Limits...
Sustained activation of Akt elicits mitochondrial dysfunction to block Plasmodium falciparum infection in the mosquito host.

Luckhart S, Giulivi C, Drexler AL, Antonova-Koch Y, Sakaguchi D, Napoli E, Wong S, Price MS, Eigenheer R, Phinney BS, Pakpour N, Pietri JE, Cheung K, Georgis M, Riehle M - PLoS Pathog. (2013)

Bottom Line: Despite an apparent increase in mitochondrial biogenesis in young females (3 d), energy deficiencies were apparent as decreased oxidative phosphorylation and increased [AMP]/[ATP] ratios.Hence, Akt-induced changes in mitochondrial dynamics perturb midgut homeostasis to enhance parasite resistance and decrease mosquito infective lifespan.Further, quality control of mitochondrial function in the midgut is necessary for the maintenance of midgut health as reflected in energy homeostasis and tissue repair and renewal.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Microbiology and Immunology, School of Medicine, University of California, Davis, Davis, California, United States of America. sluckhart@ucdavis.edu

ABSTRACT
The overexpression of activated, myristoylated Akt in the midgut of female transgenic Anopheles stephensi results in resistance to infection with the human malaria parasite Plasmodium falciparum but also decreased lifespan. In the present study, the understanding of mitochondria-dependent midgut homeostasis has been expanded to explain this apparent paradox in an insect of major medical importance. Given that Akt signaling is essential for cell growth and survival, we hypothesized that sustained Akt activation in the mosquito midgut would alter the balance of critical pathways that control mitochondrial dynamics to enhance parasite killing at some cost to survivorship. Toxic reactive oxygen and nitrogen species (RNOS) rise to high levels in the midgut after blood feeding, due to a combination of high NO production and a decline in FOXO-dependent antioxidants. Despite an apparent increase in mitochondrial biogenesis in young females (3 d), energy deficiencies were apparent as decreased oxidative phosphorylation and increased [AMP]/[ATP] ratios. In addition, mitochondrial mass was lower and accompanied by the presence of stalled autophagosomes in the posterior midgut, a critical site for blood digestion and stem cell-mediated epithelial maintenance and repair, and by functional degradation of the epithelial barrier. By 18 d, the age at which An. stephensi would transmit P. falciparum to human hosts, mitochondrial dysfunction coupled to Akt-mediated repression of autophagy/mitophagy was more evident and midgut epithelial structure was markedly compromised. Inhibition of RNOS by co-feeding of the nitric-oxide synthase inhibitor L-NAME at infection abrogated Akt-dependent killing of P. falciparum that begins within 18 h of infection in 3-5 d old mosquitoes. Hence, Akt-induced changes in mitochondrial dynamics perturb midgut homeostasis to enhance parasite resistance and decrease mosquito infective lifespan. Further, quality control of mitochondrial function in the midgut is necessary for the maintenance of midgut health as reflected in energy homeostasis and tissue repair and renewal.

Show MeSH

Related in: MedlinePlus

Resistance to P. falciparum in myrAkt HM An. stephensi was reversed by NOS inhibition.Preparation and treatments for mosquitoes is described in the Methods. (A) Prevalence of infection (percentage of mosquitoes dissected with at least one P. falciparum oocyst) of mosquitoes dissected in (B). Fisher's exact test was used to compare treatments against L-NAME; NS = not significant at alpha = 0.05. Numbers above bars reflect mean prevalences for control and treatment groups. (B) Water, D-NAME, L-NAME treatments with myrAkt HM An. stephensi. Age- and cohort-matched NTG control mosquitoes were infected side-by-side with the same parasite culture used for HM myrAkt An. stephensi. Numbers within the figure reflect mean oocysts per midgut for control and treatment groups. This experiment was repeated four times with four separate cohorts of mosquitoes. Data shown are from infected midguts (no zeros). H2O and D-NAME were outside of the 95% confidence intervals for NTG (1.79,3.05) and L-NAME (1.41,4.01); the latter groups were not different by Mann-Whitney U-test (alpha = 0.05).
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3585164&req=5

ppat-1003180-g010: Resistance to P. falciparum in myrAkt HM An. stephensi was reversed by NOS inhibition.Preparation and treatments for mosquitoes is described in the Methods. (A) Prevalence of infection (percentage of mosquitoes dissected with at least one P. falciparum oocyst) of mosquitoes dissected in (B). Fisher's exact test was used to compare treatments against L-NAME; NS = not significant at alpha = 0.05. Numbers above bars reflect mean prevalences for control and treatment groups. (B) Water, D-NAME, L-NAME treatments with myrAkt HM An. stephensi. Age- and cohort-matched NTG control mosquitoes were infected side-by-side with the same parasite culture used for HM myrAkt An. stephensi. Numbers within the figure reflect mean oocysts per midgut for control and treatment groups. This experiment was repeated four times with four separate cohorts of mosquitoes. Data shown are from infected midguts (no zeros). H2O and D-NAME were outside of the 95% confidence intervals for NTG (1.79,3.05) and L-NAME (1.41,4.01); the latter groups were not different by Mann-Whitney U-test (alpha = 0.05).

Mentions: To first assess whether overproduction of NO was responsible for resistance to P. falciparum in myrAkt An. stephensi, four separate cohorts of HM An. stephensi females were provided with water, Nω-Nitro-L-arginine methyl ester (3.7 mM, L-NAME; [69], [71]) or the biologically inactive isomer D-NAME from 72 h before blood feeding though P. falciparum infection and thereafter until dissection. Age- and cohort-matched NTG females were infected side-by-side as controls. After 10 days, females were dissected to visualize and count P. falciparum oocysts. Among those NTG An. stephensi that were fully gravid (an indicator of complete engorgement), 49% had at least one midgut oocyst (Fig. 10A). Infected NTG mosquitoes averaged 2.4 P. falciparum oocysts per midgut (Fig. 10B). As expected, HM females provided only with water or with water with D-NAME were resistant to infection (Fig. 10B). However, provision of L-NAME to HM An. stephensi reversed the phenotype of resistance to infection, resulting in a prevalence and intensity of infection that were not significantly different from control, NTG females fed on the same P. falciparum-infected blood (Figs. 10A, B). Neither L-NAME nor D-NAME had significant effects on P. falciparum growth in the absence of the mosquito (Fig. S3). Although this growth assay cannot be performed efficiently on mosquito-stage parasites, we assert that our results suggest that observed infection patterns were due to L-NAME effects on the mosquito host.


Sustained activation of Akt elicits mitochondrial dysfunction to block Plasmodium falciparum infection in the mosquito host.

Luckhart S, Giulivi C, Drexler AL, Antonova-Koch Y, Sakaguchi D, Napoli E, Wong S, Price MS, Eigenheer R, Phinney BS, Pakpour N, Pietri JE, Cheung K, Georgis M, Riehle M - PLoS Pathog. (2013)

Resistance to P. falciparum in myrAkt HM An. stephensi was reversed by NOS inhibition.Preparation and treatments for mosquitoes is described in the Methods. (A) Prevalence of infection (percentage of mosquitoes dissected with at least one P. falciparum oocyst) of mosquitoes dissected in (B). Fisher's exact test was used to compare treatments against L-NAME; NS = not significant at alpha = 0.05. Numbers above bars reflect mean prevalences for control and treatment groups. (B) Water, D-NAME, L-NAME treatments with myrAkt HM An. stephensi. Age- and cohort-matched NTG control mosquitoes were infected side-by-side with the same parasite culture used for HM myrAkt An. stephensi. Numbers within the figure reflect mean oocysts per midgut for control and treatment groups. This experiment was repeated four times with four separate cohorts of mosquitoes. Data shown are from infected midguts (no zeros). H2O and D-NAME were outside of the 95% confidence intervals for NTG (1.79,3.05) and L-NAME (1.41,4.01); the latter groups were not different by Mann-Whitney U-test (alpha = 0.05).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3585164&req=5

ppat-1003180-g010: Resistance to P. falciparum in myrAkt HM An. stephensi was reversed by NOS inhibition.Preparation and treatments for mosquitoes is described in the Methods. (A) Prevalence of infection (percentage of mosquitoes dissected with at least one P. falciparum oocyst) of mosquitoes dissected in (B). Fisher's exact test was used to compare treatments against L-NAME; NS = not significant at alpha = 0.05. Numbers above bars reflect mean prevalences for control and treatment groups. (B) Water, D-NAME, L-NAME treatments with myrAkt HM An. stephensi. Age- and cohort-matched NTG control mosquitoes were infected side-by-side with the same parasite culture used for HM myrAkt An. stephensi. Numbers within the figure reflect mean oocysts per midgut for control and treatment groups. This experiment was repeated four times with four separate cohorts of mosquitoes. Data shown are from infected midguts (no zeros). H2O and D-NAME were outside of the 95% confidence intervals for NTG (1.79,3.05) and L-NAME (1.41,4.01); the latter groups were not different by Mann-Whitney U-test (alpha = 0.05).
Mentions: To first assess whether overproduction of NO was responsible for resistance to P. falciparum in myrAkt An. stephensi, four separate cohorts of HM An. stephensi females were provided with water, Nω-Nitro-L-arginine methyl ester (3.7 mM, L-NAME; [69], [71]) or the biologically inactive isomer D-NAME from 72 h before blood feeding though P. falciparum infection and thereafter until dissection. Age- and cohort-matched NTG females were infected side-by-side as controls. After 10 days, females were dissected to visualize and count P. falciparum oocysts. Among those NTG An. stephensi that were fully gravid (an indicator of complete engorgement), 49% had at least one midgut oocyst (Fig. 10A). Infected NTG mosquitoes averaged 2.4 P. falciparum oocysts per midgut (Fig. 10B). As expected, HM females provided only with water or with water with D-NAME were resistant to infection (Fig. 10B). However, provision of L-NAME to HM An. stephensi reversed the phenotype of resistance to infection, resulting in a prevalence and intensity of infection that were not significantly different from control, NTG females fed on the same P. falciparum-infected blood (Figs. 10A, B). Neither L-NAME nor D-NAME had significant effects on P. falciparum growth in the absence of the mosquito (Fig. S3). Although this growth assay cannot be performed efficiently on mosquito-stage parasites, we assert that our results suggest that observed infection patterns were due to L-NAME effects on the mosquito host.

Bottom Line: Despite an apparent increase in mitochondrial biogenesis in young females (3 d), energy deficiencies were apparent as decreased oxidative phosphorylation and increased [AMP]/[ATP] ratios.Hence, Akt-induced changes in mitochondrial dynamics perturb midgut homeostasis to enhance parasite resistance and decrease mosquito infective lifespan.Further, quality control of mitochondrial function in the midgut is necessary for the maintenance of midgut health as reflected in energy homeostasis and tissue repair and renewal.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Microbiology and Immunology, School of Medicine, University of California, Davis, Davis, California, United States of America. sluckhart@ucdavis.edu

ABSTRACT
The overexpression of activated, myristoylated Akt in the midgut of female transgenic Anopheles stephensi results in resistance to infection with the human malaria parasite Plasmodium falciparum but also decreased lifespan. In the present study, the understanding of mitochondria-dependent midgut homeostasis has been expanded to explain this apparent paradox in an insect of major medical importance. Given that Akt signaling is essential for cell growth and survival, we hypothesized that sustained Akt activation in the mosquito midgut would alter the balance of critical pathways that control mitochondrial dynamics to enhance parasite killing at some cost to survivorship. Toxic reactive oxygen and nitrogen species (RNOS) rise to high levels in the midgut after blood feeding, due to a combination of high NO production and a decline in FOXO-dependent antioxidants. Despite an apparent increase in mitochondrial biogenesis in young females (3 d), energy deficiencies were apparent as decreased oxidative phosphorylation and increased [AMP]/[ATP] ratios. In addition, mitochondrial mass was lower and accompanied by the presence of stalled autophagosomes in the posterior midgut, a critical site for blood digestion and stem cell-mediated epithelial maintenance and repair, and by functional degradation of the epithelial barrier. By 18 d, the age at which An. stephensi would transmit P. falciparum to human hosts, mitochondrial dysfunction coupled to Akt-mediated repression of autophagy/mitophagy was more evident and midgut epithelial structure was markedly compromised. Inhibition of RNOS by co-feeding of the nitric-oxide synthase inhibitor L-NAME at infection abrogated Akt-dependent killing of P. falciparum that begins within 18 h of infection in 3-5 d old mosquitoes. Hence, Akt-induced changes in mitochondrial dynamics perturb midgut homeostasis to enhance parasite resistance and decrease mosquito infective lifespan. Further, quality control of mitochondrial function in the midgut is necessary for the maintenance of midgut health as reflected in energy homeostasis and tissue repair and renewal.

Show MeSH
Related in: MedlinePlus