Light-dependent inhibition of clathrin-mediated endocytosis in yeast unveils conserved functions of the AP2 complex

Davia Prischich; Núria Camarero; Javier Encinar Del Dedo; Maria Cambra-Pellejà; Judit Prat; Laura Nevola; Andrés Martín-Quirós; Elena Rebollo; Laura Pastor; Ernest Giralt; María Isabel Geli; Pau Gorostiza

doi:10.1016/j.isci.2023.107899

Light-dependent inhibition of clathrin-mediated endocytosis in yeast unveils conserved functions of the AP2 complex

iScience. 2023 Sep 12;26(10):107899. doi: 10.1016/j.isci.2023.107899. eCollection 2023 Oct 20.

Authors

Davia Prischich^{1

2}, Núria Camarero^{1

2}, Javier Encinar Del Dedo³, Maria Cambra-Pellejà¹, Judit Prat¹, Laura Nevola⁴, Andrés Martín-Quirós¹, Elena Rebollo⁵, Laura Pastor³, Ernest Giralt^{4

6}, María Isabel Geli³, Pau Gorostiza^{1

2

7}

Affiliations

¹ Institute for Bioengineering of Catalonia (IBEC), The Barcelona Institute of Science and Technology, Barcelona, Spain.
² Centro de Investigación Biomédica en Red - Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN), Barcelona, Spain.
³ Department of Cell Biology, Institute for Molecular Biology of Barcelona (IBMB-CSIC), Barcelona, Spain.
⁴ Institute for Research in Biomedicine (IRB Barcelona), The Barcelona Institute of Science and Technology, Barcelona, Spain.
⁵ Molecular Imaging Platform, Institute for Molecular Biology of Barcelona (IBMB-CSIC), Barcelona, Spain.
⁶ Department of Inorganic and Organic Chemistry, University of Barcelona (UB), Barcelona, Spain.
⁷ Catalan Institution for Research and Advanced Studies (ICREA), Barcelona, Spain.

Abstract

Clathrin-mediated endocytosis (CME) is an essential cellular process, conserved among eukaryotes. Yeast constitutes a powerful genetic model to dissect the complex endocytic machinery, yet there is a lack of specific pharmacological agents to interfere with CME in these organisms. TL2 is a light-regulated peptide inhibitor targeting the AP2-β-adaptin/β-arrestin interaction and that can photocontrol CME with high spatiotemporal precision in mammalian cells. Here, we study endocytic protein dynamics by live-cell imaging of the fluorescently tagged coat-associated protein Sla1-GFP, demonstrating that TL2 retains its inhibitory activity in S. cerevisiae spheroplasts. This is despite the β-adaptin/β-arrestin interaction not being conserved in yeast. Our data indicate that the AP2 α-adaptin is the functional target of activated TL2. We identified as interacting partners for the α-appendage, the Eps15 and epsin homologues Ede1 and Ent1. This demonstrates that endocytic cargo loading and sensing can be executed by conserved molecular interfaces, regardless of the proteins involved.

Keywords: Biochemistry; Biological sciences; Cell biology; Molecular biology; Natural sciences.