Limits...
Expression of Drosophila FOXO regulates growth and can phenocopy starvation.

Kramer JM, Davidge JT, Lockyer JM, Staveley BE - BMC Dev. Biol. (2003)

Bottom Line: Analysis of the wings and eyes of these small flies indicates that the reduction in size is due to decreases in cell size and cell number.Overexpression of dFOXO in the developing eye leads to a characteristic phenotype with reductions in cell size and cell number.This phenotype can be rescued by co-expression of upstream insulin signaling components, dPI3K and dAkt, however, this rescue is not seen when FOXO is mutated to a constitutively active form. dFOXO is conserved in both sequence and regulatory mechanisms when compared with other FOXO homologues.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biology, Memorial University of Newfoundland, St, John's, Newfoundland, (A1B 3X9), Canada. x04jmk@mun.ca

ABSTRACT

Background: Components of the insulin signaling pathway are important regulators of growth. The FOXO (forkhead box, sub-group "O") transcription factors regulate cellular processes under conditions of low levels of insulin signaling. Studies in mammalian cell culture show that activation of FOXO transcription factors causes cell death or cell cycle arrest. The Caenorhabditis elegans homologue of FOXO, Daf-16, is required for the formation of dauer larvae in response to nutritional stress. In addition, FOXO factors have been implicated in stress resistance and longevity.

Results: We have identified the Drosophila melanogaster homologue of FOXO (dFOXO), which is conserved in amino acid sequence compared with the mammalian FOXO homologues and Daf-16. Expression of dFOXO during early larval development causes inhibition of larval growth and alterations in feeding behavior. Inhibition of larval growth is reversible upon discontinuation of dFOXO expression. Expression of dFOXO during the third larval instar or at low levels during development leads to the generation of adults that are reduced in size. Analysis of the wings and eyes of these small flies indicates that the reduction in size is due to decreases in cell size and cell number. Overexpression of dFOXO in the developing eye leads to a characteristic phenotype with reductions in cell size and cell number. This phenotype can be rescued by co-expression of upstream insulin signaling components, dPI3K and dAkt, however, this rescue is not seen when FOXO is mutated to a constitutively active form.

Conclusions: dFOXO is conserved in both sequence and regulatory mechanisms when compared with other FOXO homologues. The establishment of Drosophila as a model for the study of FOXO transcription factors should prove beneficial to determining the biological role of these signaling molecules. The alterations in larval development seen upon overexpression of dFOXO closely mimic the phenotypic effects of starvation, suggesting a role for dFOXO in the response to nutritional adversity. This work has implications in the understanding of cancer and insulin related disorders, such as diabetes and obesity.

Show MeSH

Related in: MedlinePlus

dFOXO responds to dRas2 signaling, but not to inhibitorsof apoptosis. GMR-Gal4 was used to drive the expressionof UAS-dFOXO (A) alone, and in the presence of (B) UAS-p35,(D) UAS-dEGFR, (F) UAS-Ras2V14. UAS-Ras2V14 wasalso expressed in combination with UAS-mFoxo1 (G) and UAS-mFoxo1-AA (H).Scale bars equal 150 μm. Genotypes are: (A) w; GMR-Gal4/+; UAS-dFOXO/+,(B) w; GMR-Gal4/UAS-p35; UAS-dFOXO/+,(C) w; GMR-Gal4/UAS-dEGFR, (D) w; GMR-Gal4/UAS-dEGFR; UAS-dFOXO/+,(E) w; GMR-Gal4/UAS-RasV14, (F) w; GMR-Gal4/UAS-Ras2V14;UAS-dFOXO/+, (G) w; GMR-Gal4, UAS-mFoxo1/UAS-Ras2V14,and (H) w, UAS-mFoxo1-AA/w; GMR-Gal4/ UAS-Ras2V14.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC183841&req=5

Figure 6: dFOXO responds to dRas2 signaling, but not to inhibitorsof apoptosis. GMR-Gal4 was used to drive the expressionof UAS-dFOXO (A) alone, and in the presence of (B) UAS-p35,(D) UAS-dEGFR, (F) UAS-Ras2V14. UAS-Ras2V14 wasalso expressed in combination with UAS-mFoxo1 (G) and UAS-mFoxo1-AA (H).Scale bars equal 150 μm. Genotypes are: (A) w; GMR-Gal4/+; UAS-dFOXO/+,(B) w; GMR-Gal4/UAS-p35; UAS-dFOXO/+,(C) w; GMR-Gal4/UAS-dEGFR, (D) w; GMR-Gal4/UAS-dEGFR; UAS-dFOXO/+,(E) w; GMR-Gal4/UAS-RasV14, (F) w; GMR-Gal4/UAS-Ras2V14;UAS-dFOXO/+, (G) w; GMR-Gal4, UAS-mFoxo1/UAS-Ras2V14,and (H) w, UAS-mFoxo1-AA/w; GMR-Gal4/ UAS-Ras2V14.

Mentions: The lack of ommatidia and mechanosensory bristles caused by dFOXOexpression suggest a reduction in cell number during eye development(Figure 6A).Reduction of cell number can occur through either increased cell death,or decreased of cell proliferation. The Drosophila inhibitors ofapoptosis, Diap1 and Diap2 (data not shown), and the baculovirusinhibitor of apoptosis, p35 (Figure 6B),were unable to rescue the phenotype caused by dFOXO expression.In addition, acridine orange staining of eye imaginal discs expressingdFOXO showed no increase in apoptosis when compared to controls(data not shown). Drosophila Epidermal Growth Factor Receptor (dEGFR)signaling acts to protect differentiated cells from death duringeye development [51]. We thought that thepro-survival effects of dEGFR may be sufficient to suppress thephenotype caused by dFOXO overexpression. Co-expression of dEGFRwith dFOXO, however, does not rescue the dFOXO phenotype as ommatidiaand bristles are clearly still missing (Figure 6D). Conversely, dFOXO doesnot appear to affect the phenotype of dEGFR overexpression as thegeneral disorganization of the ommatidia appears to be the same(Figure 6C).Thus, it appears that these two mechanisms are acting independently.Taken together, these results suggest that dFOXO overexpression doesnot cause cell death during eye development as direct inhibitorsof the apoptotic machinery (p35 and Diap1/2) and a known cell survivalfactor (dEGFR) were unable to rescue the dFOXO phenotype.


Expression of Drosophila FOXO regulates growth and can phenocopy starvation.

Kramer JM, Davidge JT, Lockyer JM, Staveley BE - BMC Dev. Biol. (2003)

dFOXO responds to dRas2 signaling, but not to inhibitorsof apoptosis. GMR-Gal4 was used to drive the expressionof UAS-dFOXO (A) alone, and in the presence of (B) UAS-p35,(D) UAS-dEGFR, (F) UAS-Ras2V14. UAS-Ras2V14 wasalso expressed in combination with UAS-mFoxo1 (G) and UAS-mFoxo1-AA (H).Scale bars equal 150 μm. Genotypes are: (A) w; GMR-Gal4/+; UAS-dFOXO/+,(B) w; GMR-Gal4/UAS-p35; UAS-dFOXO/+,(C) w; GMR-Gal4/UAS-dEGFR, (D) w; GMR-Gal4/UAS-dEGFR; UAS-dFOXO/+,(E) w; GMR-Gal4/UAS-RasV14, (F) w; GMR-Gal4/UAS-Ras2V14;UAS-dFOXO/+, (G) w; GMR-Gal4, UAS-mFoxo1/UAS-Ras2V14,and (H) w, UAS-mFoxo1-AA/w; GMR-Gal4/ UAS-Ras2V14.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC183841&req=5

Figure 6: dFOXO responds to dRas2 signaling, but not to inhibitorsof apoptosis. GMR-Gal4 was used to drive the expressionof UAS-dFOXO (A) alone, and in the presence of (B) UAS-p35,(D) UAS-dEGFR, (F) UAS-Ras2V14. UAS-Ras2V14 wasalso expressed in combination with UAS-mFoxo1 (G) and UAS-mFoxo1-AA (H).Scale bars equal 150 μm. Genotypes are: (A) w; GMR-Gal4/+; UAS-dFOXO/+,(B) w; GMR-Gal4/UAS-p35; UAS-dFOXO/+,(C) w; GMR-Gal4/UAS-dEGFR, (D) w; GMR-Gal4/UAS-dEGFR; UAS-dFOXO/+,(E) w; GMR-Gal4/UAS-RasV14, (F) w; GMR-Gal4/UAS-Ras2V14;UAS-dFOXO/+, (G) w; GMR-Gal4, UAS-mFoxo1/UAS-Ras2V14,and (H) w, UAS-mFoxo1-AA/w; GMR-Gal4/ UAS-Ras2V14.
Mentions: The lack of ommatidia and mechanosensory bristles caused by dFOXOexpression suggest a reduction in cell number during eye development(Figure 6A).Reduction of cell number can occur through either increased cell death,or decreased of cell proliferation. The Drosophila inhibitors ofapoptosis, Diap1 and Diap2 (data not shown), and the baculovirusinhibitor of apoptosis, p35 (Figure 6B),were unable to rescue the phenotype caused by dFOXO expression.In addition, acridine orange staining of eye imaginal discs expressingdFOXO showed no increase in apoptosis when compared to controls(data not shown). Drosophila Epidermal Growth Factor Receptor (dEGFR)signaling acts to protect differentiated cells from death duringeye development [51]. We thought that thepro-survival effects of dEGFR may be sufficient to suppress thephenotype caused by dFOXO overexpression. Co-expression of dEGFRwith dFOXO, however, does not rescue the dFOXO phenotype as ommatidiaand bristles are clearly still missing (Figure 6D). Conversely, dFOXO doesnot appear to affect the phenotype of dEGFR overexpression as thegeneral disorganization of the ommatidia appears to be the same(Figure 6C).Thus, it appears that these two mechanisms are acting independently.Taken together, these results suggest that dFOXO overexpression doesnot cause cell death during eye development as direct inhibitorsof the apoptotic machinery (p35 and Diap1/2) and a known cell survivalfactor (dEGFR) were unable to rescue the dFOXO phenotype.

Bottom Line: Analysis of the wings and eyes of these small flies indicates that the reduction in size is due to decreases in cell size and cell number.Overexpression of dFOXO in the developing eye leads to a characteristic phenotype with reductions in cell size and cell number.This phenotype can be rescued by co-expression of upstream insulin signaling components, dPI3K and dAkt, however, this rescue is not seen when FOXO is mutated to a constitutively active form. dFOXO is conserved in both sequence and regulatory mechanisms when compared with other FOXO homologues.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biology, Memorial University of Newfoundland, St, John's, Newfoundland, (A1B 3X9), Canada. x04jmk@mun.ca

ABSTRACT

Background: Components of the insulin signaling pathway are important regulators of growth. The FOXO (forkhead box, sub-group "O") transcription factors regulate cellular processes under conditions of low levels of insulin signaling. Studies in mammalian cell culture show that activation of FOXO transcription factors causes cell death or cell cycle arrest. The Caenorhabditis elegans homologue of FOXO, Daf-16, is required for the formation of dauer larvae in response to nutritional stress. In addition, FOXO factors have been implicated in stress resistance and longevity.

Results: We have identified the Drosophila melanogaster homologue of FOXO (dFOXO), which is conserved in amino acid sequence compared with the mammalian FOXO homologues and Daf-16. Expression of dFOXO during early larval development causes inhibition of larval growth and alterations in feeding behavior. Inhibition of larval growth is reversible upon discontinuation of dFOXO expression. Expression of dFOXO during the third larval instar or at low levels during development leads to the generation of adults that are reduced in size. Analysis of the wings and eyes of these small flies indicates that the reduction in size is due to decreases in cell size and cell number. Overexpression of dFOXO in the developing eye leads to a characteristic phenotype with reductions in cell size and cell number. This phenotype can be rescued by co-expression of upstream insulin signaling components, dPI3K and dAkt, however, this rescue is not seen when FOXO is mutated to a constitutively active form.

Conclusions: dFOXO is conserved in both sequence and regulatory mechanisms when compared with other FOXO homologues. The establishment of Drosophila as a model for the study of FOXO transcription factors should prove beneficial to determining the biological role of these signaling molecules. The alterations in larval development seen upon overexpression of dFOXO closely mimic the phenotypic effects of starvation, suggesting a role for dFOXO in the response to nutritional adversity. This work has implications in the understanding of cancer and insulin related disorders, such as diabetes and obesity.

Show MeSH
Related in: MedlinePlus