Arabidopsis CaLB1 undergoes phase separation with the ESCRT protein ALIX and modulates autophagosome maturation

• Published in: Nature communications 2024-06, Vol.15 (1), p.5188-20, Article 5188
• Main Authors: Mosesso, Niccolò, Lerner, Niharika Savant, Bläske, Tobias, Groh, Felix, Maguire, Shane, Niedermeier, Marie Laura, Landwehr, Eliane, Vogel, Karin, Meergans, Konstanze, Nagel, Marie-Kristin, Drescher, Malte, Stengel, Florian, Hauser, Karin, Isono, Erika
• Format: Article
• Language: English
• Subjects: 14/1; 14/19; 14/28; 14/35; 45/29; 45/44; 631/449/2661/1797; 631/449/448/2024; 631/449/448/2189; 82/29; 82/80; 82/83; Arabidopsis; Arabidopsis - genetics; Arabidopsis - metabolism; Arabidopsis Proteins - genetics; Arabidopsis Proteins - metabolism; Autophagosomes - metabolism; Autophagy; Autophagy-Related Protein 8 Family - genetics; Autophagy-Related Protein 8 Family - metabolism; Calcium ions; Calcium-binding protein; Calcium-Binding Proteins - genetics; Calcium-Binding Proteins - metabolism; Cell membranes; Cleavage; Endosomal Sorting Complexes Required for Transport - metabolism; Filaments; Humanities and Social Sciences; Lipids; Maturation; Membranes; Molecular modelling; multidisciplinary; Phagosomes; Phase Separation; Phosphatidylinositol Phosphates - metabolism; Phospholipids; Protein X; Proteins; Salts; Science; Science (multidisciplinary); Vacuoles - metabolism
• ISSN: 2041-1723; 2041-1723
• DOI: 10.1038/s41467-024-49485-6

Autophagy is relevant for diverse processes in eukaryotic cells, making its regulation of fundamental importance. The formation and maturation of autophagosomes require a complex choreography of numerous factors. The endosomal sorting complex required for transport (ESCRT) is implicated in the final step of autophagosomal maturation by sealing of the phagophore membrane. ESCRT-III components were shown to mediate membrane scission by forming filaments that interact with cellular membranes. However, the molecular mechanisms underlying the recruitment of ESCRTs to non-endosomal membranes remain largely unknown. Here we focus on the ESCRT-associated protein ALG2-interacting protein X (ALIX) and identify Ca 2+ -dependent lipid binding protein 1 (CaLB1) as its interactor. Our findings demonstrate that CaLB1 interacts with AUTOPHAGY8 (ATG8) and PI(3)P, a phospholipid found in autophagosomal membranes. Moreover, CaLB1 and ALIX localize with ATG8 on autophagosomes upon salt treatment and assemble together into condensates. The depletion of CaLB1 impacts the maturation of salt-induced autophagosomes and leads to reduced delivery of autophagosomes to the vacuole. Here, we propose a crucial role of CaLB1 in augmenting phase separation of ALIX, facilitating the recruitment of ESCRT-III to the site of phagophore closure thereby ensuring efficient maturation of autophagosomes. The study demonstrates that the C2 domain-containing protein CaLB1 interacts with and induces phase separation of the ESCRT protein ALIX on autophagosomes. This modulates autophagosome maturation, particularly in response to salt stress, in Arabidopsis .