Limits...
A drosophila model for EGFR-Ras and PI3K-dependent human glioma.

Read RD, Cavenee WK, Furnari FB, Thomas JB - PLoS Genet. (2009)

Bottom Line: This network acts synergistically to coordinately stimulate cell cycle entry and progression, protein translation, and inappropriate cellular growth and migration.In particular, we found that the fly orthologs of CyclinE, Cdc25, and Myc are key rate-limiting genes required for glial neoplasia.Moreover, orthologs of Sin1, Rictor, and Cdk4 are genes required only for abnormal neoplastic glial proliferation but not for glial development.

View Article: PubMed Central - PubMed

Affiliation: Molecular Neurobiology Laboratory, Salk Institute for Biological Studies, La Jolla, California, United States of America. rread@salk.edu

ABSTRACT
Gliomas, the most common malignant tumors of the nervous system, frequently harbor mutations that activate the epidermal growth factor receptor (EGFR) and phosphatidylinositol-3 kinase (PI3K) signaling pathways. To investigate the genetic basis of this disease, we developed a glioma model in Drosophila. We found that constitutive coactivation of EGFR-Ras and PI3K pathways in Drosophila glia and glial precursors gives rise to neoplastic, invasive glial cells that create transplantable tumor-like growths, mimicking human glioma. Our model represents a robust organotypic and cell-type-specific Drosophila cancer model in which malignant cells are created by mutations in signature genes and pathways thought to be driving forces in a homologous human cancer. Genetic analyses demonstrated that EGFR and PI3K initiate malignant neoplastic transformation via a combinatorial genetic network composed primarily of other pathways commonly mutated or activated in human glioma, including the Tor, Myc, G1 Cyclins-Cdks, and Rb-E2F pathways. This network acts synergistically to coordinately stimulate cell cycle entry and progression, protein translation, and inappropriate cellular growth and migration. In particular, we found that the fly orthologs of CyclinE, Cdc25, and Myc are key rate-limiting genes required for glial neoplasia. Moreover, orthologs of Sin1, Rictor, and Cdk4 are genes required only for abnormal neoplastic glial proliferation but not for glial development. These and other genes within this network may represent important therapeutic targets in human glioma.

Show MeSH

Related in: MedlinePlus

Coactivation of EGFR-Ras and PI3K in Drosophila glia causes neoplasia.(A) Optical projections of whole brain-ventral nerve cord complexes from late 3rd instar larvae approximately 130 hr AED, displayed at the same scale. Dorsal view; anterior up. Glia are labeled with CD8GFP (green) driven by repo-Gal4. Each brain is composed of 2 symmetrical hemispheres attached to the ventral nerve cord (VNC). In repo>dEGFRλ;dp110CAAX larvae, both brain hemispheres and the VNC are enlarged and elongated relative to other genotypes. (B–H) 2 µm optical sections of larval brain hemispheres from late 3rd instar larvae approximately 130 hr AED, displayed at the same scale. 20 µm scale bars. Frontal sections, midway through brains. Anterior up; midline to left. Glial cell nuclei are labeled with Repo (red). Glial cell bodies and membranes are labeled with CD8GFP (green) driven by repo-Gal4. In repo>dEGFRλ;dp110CAAX brains (B) there is a dramatic increase in number of glial nuclei relative to wild-type (C), while in repo>dRasN17;dEGFRλ;dp110CAAX (D) and repo>PTEN; dEGFRλ;dp110CAAX (E) brains there are fewer excess glial nuclei. In wild-type animals (C), glia extend processes throughout the entire brain, enveloping nearly every cell with glial membranes. In contrast, dEGFRλ;dp110CAAX glia (B) show loss of fine projections and instead have compact cell bodies. Brains are counter-stained with anti-HRP (blue), which reveals neuronal fiber tracts (neuropil) at high intensity and some cell bodies of neurons and neuronal precursors at low intensity. HRP staining varies slightly according to exact plane of section, brain orientation, and mutant phenotype. Dark areas within brains contain unstained cells, which are typically optic lobe neural precursors. Genotypes: (A) Clockwise from top left: repo-Gal4 UAS-CD8GFP/+, UAS-dEGFRλ UAS-dp110CAAX/+; repo-Gal4 UAS-CD8GFP/+, UAS-dp110CAAX/+; repo-Gal4 UAS-CD8GFP/+, UAS-dEGFRλ/+; repo-Gal4 UAS-CD8GFP/+, (B) UAS-dEGFRλ UAS-dp110CAAX/+; repo-Gal4 UAS-CD8GFP/+, (C) repo-Gal4 UAS-CD8GFP/+, (D) UAS-dEGFRλ UAS-dp110CAAX/UAS-dRas85DN17; repo-Gal4 UAS-CD8GFP/+, (E) UAS-dEGFRλ UAS-dp110CAAX/+; UAS-PTEN/+; repo-Gal4 UAS-CD8GFP/+, (F) UAS-dEGFRλ/+; repo-Gal4 UAS-CD8GFP/+, (G) repo-Gal4 UAS-CD8GFP/UAS-dRas85DV12, (H) UAS-dp110CAAX/+; repo-Gal4 UAS-CD8GFP/+.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC2636203&req=5

pgen-1000374-g001: Coactivation of EGFR-Ras and PI3K in Drosophila glia causes neoplasia.(A) Optical projections of whole brain-ventral nerve cord complexes from late 3rd instar larvae approximately 130 hr AED, displayed at the same scale. Dorsal view; anterior up. Glia are labeled with CD8GFP (green) driven by repo-Gal4. Each brain is composed of 2 symmetrical hemispheres attached to the ventral nerve cord (VNC). In repo>dEGFRλ;dp110CAAX larvae, both brain hemispheres and the VNC are enlarged and elongated relative to other genotypes. (B–H) 2 µm optical sections of larval brain hemispheres from late 3rd instar larvae approximately 130 hr AED, displayed at the same scale. 20 µm scale bars. Frontal sections, midway through brains. Anterior up; midline to left. Glial cell nuclei are labeled with Repo (red). Glial cell bodies and membranes are labeled with CD8GFP (green) driven by repo-Gal4. In repo>dEGFRλ;dp110CAAX brains (B) there is a dramatic increase in number of glial nuclei relative to wild-type (C), while in repo>dRasN17;dEGFRλ;dp110CAAX (D) and repo>PTEN; dEGFRλ;dp110CAAX (E) brains there are fewer excess glial nuclei. In wild-type animals (C), glia extend processes throughout the entire brain, enveloping nearly every cell with glial membranes. In contrast, dEGFRλ;dp110CAAX glia (B) show loss of fine projections and instead have compact cell bodies. Brains are counter-stained with anti-HRP (blue), which reveals neuronal fiber tracts (neuropil) at high intensity and some cell bodies of neurons and neuronal precursors at low intensity. HRP staining varies slightly according to exact plane of section, brain orientation, and mutant phenotype. Dark areas within brains contain unstained cells, which are typically optic lobe neural precursors. Genotypes: (A) Clockwise from top left: repo-Gal4 UAS-CD8GFP/+, UAS-dEGFRλ UAS-dp110CAAX/+; repo-Gal4 UAS-CD8GFP/+, UAS-dp110CAAX/+; repo-Gal4 UAS-CD8GFP/+, UAS-dEGFRλ/+; repo-Gal4 UAS-CD8GFP/+, (B) UAS-dEGFRλ UAS-dp110CAAX/+; repo-Gal4 UAS-CD8GFP/+, (C) repo-Gal4 UAS-CD8GFP/+, (D) UAS-dEGFRλ UAS-dp110CAAX/UAS-dRas85DN17; repo-Gal4 UAS-CD8GFP/+, (E) UAS-dEGFRλ UAS-dp110CAAX/+; UAS-PTEN/+; repo-Gal4 UAS-CD8GFP/+, (F) UAS-dEGFRλ/+; repo-Gal4 UAS-CD8GFP/+, (G) repo-Gal4 UAS-CD8GFP/UAS-dRas85DV12, (H) UAS-dp110CAAX/+; repo-Gal4 UAS-CD8GFP/+.

Mentions: Glial-specific coactivation of EGFR-Ras and PI3K stimulated glial neoplasia, giving rise to CNS enlargement and malformation, neurologic defects, and late larval lethality. repo-Gal4-driven co-overexpression of activated dEGFR (dEGFRλ) and dp110 (dp110CAAX) induced progressive accumulation of ∼50-fold excess glia (Figure 1A and 1B) [14],[15]. dEGFRλ is a constitutively active dEGFR variant in which a lambda dimerization domain replaces the extracellular domain [14] (Figure S1). Co-overexpression of combinations of dEGFRλ and core components of the PI3K pathway, such as dAkt, induced phenotypes similar to repo>dEGFRλ;dp110CAAX, although phenotypes varied somewhat depending on strength of pathway activation and transgene expression (Figure S2 and Table S1). Dramatic glial overgrowth also occurred upon co-overexpression of constitutively active dRas (dRas85DV12) or its effector dRaf (dRafgof) with dp110CAAX, dAkt, or a dPTENdsRNA, which partially knocked-down dPTEN (Figure S2 and Table S1). Finally, glial overgrowth in repo>dEGFRλ;dp110CAAX larvae was strongly suppressed by co-overexpression of dPTEN or more moderately by dominant negative dRas85D (dRas85DN17) (Figure 1D and 1E), indicating that Ras activity and excess phospho-inositols are essential for neoplasia.


A drosophila model for EGFR-Ras and PI3K-dependent human glioma.

Read RD, Cavenee WK, Furnari FB, Thomas JB - PLoS Genet. (2009)

Coactivation of EGFR-Ras and PI3K in Drosophila glia causes neoplasia.(A) Optical projections of whole brain-ventral nerve cord complexes from late 3rd instar larvae approximately 130 hr AED, displayed at the same scale. Dorsal view; anterior up. Glia are labeled with CD8GFP (green) driven by repo-Gal4. Each brain is composed of 2 symmetrical hemispheres attached to the ventral nerve cord (VNC). In repo>dEGFRλ;dp110CAAX larvae, both brain hemispheres and the VNC are enlarged and elongated relative to other genotypes. (B–H) 2 µm optical sections of larval brain hemispheres from late 3rd instar larvae approximately 130 hr AED, displayed at the same scale. 20 µm scale bars. Frontal sections, midway through brains. Anterior up; midline to left. Glial cell nuclei are labeled with Repo (red). Glial cell bodies and membranes are labeled with CD8GFP (green) driven by repo-Gal4. In repo>dEGFRλ;dp110CAAX brains (B) there is a dramatic increase in number of glial nuclei relative to wild-type (C), while in repo>dRasN17;dEGFRλ;dp110CAAX (D) and repo>PTEN; dEGFRλ;dp110CAAX (E) brains there are fewer excess glial nuclei. In wild-type animals (C), glia extend processes throughout the entire brain, enveloping nearly every cell with glial membranes. In contrast, dEGFRλ;dp110CAAX glia (B) show loss of fine projections and instead have compact cell bodies. Brains are counter-stained with anti-HRP (blue), which reveals neuronal fiber tracts (neuropil) at high intensity and some cell bodies of neurons and neuronal precursors at low intensity. HRP staining varies slightly according to exact plane of section, brain orientation, and mutant phenotype. Dark areas within brains contain unstained cells, which are typically optic lobe neural precursors. Genotypes: (A) Clockwise from top left: repo-Gal4 UAS-CD8GFP/+, UAS-dEGFRλ UAS-dp110CAAX/+; repo-Gal4 UAS-CD8GFP/+, UAS-dp110CAAX/+; repo-Gal4 UAS-CD8GFP/+, UAS-dEGFRλ/+; repo-Gal4 UAS-CD8GFP/+, (B) UAS-dEGFRλ UAS-dp110CAAX/+; repo-Gal4 UAS-CD8GFP/+, (C) repo-Gal4 UAS-CD8GFP/+, (D) UAS-dEGFRλ UAS-dp110CAAX/UAS-dRas85DN17; repo-Gal4 UAS-CD8GFP/+, (E) UAS-dEGFRλ UAS-dp110CAAX/+; UAS-PTEN/+; repo-Gal4 UAS-CD8GFP/+, (F) UAS-dEGFRλ/+; repo-Gal4 UAS-CD8GFP/+, (G) repo-Gal4 UAS-CD8GFP/UAS-dRas85DV12, (H) UAS-dp110CAAX/+; repo-Gal4 UAS-CD8GFP/+.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2636203&req=5

pgen-1000374-g001: Coactivation of EGFR-Ras and PI3K in Drosophila glia causes neoplasia.(A) Optical projections of whole brain-ventral nerve cord complexes from late 3rd instar larvae approximately 130 hr AED, displayed at the same scale. Dorsal view; anterior up. Glia are labeled with CD8GFP (green) driven by repo-Gal4. Each brain is composed of 2 symmetrical hemispheres attached to the ventral nerve cord (VNC). In repo>dEGFRλ;dp110CAAX larvae, both brain hemispheres and the VNC are enlarged and elongated relative to other genotypes. (B–H) 2 µm optical sections of larval brain hemispheres from late 3rd instar larvae approximately 130 hr AED, displayed at the same scale. 20 µm scale bars. Frontal sections, midway through brains. Anterior up; midline to left. Glial cell nuclei are labeled with Repo (red). Glial cell bodies and membranes are labeled with CD8GFP (green) driven by repo-Gal4. In repo>dEGFRλ;dp110CAAX brains (B) there is a dramatic increase in number of glial nuclei relative to wild-type (C), while in repo>dRasN17;dEGFRλ;dp110CAAX (D) and repo>PTEN; dEGFRλ;dp110CAAX (E) brains there are fewer excess glial nuclei. In wild-type animals (C), glia extend processes throughout the entire brain, enveloping nearly every cell with glial membranes. In contrast, dEGFRλ;dp110CAAX glia (B) show loss of fine projections and instead have compact cell bodies. Brains are counter-stained with anti-HRP (blue), which reveals neuronal fiber tracts (neuropil) at high intensity and some cell bodies of neurons and neuronal precursors at low intensity. HRP staining varies slightly according to exact plane of section, brain orientation, and mutant phenotype. Dark areas within brains contain unstained cells, which are typically optic lobe neural precursors. Genotypes: (A) Clockwise from top left: repo-Gal4 UAS-CD8GFP/+, UAS-dEGFRλ UAS-dp110CAAX/+; repo-Gal4 UAS-CD8GFP/+, UAS-dp110CAAX/+; repo-Gal4 UAS-CD8GFP/+, UAS-dEGFRλ/+; repo-Gal4 UAS-CD8GFP/+, (B) UAS-dEGFRλ UAS-dp110CAAX/+; repo-Gal4 UAS-CD8GFP/+, (C) repo-Gal4 UAS-CD8GFP/+, (D) UAS-dEGFRλ UAS-dp110CAAX/UAS-dRas85DN17; repo-Gal4 UAS-CD8GFP/+, (E) UAS-dEGFRλ UAS-dp110CAAX/+; UAS-PTEN/+; repo-Gal4 UAS-CD8GFP/+, (F) UAS-dEGFRλ/+; repo-Gal4 UAS-CD8GFP/+, (G) repo-Gal4 UAS-CD8GFP/UAS-dRas85DV12, (H) UAS-dp110CAAX/+; repo-Gal4 UAS-CD8GFP/+.
Mentions: Glial-specific coactivation of EGFR-Ras and PI3K stimulated glial neoplasia, giving rise to CNS enlargement and malformation, neurologic defects, and late larval lethality. repo-Gal4-driven co-overexpression of activated dEGFR (dEGFRλ) and dp110 (dp110CAAX) induced progressive accumulation of ∼50-fold excess glia (Figure 1A and 1B) [14],[15]. dEGFRλ is a constitutively active dEGFR variant in which a lambda dimerization domain replaces the extracellular domain [14] (Figure S1). Co-overexpression of combinations of dEGFRλ and core components of the PI3K pathway, such as dAkt, induced phenotypes similar to repo>dEGFRλ;dp110CAAX, although phenotypes varied somewhat depending on strength of pathway activation and transgene expression (Figure S2 and Table S1). Dramatic glial overgrowth also occurred upon co-overexpression of constitutively active dRas (dRas85DV12) or its effector dRaf (dRafgof) with dp110CAAX, dAkt, or a dPTENdsRNA, which partially knocked-down dPTEN (Figure S2 and Table S1). Finally, glial overgrowth in repo>dEGFRλ;dp110CAAX larvae was strongly suppressed by co-overexpression of dPTEN or more moderately by dominant negative dRas85D (dRas85DN17) (Figure 1D and 1E), indicating that Ras activity and excess phospho-inositols are essential for neoplasia.

Bottom Line: This network acts synergistically to coordinately stimulate cell cycle entry and progression, protein translation, and inappropriate cellular growth and migration.In particular, we found that the fly orthologs of CyclinE, Cdc25, and Myc are key rate-limiting genes required for glial neoplasia.Moreover, orthologs of Sin1, Rictor, and Cdk4 are genes required only for abnormal neoplastic glial proliferation but not for glial development.

View Article: PubMed Central - PubMed

Affiliation: Molecular Neurobiology Laboratory, Salk Institute for Biological Studies, La Jolla, California, United States of America. rread@salk.edu

ABSTRACT
Gliomas, the most common malignant tumors of the nervous system, frequently harbor mutations that activate the epidermal growth factor receptor (EGFR) and phosphatidylinositol-3 kinase (PI3K) signaling pathways. To investigate the genetic basis of this disease, we developed a glioma model in Drosophila. We found that constitutive coactivation of EGFR-Ras and PI3K pathways in Drosophila glia and glial precursors gives rise to neoplastic, invasive glial cells that create transplantable tumor-like growths, mimicking human glioma. Our model represents a robust organotypic and cell-type-specific Drosophila cancer model in which malignant cells are created by mutations in signature genes and pathways thought to be driving forces in a homologous human cancer. Genetic analyses demonstrated that EGFR and PI3K initiate malignant neoplastic transformation via a combinatorial genetic network composed primarily of other pathways commonly mutated or activated in human glioma, including the Tor, Myc, G1 Cyclins-Cdks, and Rb-E2F pathways. This network acts synergistically to coordinately stimulate cell cycle entry and progression, protein translation, and inappropriate cellular growth and migration. In particular, we found that the fly orthologs of CyclinE, Cdc25, and Myc are key rate-limiting genes required for glial neoplasia. Moreover, orthologs of Sin1, Rictor, and Cdk4 are genes required only for abnormal neoplastic glial proliferation but not for glial development. These and other genes within this network may represent important therapeutic targets in human glioma.

Show MeSH
Related in: MedlinePlus