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Lipoprotein particles are required for Hedgehog and Wingless signalling

Abstract

Wnt and Hedgehog family proteins are secreted signalling molecules (morphogens) that act at both long and short range to control growth and patterning during development. Both proteins are covalently modified by lipid, and the mechanism by which such hydrophobic molecules might spread over long distances is unknown. Here we show that Wingless, Hedgehog and glycophosphatidylinositol-linked proteins copurify with lipoprotein particles, and co-localize with them in the developing wing epithelium of Drosophila. In larvae with reduced lipoprotein levels, Hedgehog accumulates near its site of production, and fails to signal over its normal range. Similarly, the range of Wingless signalling is narrowed. We propose a novel function for lipoprotein particles, in which they act as vehicles for the movement of lipid-linked morphogens and glycophosphatidylinositol-linked proteins.

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Figure 1: Lipid-linked proteins co-fractionate with lipophorin.
Figure 2: Wingless and Hedgehog co-localize with Alexa488lipophorin.
Figure 3: Lipophorin–RNAi perturbs lipid transport.
Figure 4: Lipophorin–RNAi alters Hedgehog distribution and signalling.
Figure 5: Hedgehog signalling is unaffected by lipid-depletion.
Figure 6: Lipophorin–RNAi narrows the range of Wingless signalling.

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Acknowledgements

We thank T. Kurzchalia for advice regarding lipid depletion. We thank S. Cohen, I. Guerrero, P. Ingham, Y. Hiromi, M. Horscht, John Incardona and R. White for gifts of antibodies, and G. Griffiths for the CD63:GFP fusion construct. We are grateful to A. Mahmoud and S. Bowman for developing CFP:Rab5-expressing flies, and to V. Greco for helping to initiate these studies. We thank D. Backasch for performing embryo injections. K. Simons, M. Zerial, T. Kurzchalia and C. Dahmann provided comments on the manuscript.Author contributions This work has been a collaborative effort between the groups of C. Thiele and S. Eaton, the Thiele laboratory contributing biochemical expertise and the Eaton laboratory expertise in working with Drosophila. First authors appear in alphabetical order.

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Correspondence to Christoph Thiele or Suzanne Eaton.

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Supplementary information

Supplementary Figure S1

Supplementary Figure S1 describes and characterizes the marker used for exosomes (CD63:GFP) and Lipophorin (anti-ApoLI and anti-ApoLII). (JPG 81 kb)

Supplementary Figure S2

Hedgehog and Fasciclin I can be co-immunoprecipitated with Lipophorin from the top, low-density fraction of KBr gradients. (PDF 240 kb)

Supplementary Figure S3

The rate of reduction of ApoLI protein levels by Lipophorin-RNAi. (PDF 196 kb)

Supplementary Figure S4

Insulin signalling is not reduced in Lipophorin RNAi discs and that apoptosis is not significantly elevated. (JPG 47 kb)

Supplementary Figure S5

Hedgehog and Patched in Lipophorin-RNAi larvae accumulate in endosomes. (JPG 44 kb)

Supplementary Figure S6

Quantification of Distalless staining intensity in individual wild type and Lipophorin-RNAi discs. (JPG 24 kb)

Supplementary Figure S7

Hedgehog and Patched accumulation in Lipophorin RNAi discs can be reversed by adding purified Lipophorin particles to explanted discs. (PDF 1817 kb)

Supplementary Figure S7

Legend to accompany the above figure. (DOC 23 kb)

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Panáková, D., Sprong, H., Marois, E. et al. Lipoprotein particles are required for Hedgehog and Wingless signalling. Nature 435, 58–65 (2005). https://doi.org/10.1038/nature03504

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