QUICK SEARCH:   [advanced] Author: Keyword(s):
Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents

First published online 12 September 2007
doi: 10.1242/jcs.005926

This Article Summary Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Related articles in JCS Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Devergne, O. Articles by Noselli, S. Search for Related Content PubMed PubMed Citation Articles by Devergne, O. Articles by Noselli, S. Social Bookmarking                         What's this?

The endocytic control of JAK/STAT signalling in Drosophila

Institute of Developmental Biology and Cancer, CNRS-UMR 6543, University of Nice Sophia-Antipolis, Parc Valrose, 06108 Nice cedex 2, France

View larger version (62K): [in this window] [in a new window]   Fig. 1. Dome internalization is dependent on Upd ligand binding. (A) Schematic representation of a stage 8 egg chamber. The gradient of apically located vesicles is represented by red dots. Polar cells are in black. (B,C) Dome is expressed in a gradient (white wedges) of apically located vesicles at the anterior (B) and posterior (C) poles of the egg chamber. The abundance and distribution of Dome vesicles depend on the distance from the polar cells (marked with A101--galactosidase; green), the source of the Upd ligand. (D) When polar cells (arrow; marked with Fas III, membrane marker, green) are wild type for Upd (i.e. nuclear-GFP positive), Dome is present in intracellular vesicles. (E) By contrast, when polar cells (arrow) are mutant for Upd (i.e. nuclear-GFP negative), Dome is not detected in intracellular vesicles. In D and E, polar cells are marked using the FasIII antibody (membrane green staining). The clonal marker is GFP (nuclear green staining). Bars, 10 µm.

View larger version (50K): [in this window] [in a new window]   Fig. 2. Ligand diffusion and internalization leads to STAT nuclear localization in S2 cells. (A-C) Cells expressing either Upd-GFP or Dome were mixed and cultured together for 0.5-4 hours. The 0 hour time-point corresponds to cells transfected with Dome only. Upd-GFP diffuses in the medium and binds to Dome-expressing cells in a time-dependent manner. Cells that do not express Dome are outlined by broken lines. Upd-GFP is first found at the membrane of Dome-expressing cells, and (B) becomes internalized in Dome-containing vesicles (arrows) after 6 hours. (C) Binding of Upd-GFP leads to the nuclear accumulation of Stat in a Dome- and time-dependent manner. Dome-expressing cells in C are stained with a -galactosidase marker (purple). Bars, 10 µm.

View larger version (80K): [in this window] [in a new window]   Fig. 3. Nature of Dome-containing vesicles. (A) Co-localization of Dome with the human transferrin receptor (hTfR), which was expressed in border cells using the slbo-Gal4 driver. (B-D) Dome co-localizes with specific markers (green) of the early (B, 2XFYVE-GFP; C, Rab5-GFP) and late (D, Rab7-GFP) endosomal compartments in the border cells. Arrowheads in insets show double-labelled intracellular vesicles. Bars, 10 µm.

View larger version (99K): [in this window] [in a new window]   Fig. 4. Localization of Dome in mutants with affected intracellular trafficking. (A-D) Immunostaining using anti-Dome antibodies (red) showing Dome localization in Clathrin heavy chain (Chc, A), rab5 (B) and dor (C,D) mutant cells. (A) Mosaic egg chamber containing Chc mutant clones (mutant cells are shown enclosed by a broken line and do not express the GFP clonal marker, green). In Chc mutant clones, Dome is concentrated at the membrane, indicating that Dome is internalized in clathrin-coated vesicles. (B) Mutations in rab5 induce Dome accumulation in follicle cells. Notice the formation of a multi-layered epithelium, as previously observed in follicle cells of rab5 mutant clones (Lu and Bilder, 2005). (C,D) Loss of dor function leads to a strong accumulation of Dome in large intracellular structures corresponding to enlarged multivesicular bodies (MVBs). This phenotype is only seen in dor mutant cells close to the ligand source at the posterior (C) and anterior (D) poles of the egg chamber. Bars, 10 µm.

View larger version (79K): [in this window] [in a new window]   Fig. 5. Endocytosis controls STAT nuclear localization and pathway activity. (A-G) STAT intracellular localization was analyzed using anti-STAT antibodies (red) to measure JAK/STAT pathway activation in wild-type egg chambers (A) and in dome (B), Chc (C), rab5 (D), rab11 (E), dor (F) and Hrs (G) mutant backgrounds. In cells mutant for Chc (C), rab5 (D) or Hrs (G), Stat disappears from the cytoplasm and nucleus, as in dome mutant cells (B), indicating that JAK/STAT signalling is strongly affected in these conditions. Mutations in dor (F) also reduce JAK/STAT signalling but to a lesser extent, as evidenced by weak residual STAT staining in the periphery of the nucleus (F). By contrast, cells mutant for rab11 (E) show normal Stat staining, suggesting that it is not required for JAK/STAT signalling. Broken lines mark the mutant cells. Bars, 10 µm.

View larger version (64K): [in this window] [in a new window]   Fig. 6. Expression of the STAT target gene pointed (pnt) is affected in Chc and dor mutant cells. Expression of STAT (blue) and the activity of the JAK/STAT pathway, monitored using a pointed-lacZ reporter gene (red). Staining of wild-type (A-A''), Chc1 (B-B'') and dor8 (C-C'') egg chambers. The clonal marker is GFP (green in B-C''). Broken lines mark the mutant cells. Bar, 10 µm.

View larger version (16K): [in this window] [in a new window]   Fig. 7. Model of endocytic control of Dome trafficking and JAK/STAT signalling. Dome intracellular trafficking in egg chambers. Following ligand binding, Dome is directed to the early endosome and is then sorted to the lysosome, via the multivesicular body (MVB) and the late endosomal compartment. The off/on/off model (right): to become active, the JAK/STAT signalling pathway requires the formation of the ligand-receptor complex at the cell membrane and the formation of Chc-containing budding vesicles. These two conditions allow JAK/STAT signalling to be activated only when the receptor is on its way to degradation. Finally, the degradation of Dome in the MVB/lysosome eliminates active receptors, thus dynamically controlling JAK/STAT pathway activity.

CiteULike

Complore

Connotea

Del.icio.us

Digg

Reddit

Technorati

Twitter