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TMED10 mediates the loading of neosynthesised Sonic Hedgehog in COPII vesicles for efficient secretion and signalling

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Abstract

The morphogen Sonic Hedgehog (SHH) plays an important role in coordinating embryonic development. Short- and long-range SHH signalling occurs through a variety of membrane-associated and membrane-free forms. However, the molecular mechanisms that govern the early events of the trafficking of neosynthesised SHH in mammalian cells are still poorly understood. Here, we employed the retention using selective hooks (RUSH) system to show that newly-synthesised SHH is trafficked through the classical biosynthetic secretory pathway, using TMED10 as an endoplasmic reticulum (ER) cargo receptor for efficient ER-to-Golgi transport and Rab6 vesicles for Golgi-to-cell surface trafficking. TMED10 and SHH colocalized at ER exit sites (ERES), and TMED10 depletion significantly delays SHH loading onto ERES and subsequent exit leading to significant SHH release defects. Finally, we utilised the Drosophila wing imaginal disc model to demonstrate that the homologue of TMED10, Baiser (Bai), participates in Hedgehog (Hh) secretion and signalling in vivo. In conclusion, our work highlights the role of TMED10 in cargo-specific egress from the ER and sheds light on novel important partners of neosynthesised SHH secretion with potential impact on embryonic development.

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Acknowledgements

We acknowledge the MRI imaging facility, member of the national infrastructure France-BioImaging, for advice and training. We thank Drs. Adrian Salic, Franck Perez, Gregory Lavieu and Robert Blum for the gift of constructs used in this study

Funding

This work was supported by the Agence Nationale de la Recherche (ANR-18-CE13-0003–01) to RG and PT. YB obtained a post-doctoral fellowship from Fondation pour la Recherche Médicale (FRM, SPF202110014043).

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Authors and Affiliations

  1. Institut de Recherche en Infectiologie de Montpellier (IRIM) CNRS, 1919 Route de Mende, 34293, Montpellier, France

    Yonis Bare, Maika S. Deffieu & Raphael Gaudin

  2. Université de Montpellier, 34090, Montpellier, France

    Yonis Bare, Maika S. Deffieu & Raphael Gaudin

  3. Université Côte d’Azur, UMR7277 CNRS, Inserm 1091, Institut de Biologie de Valrose (iBV), Parc Valrose, Nice, France

    Tamás Matusek & Pascal P. Thérond

  4. Laboratoire des Biomolécules (LBM), Département de Chimie, École Normale Supérieure, PSL University, Sorbonne Université, CNRS, 75005, Paris, France

    Sophie Vriz

  5. Faculty of Science, Université de Paris, Paris, France

    Sophie Vriz

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  1. Yonis Bare
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  2. Tamás Matusek
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  3. Sophie Vriz
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  4. Maika S. Deffieu
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  5. Pascal P. Thérond
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  6. Raphael Gaudin
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Contributions

Conceptualisation: YB, RG. Methodology: YB, TM, SV, PT, MSD, RG. Investigation: YB, TM, PT, RG. Analysis: YB, TM, PT, RG. Supervision: PT, RG. Writing—original draft: YB, RG. Writing—review & editing: YB, TM, SV, PT, MSD, RG.

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Correspondence to Raphael Gaudin.

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Video S1: Dual-colour live-cell imaging of SHH-RUSH trafficking through the Golgi complex, related to Figure 1. SVG-A cell co-transfected with SHH-RUSH (magenta) and the medial Golgi marker, Mannosidase-II-EGFP (green) and imaged using 3D spinning disk confocal microscopy. Acquisition started 5 mins after the addition of 40 µM biotin. Frames were taken every 5 mins for 90 mins. Scale bar – 10µm

Video S2: Zoomed inset of Video S1, related to Figure 1. SHH-RUSH (magenta) transported into and exits from the Golgi (green). Frames taken every 5 mins for 90 min. Scale bar – 5 µm

Video S3: Two colour TIRF imaging of SHH-RUSH and GFP-Rab6a, related to Figure 2. SVG-A cell co-transfected with SHH-RUSH (magenta) and GFP-Rab6a (green). Acquisition started 30 mins after the addition of 40 µM biotin. Frames were taken every 2 seconds for an additional 30 mins. Scale bar – 10 µM

Video S4: Zoomed inset of Video S3, related to Figure 2. Video corresponds to 1890s to 1920s post biotin addition and demonstrates a double-positive vesicle of SHH-RUSH (magenta) and GFP-Rab6a (green) arriving at the plasma membrane. Scale bar – 2 µm

Video S5: Dual-colour live-cell imaging of SHH-RUSH and GFP-TMED10, related to Figure 3. SVG-A cell co-transfected with SHH-RUSH (magenta) and GFP-TMED10 (green) and imaged using 3D spinning disk confocal microscopy. Acquisition started immediately after the addition of 40 µM biotin. Frames were taken every 500 ms for 10 mins. Scale bar – 10 µM

Video S6: Zoomed inset of Video S5, related to Figure 3. A double-positive vesicle of SHH-RUSH (magenta) and GFP-TMED10 (green) trafficking towards a perinuclear region. Video corresponds to 49s to 99s post biotin addition. Scale bar – 5 µm

Video S7: Dual colour live-cell imaging of SHH-RUSH and SEC24D-EGFP, related to Figure 4. SVG-A cells co-transfected with SHH-RUSH (magenta) and SEC24D-EGFP (green) and imaged using 3D spinning disk confocal microscopy. Acquisition started immediately after the addition of 40 µM biotin. Frames were taken every 2 s for 15 min. Scale bar – 10 µm

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Bare, Y., Matusek, T., Vriz, S. et al. TMED10 mediates the loading of neosynthesised Sonic Hedgehog in COPII vesicles for efficient secretion and signalling. Cell. Mol. Life Sci. 80, 266 (2023). https://doi.org/10.1007/s00018-023-04918-1

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  • DOI: https://doi.org/10.1007/s00018-023-04918-1

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