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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.

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Midgut metabolite profiles in HM myrAkt An. stephensi suggested marked energy deficiencies relative to NTG mosquitoes at 3 d and 18 d post-emergence.All metabolites were evaluated by HPLC as described under Methods. ATP, ADP, AMP, AN (ATP+ADP+AMP), NAD, NADH and NAD+NADH were calculated as nmol/midgut and data are represented as fold of NTG values. The NTG values were the average of 3 d and 18 d given that no statistical differences were observed between these two days. Data were analyzed relative to NTG values with Student's t-test (alpha = 0.05).*P<0.05 relative to NTG values.
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ppat-1003180-g007: Midgut metabolite profiles in HM myrAkt An. stephensi suggested marked energy deficiencies relative to NTG mosquitoes at 3 d and 18 d post-emergence.All metabolites were evaluated by HPLC as described under Methods. ATP, ADP, AMP, AN (ATP+ADP+AMP), NAD, NADH and NAD+NADH were calculated as nmol/midgut and data are represented as fold of NTG values. The NTG values were the average of 3 d and 18 d given that no statistical differences were observed between these two days. Data were analyzed relative to NTG values with Student's t-test (alpha = 0.05).*P<0.05 relative to NTG values.

Mentions: Examination of OXPHOS capacity revealed that in HM mosquitoes, activities of Complex I, Complex II–III and Complex V were 75% (for Complex I average of both NQR and NFR activities), 30% and 70% of NTG controls at 3 d, respectively, whereas at 18 d, these values were 50%, 40% and 50% of NTG controls, respectively (Table 3). These activities were still lower than controls when normalized to citrate synthase, making them independent of the number of mitochondria present at any given time point in whole midguts (Table 3). Relatively lower ETC activities can result in lower OXPHOS and energy deficits. To test this hypothesis, the energy charge potential (ECP) – defined as ([ATP]+0.5*[ADP])/([ATP]+[ADP]+[AMP]; [64]) – was evaluated in NTG and HM female midguts at 3 d and 18 d post-emergence and in whole bodies of 3 d NTG and HM females. In midguts from HM females at 3 d, the ECP showed some decline (94% of controls), which was clearly lower at 18 d (89% of controls; Table 3,Fig. 7). In addition to local tissue effects, overexpression of myrAkt in the An. stephensi midgut was associated with significantly reduced whole body total adenosine metabolites. In particular, [ADP] (in percentage of total nucleotides) at 3 d was 2-fold higher, [AMP] was 7.7-fold higher, and ECP was significantly reduced relative to age-matched NTG controls (Table 4) despite the fact that Akt overexpression was targeted to the midgut of An. stephensi. These results (decreased ECP, higher [ADP]/[ATP] and [AMP]/[ATP] ratios) indicated clear energy deficiencies locally in the midgut and systemically in the body of HM mosquitoes, suggesting that (i) mitochondrial biogenesis could not be correctly completed (defects at import/assembly) or (ii) oxidative/nitrative stress-mediated damage overpowered this putative compensatory response.


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)

Midgut metabolite profiles in HM myrAkt An. stephensi suggested marked energy deficiencies relative to NTG mosquitoes at 3 d and 18 d post-emergence.All metabolites were evaluated by HPLC as described under Methods. ATP, ADP, AMP, AN (ATP+ADP+AMP), NAD, NADH and NAD+NADH were calculated as nmol/midgut and data are represented as fold of NTG values. The NTG values were the average of 3 d and 18 d given that no statistical differences were observed between these two days. Data were analyzed relative to NTG values with Student's t-test (alpha = 0.05).*P<0.05 relative to NTG values.
© Copyright Policy
Related In: Results  -  Collection

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

ppat-1003180-g007: Midgut metabolite profiles in HM myrAkt An. stephensi suggested marked energy deficiencies relative to NTG mosquitoes at 3 d and 18 d post-emergence.All metabolites were evaluated by HPLC as described under Methods. ATP, ADP, AMP, AN (ATP+ADP+AMP), NAD, NADH and NAD+NADH were calculated as nmol/midgut and data are represented as fold of NTG values. The NTG values were the average of 3 d and 18 d given that no statistical differences were observed between these two days. Data were analyzed relative to NTG values with Student's t-test (alpha = 0.05).*P<0.05 relative to NTG values.
Mentions: Examination of OXPHOS capacity revealed that in HM mosquitoes, activities of Complex I, Complex II–III and Complex V were 75% (for Complex I average of both NQR and NFR activities), 30% and 70% of NTG controls at 3 d, respectively, whereas at 18 d, these values were 50%, 40% and 50% of NTG controls, respectively (Table 3). These activities were still lower than controls when normalized to citrate synthase, making them independent of the number of mitochondria present at any given time point in whole midguts (Table 3). Relatively lower ETC activities can result in lower OXPHOS and energy deficits. To test this hypothesis, the energy charge potential (ECP) – defined as ([ATP]+0.5*[ADP])/([ATP]+[ADP]+[AMP]; [64]) – was evaluated in NTG and HM female midguts at 3 d and 18 d post-emergence and in whole bodies of 3 d NTG and HM females. In midguts from HM females at 3 d, the ECP showed some decline (94% of controls), which was clearly lower at 18 d (89% of controls; Table 3,Fig. 7). In addition to local tissue effects, overexpression of myrAkt in the An. stephensi midgut was associated with significantly reduced whole body total adenosine metabolites. In particular, [ADP] (in percentage of total nucleotides) at 3 d was 2-fold higher, [AMP] was 7.7-fold higher, and ECP was significantly reduced relative to age-matched NTG controls (Table 4) despite the fact that Akt overexpression was targeted to the midgut of An. stephensi. These results (decreased ECP, higher [ADP]/[ATP] and [AMP]/[ATP] ratios) indicated clear energy deficiencies locally in the midgut and systemically in the body of HM mosquitoes, suggesting that (i) mitochondrial biogenesis could not be correctly completed (defects at import/assembly) or (ii) oxidative/nitrative stress-mediated damage overpowered this putative compensatory response.

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