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First published online 1 November 2006
doi: 10.1242/dev.02659

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Cytoplasmic activated protein kinase Akt regulates lipid-droplet accumulation in Drosophila nurse cells

Department of Physiology, Anatomy and Genetics, Le Gros Clark Building, University of Oxford, South Parks Road, Oxford OX1 3QX, UK.

View larger version (101K): [in a new window]   Fig. 1. Increased IIS produces lipid-droplet accumulation defects in nurse cells. (A,B) Nile Red-positive small neutral lipid droplets (red) accumulate in wild-type nurse cells, particularly in a perinuclear region (arrowheads in B), which appears to co-localize with the endoplasmic reticulum (Teixeira et al., 2003). By contrast, non-GFP-labelled (not green) Pten mutant cells (D-I) contain large aggregated lipid droplets of up to 15 µm diameter (arrowheads in E,H; shown at high magnification in F,I) and contain far fewer small droplets. (C) This phenotype is completely suppressed in Pten mutant animals rescued by a specific combination of Akt alleles (Stocker et al., 2002). (J-O) In some egg chambers, overexpression of Dp110 in nurse cells induces sporadic formation of superficial, elongated lipid-containing structures close to the plasma membrane (arrowheads in K,N). The profile of these cells is outlined by intense GFP expression in overlying somatic stretched follicular cells (green in L,O; see also deeper section in Fig. 3C). Scale bars: 40 µm in A-C,F,I; 40 µm in E for D,E; 40 µm in G for G,H; 40 µm in J for J,K,L; and 40 µm in M for M,N.O.

View larger version (133K): [in a new window]   Fig. 2. Cytoplasmic activated Akt is greatly increased in Pten-mutant nurse cells. (A,B) Nurse cells from wild-type egg chambers show a modest accumulation of activated P-Akt (blue) in the cytoplasm. The actin cytoskeleton of these cells is labelled with TRITC-phalloidin (red). (E-L) Non-GFP-labelled (not green in H,L) Pten mutant cells have no obvious cortical cytoskeletal abnormalities (G,K), but contain much higher levels of cytoplasmic P-Akt (F,J). (C,D) Elevated levels of P-Akt are observed in the cytoplasm of mutant cells but not in nuclei, which are stained with propidium iodide (red) in this egg chamber. The positions of Pten mutant nuclei are marked in red in D. (M-P) Nurse cells overexpressing Dp110 produce increased cell-surface P-Akt (N), co-localizing with cortical actin (O). Intensely GFP-positive nuclei of peripheral Dp110-overexpressing stretched follicle cells overlying the nurse cells are also observed in this image (P, arrowheads). Scale bar: 40 µm in A for A,B; 40 µm in D for C,D; 40 µm in E for E-H; 40 µm in I for I-L; and 40 µm in M for M-P.

View larger version (98K): [in a new window]   Fig. 3. Cytoplasmic activated Akt controls Lsd2 expression and specific IIS-dependent functions. (A,B) Perilipin homologue LSD2 (B, blue) is expressed relatively homogeneously in the nurse cell cytoplasm, which is stained for lipid with Nile Red (A, red). (C-I) Nile Red-positive droplets (red; arrows) in Pten-mutant (GFP-negative cell in E-I; I,H are magnified views of anterior nurse cell in E) and Dp110-overexpressing (C,D) nurse cells exclude the perilipin homologue LSD2. In G,H, Nile Red is also detected in the green channel in intensely stained large lipid droplets. LSD2 levels are highly elevated in Pten mutant cells (F,I); normal perilipin levels are not visible at this low imaging sensitivity. There is evidence for elevated LSD2 accumulation around some lipid droplets (arrow in H). Arrowhead in I points to a region in the mutant cell that does not contain LSD2 - which is presumably the nucleus. (J) Schematic showing the established InR/PI3-kinase/Akt signalling cassette, involving InR-dependent activation of cell-surface Akt (orange), a process antagonized by PTEN. Our data reveal a role for cytoplasmic activated Akt (pink) in regulating the accumulation of large lipid droplets (red) in Drosophila nurse cells via a mechanism that, in part, involves LSD2. Note potential negative-feedback mechanisms (green), which might be controlled by cytoplasmic as well as surface P-Akt, and could therefore induce differential sensitivity to insulin in different subcellular domains. Scale bars: 40 µm.