Structural insights into the recognition of phosphorylated FUNDC1 by LC3B in mitophagy

Abstract

Mitophagy is an essential intracellular process that eliminates dysfunctional mitochondria and maintains cellular homeostasis. Mitophagy is regulated by the post-translational modification of mitophagy receptors. Fun14 domain-containing protein 1 (FUNDC1) was reported to be a new receptor for hypoxia-induced mitophagy in mammalian cells and interact with microtubule-associated protein light chain 3 beta (LC3B) through its LC3 interaction region (LIR). Moreover, the phosphorylation modification of FUNDC1 affects its binding affinity for LC3B and regulates selective mitophagy. However, the structural basis of this regulation mechanism remains unclear. Here, we present the crystal structure of LC3B in complex with a FUNDC1 LIR peptide phosphorylated at Ser17 (pS17), demonstrating the key residues of LC3B for the specific recognition of the phosphorylated or dephosphorylated FUNDC1. Intriguingly, the side chain of LC3B Lys49 shifts remarkably and forms a hydrogen bond and electrostatic interaction with the phosphate group of FUNDC1 pS17. Alternatively, phosphorylated Tyr18 (pY18) and Ser13 (pS13) in FUNDC1 significantly obstruct their interaction with the hydrophobic pocket and Arg10 of LC3B, respectively. Structural observations are further validated by mutation and isothermal titration calorimetry (ITC) assays. Therefore, our structural and biochemical results reveal a working model for the specific recognition of FUNDC1 by LC3B and imply that the reversible phosphorylation modification of mitophagy receptors may be a switch for selective mitophagy.Electronic supplementary materialThe online version of this article (doi:10.1007/s13238-016-0328-8) contains supplementary material, which is available to authorized users.

Silencing FUNDC1 alleviates chronic obstructive pulmonary disease by inhibiting mitochondrial autophagy and bronchial epithelium cell apoptosis under hypoxic environment

“…23,24 FUNDC1 shows significant impacts on the mitophagy induced by hypoxia in mammalian cells. 25 Another study has also indicated that accumulation of FUNDC1 is presented at the MAM in the hypoxia environment. 26 Furthermore, FUNDC1 was also verified to upregulate in COPD patients, 14 which was consistent with the findings in this study.…”

The conformational and mutational landscape of the ubiquitin-like marker for the autophagosome formation in cancer

“…Y113C was recently shown to inhibit the enzymatic activity of Atg7 (E1-like enzyme) but not the E2-like activity (Nuta et al, 2018). Mutations at K49 alter the binding to the phosphorylated variants of the LIR-containing LC3B interactor, FUNDC1 (Lv et al, 2016), whereas if this residue is mutated to alanine can increase the binding of another LIR-containing protein, i.e. Nix (Rogov et al, 2017).…”

“…Several three-dimensional (3D) structures of LC3B have been solved by X-ray or NMR in the free state (Kouno et al, 2005;Rogov et al, 2013) or in complex with different biological partners (Ichimura et al, 2008;Jemal et al, 2011;Kuang et al, 2016b;Kwon et al, 2017;Lv et al, 2016;McEwan et al, 2015;Olsvik et al, 2015;Qiu et al, 2017;Rogov et al, 2017Rogov et al, , 2013Stadel et al, 2015;Suzuki et al, 2014;Yang et al, 2017). This provides a valuable source of information for investigations using biomolecular simulations and other structural computational methods of both the LC3B wild-type and its mutated variant.…”

How to get rid of mitochondria: crosstalk and regulation of multiple mitophagy pathways

“…ULK1, as the other major regulator, was shown to be recruited to fragmented mitochondria under hypoxia and to phosphorylate Ser17 of FUNDC1, enhancing the binding to LC3 (Wu et al, 2014b). Although these phosphorylation sites are just a few residues apart, their impact on the interaction of the protein to LC3 is remarkable and the structural basis of this interaction was intensively studied (Lv et al, 2017).…”