ABSTRACT
The intracellular parasite Toxoplasma gondii induces host AKT activation to prevent autophagy-mediated clearance; however, the molecular underpinnings are not fully understood. Autophagy can be negatively regulated through AKT-sensitive phosphorylation and nuclear export of the transcription factor Forkhead box O3a (FOXO3a). Using a combination of pharmacological and genetic approaches, herein we investigated whether T. gondii hinders host autophagy through AKT-dependent inactivation of FOXO3a. We found that infection by type I and II strains of T. gondii promotes gradual and sustained AKT-dependent phosphorylation of FOXO3a at residues S253 and T32 in human foreskin fibroblasts (HFF) and murine 3T3 fibroblasts. Mechanistically, AKT-sensitive phosphorylation of FOXO3a by T. gondii required live infection and the activity of PI3K but was independent of the plasma membrane receptor EGFR and the kinase PKCα. Phosphorylation of FOXO3a at AKT-sensitive residues was paralleled by its nuclear exclusion in T. gondii-infected HFF. Importantly, the parasite was unable to drive cytoplasmic localization of FOXO3a upon pharmacological blockade of AKT or overexpression of an AKT-insensitive mutant form of FOXO3a. Transcription of a subset of bona fide autophagy-related targets of FOXO3a was reduced during T. gondii infection in an AKT-dependent fashion. However, parasite-directed repression of autophagy-related genes was AKT-resistant in cells deficient in FOXO3a. Consistent with this, T. gondii failed to inhibit the recruitment of acidic organelles and LC3, an autophagy marker, to the parasitophorous vacuole upon chemically or genetically induced nuclear retention of FOXO3a. In all, we provide evidence that T. gondii suppresses FOXO3a-regulated transcriptional programs to prevent autophagy-mediated killing. IMPORTANCE The parasite Toxoplasma gondii is the etiological agent of toxoplasmosis, an opportunistic infection commonly transmitted by ingestion of contaminated food or water. To date, no effective vaccines in humans have been developed and no promising drugs are available to treat chronic infection or prevent congenital infection. T. gondii targets numerous host cell processes to establish a favorable replicative niche. Of note, T. gondii activates the host AKT signaling pathway to prevent autophagy-mediated killing. Herein, we report that T. gondii inhibits FOXO3a, a transcription factor that regulates the expression of autophagy-related genes, through AKT-dependent phosphorylation. The parasite's ability to block the recruitment of the autophagy machinery to the parasitophorous vacuole is impeded upon pharmacological inhibition of AKT or overexpression of an AKT-insensitive form of FOXO3a. Thus, our study provides greater granularity in the role of FOXO3a during infection and reinforces the potential of targeting autophagy as a therapeutic strategy against T. gondii.
ABSTRACT
Introducción: en la actualidad, nuevas tendencias tecnológicas e iniciativas se están presentando en el desarrollo de productos insecticidas derivados de productos naturales, y de nuevos agentes antimicrobianos, dado que poseen bioactivos que son selectivos, biodegradables y tienen menores efectos adversos. La especie Ambrosia peruviana Willd. es de gran interés en el estudio por su gran potencial biológico y etnobotánico. Objetivo: evaluar la actividad larvicida sobre Aedes aegypty L. y la actividad antibacteriana sobre bacterias Gram positivas y Gram negativas de extractos de A. peruviana. Métodos: a partir del material vegetal seco (hojas), se obtuvieron cinco extractos de diferente polaridad en hexano (H), diclorometano (D), acetato de etilo (A) y etanol (E) y aceites esenciales (AE), los cuales fueron evaluados mediante la inhibición del crecimiento de larvas por el método recomendado de la OMS y la inhibición de las bacterias por el método de difusión en agar de Kirby-Bauer. Resultados: la tasa de mortalidad encontrada a las 24 h a una concentración de 200 ppm para todos los extractos fue del 10 por ciento. Al evaluar el paso de los insectos de larvas a adultos a las 144 h se observó a esta misma concentración una mortalidad del 100 por ciento con todos los extractos. Por otra parte, los extractos de A. peruviana presentaron inhibición sobre Bacillus cereus Frankland & Frankland y Bacillus subtilis (Ehrenberg) Cohn con halos de inhibición del extracto de diclorometano (APExtD) de 10,5 y 15,0 mm de diámetro respectivamente, al contrario sobre las cepas Serratia marcescens Bizio, Proteus mirabilis Hauser, Enterobacter cloacae (Jordan) Hormaeche & Edwards y Staphylococcus aureus Rosenbach no se presentó actividad antibacteriana. Conclusiones: esta investigación es el primer reporte de actividad larvicida sobre A. aegypty y de actividad antibacteriana sobre B. cereus y B. subtilis de varios extractos de A. peruviana con promisorios resultados en estos modelos(AU)
Introduction: New technological trends and initiatives are currently being put forth concerning the development of insecticidal products and antimicrobial agents of natural origin, since their bioactive components are selective and biodegradable, and cause fewer adverse effects. The species Ambrosia peruviana Willd. was of great interest to the present study, due to its great biological and ethnobotanical potential. Objective: Evaluate the larvicidal activity of A. peruviana extracts against Aedes aegypti L., and its antibacterial activity against gram-positive and gram-negative bacteria. Methods: Dry plant material (leaves) was processed to obtain five extracts of different polarity in hexane (H), dichloromethane (D), ethyl acetate (A), ethanol (E) and essential oils (AE), which were evaluated for larval growth inhibition with the method recommended by WHO, and for bacterial inhibition with the Kirby-Bauer agar diffusion method. Results: The mortality rate at 24 h and a concentration of 200 ppm was 10 percent for all extracts. Examination of the transition of larvae into adults at 144 h and the same concentration revealed a mortality of 100 percent with all extracts. On the other hand, the extracts of A. peruviana displayed inhibition capacity against Bacillus cereus Frankland & Frankland and Bacillus subtilis (Ehrenberg) Cohn with inhibition haloes for the dichloromethane extract (APExtD) of 10.5 and 15.0 mm in diameter, respectively, whereas no antibacterial activity was found against the strains Serratia marcescens Bizio, Proteus mirabilis Hauser, Enterobacter cloacae (Jordan) Hormaeche & Edwards and Staphylococcus aureus. Conclusions: This study is the first report of larvicidal activity againstA. aegypti and antibacterial activity against B. cereus and B. subtilis by several extracts of A. peruviana with promising results in these models(AU)
Subject(s)
Animals , Aedes/pathogenicity , Insecticides/therapeutic use , Pest Control, Biological/methods , Teucrium , Teucrium/poisoning , Vector Control of DiseasesABSTRACT
Introducción: en la actualidad, nuevas tendencias tecnológicas e iniciativas se están presentando en el desarrollo de productos insecticidas derivados de productos naturales, y de nuevos agentes antimicrobianos, dado que poseen bioactivos que son selectivos, biodegradables y tienen menores efectos adversos. La especie Ambrosia peruviana Willd. es de gran interés en el estudio por su gran potencial biológico y etnobotánico. Objetivo: evaluar la actividad larvicida sobre Aedes aegypty L. y la actividad antibacteriana sobre bacterias Gram positivas y Gram negativas de extractos de A. peruviana. Métodos: a partir del material vegetal seco (hojas), se obtuvieron cinco extractos de diferente polaridad en hexano (H), diclorometano (D), acetato de etilo (A) y etanol (E) y aceites esenciales (AE), los cuales fueron evaluados mediante la inhibición del crecimiento de larvas por el método recomendado de la OMS y la inhibición de las bacterias por el método de difusión en agar de Kirby-Bauer. Resultados: la tasa de mortalidad encontrada a las 24 h a una concentración de 200 ppm para todos los extractos fue del 10 por ciento. Al evaluar el paso de los insectos de larvas a adultos a las 144 h se observó a esta misma concentración una mortalidad del 100 por ciento con todos los extractos. Por otra parte, los extractos de A. peruviana presentaron inhibición sobre Bacillus cereus Frankland & Frankland y Bacillus subtilis (Ehrenberg) Cohn con halos de inhibición del extracto de diclorometano (APExtD) de 10,5 y 15,0 mm de diámetro respectivamente, al contrario sobre las cepas Serratia marcescens Bizio, Proteus mirabilis Hauser, Enterobacter cloacae (Jordan) Hormaeche & Edwards y Staphylococcus aureus Rosenbach no se presentó actividad antibacteriana. Conclusiones: esta investigación es el primer reporte de actividad larvicida sobre A. aegypty y de actividad antibacteriana sobre B. cereus y B. subtilis de varios extractos de A. peruviana con promisorios resultados en estos modelos(AU)
Introduction: New technological trends and initiatives are currently being put forth concerning the development of insecticidal products and antimicrobial agents of natural origin, since their bioactive components are selective and biodegradable, and cause fewer adverse effects. The species Ambrosia peruviana Willd. was of great interest to the present study, due to its great biological and ethnobotanical potential. Objective: Evaluate the larvicidal activity of A. peruviana extracts against Aedes aegypti L., and its antibacterial activity against gram-positive and gram-negative bacteria. Methods: Dry plant material (leaves) was processed to obtain five extracts of different polarity in hexane (H), dichloromethane (D), ethyl acetate (A), ethanol (E) and essential oils (AE), which were evaluated for larval growth inhibition with the method recommended by WHO, and for bacterial inhibition with the Kirby-Bauer agar diffusion method. Results: The mortality rate at 24 h and a concentration of 200 ppm was 10 percent for all extracts. Examination of the transition of larvae into adults at 144 h and the same concentration revealed a mortality of 100 percent with all extracts. On the other hand, the extracts of A. peruviana displayed inhibition capacity against Bacillus cereus Frankland & Frankland and Bacillus subtilis (Ehrenberg) Cohn with inhibition haloes for the dichloromethane extract (APExtD) of 10.5 and 15.0 mm in diameter, respectively, whereas no antibacterial activity was found against the strains Serratia marcescens Bizio, Proteus mirabilis Hauser, Enterobacter cloacae (Jordan) Hormaeche & Edwards and Staphylococcus aureus. Conclusions: This study is the first report of larvicidal activity againstA. aegypti and antibacterial activity against B. cereus and B. subtilis by several extracts of A. peruviana with promising results in these models(AU)
