 1.16, p <0.001) and the presence of atherosclerosis (odds ratio 5.7, 95% CI 2.02 to 16.15, p <0.001) independently predicted adverse aortic events. We find that DTA with or without associated AA appears to be a disease more highly associated with atherosclerosis, hypertension, and advanced age. In contrast, isolated AA appears to be a clinically distinct entity with a greater burden of genetically triggered disease.
Asunto(s)
Aorta Torácica/diagnóstico por imagen , Aneurisma de la Aorta Torácica/mortalidad , Disección Aórtica/mortalidad , Rotura de la Aorta/mortalidad , Anciano , Disección Aórtica/diagnóstico , Aneurisma de la Aorta Torácica/diagnóstico , Rotura de la Aorta/diagnóstico , Femenino , Estudios de Seguimiento , Humanos , Imagen por Resonancia Cinemagnética , Masculino , Massachusetts/epidemiología , Persona de Mediana Edad , Pronóstico , Estudios Prospectivos , Factores de Riesgo , Tasa de Supervivencia/tendencias , Tomografía Computarizada por Rayos XRESUMEN
Elongating DNA polymerases frequently encounter lesions or structures that impede progress and require repair before DNA replication can be completed. Therefore, directing repair factors to a blocked fork, without interfering with normal replication, is important for proper cell function, and it is a process that is not well understood. To study this process, we have employed the chain-terminating nucleoside analog, 3' azidothymidine (AZT) and the E. coli genetic system, for which replication and repair factors have been well-defined. By using high-expression suppressor screens, we identified yoaA, encoding a putative helicase, and holC, encoding the Chi component of the replication clamp loader, as genes that promoted tolerance to AZT. YoaA is a putative Fe-S helicase in the XPD/RAD3 family for which orthologs can be found in most bacterial genomes; E. coli has a paralog to YoaA, DinG, which possesses 5' to 3' helicase activity and an Fe-S cluster essential to its activity. Mutants in yoaA are sensitive to AZT exposure; dinG mutations cause mild sensitivity to AZT and exacerbate the sensitivity of yoaA mutant strains. Suppression of AZT sensitivity by holC or yoaA was mutually codependent and we provide evidence here that YoaA and Chi physically interact. Interactions of Chi with single-strand DNA binding protein (SSB) and with Psi were required to aid AZT tolerance, as was the proofreading 3' exonuclease, DnaQ. Our studies suggest that repair is coupled to blocked replication through these interactions. We hypothesize that SSB, through Chi, recruits the YoaA helicase to replication gaps and that unwinding of the nascent strand promotes repair and AZT excision. This recruitment prevents the toxicity of helicase activity and aids the handoff of repair with replication factors, ensuring timely repair and resumption of replication.