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Structural determinants in GABARAP required for the selective binding and recruitment of ALFY to LC3B-positive structures.

ABSTRACT: Several autophagy proteins contain an LC3-interacting region (LIR) responsible for their interaction with Atg8 homolog proteins. Here, we show that ALFY binds selectively to LC3C and the GABARAPs through a LIR in its WD40 domain. Binding of ALFY to GABARAP is indispensable for its recruitment to LC3B-positive structures and, thus, for the clearance of certain p62 structures by autophagy. In addition, the crystal structure of the GABARAP-ALFY-LIR peptide complex identifies three conserved residues in the GABARAPs that are responsible for binding to ALFY. Interestingly, introduction of these residues in LC3B is sufficient to enable its interaction with ALFY, indicating that residues outside the LIR-binding hydrophobic pockets confer specificity to the interactions with Atg8 homolog proteins.

SUBMITTER: Lystad AH

PROVIDER: S-EPMC4210083 | biostudies-literature | 2014 May

REPOSITORIES: biostudies-literature

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Structural determinants in GABARAP required for the selective binding and recruitment of ALFY to LC3B-positive structures.

Lystad Alf Håkon AH Ichimura Yoshinobu Y Takagi Kenji K Yang Yinjie Y Pankiv Serhiy S Kanegae Yumi Y Kageyama Shun S Suzuki Mariko M Saito Izumu I Mizushima Tsunehiro T Komatsu Masaaki M Simonsen Anne A

EMBO reports 20140325 5

Several autophagy proteins contain an LC3-interacting region (LIR) responsible for their interaction with Atg8 homolog proteins. Here, we show that ALFY binds selectively to LC3C and the GABARAPs through a LIR in its WD40 domain. Binding of ALFY to GABARAP is indispensable for its recruitment to LC3B-positive structures and, thus, for the clearance of certain p62 structures by autophagy. In addition, the crystal structure of the GABARAP-ALFY-LIR peptide complex identifies three conserved residue ...[more]

PMID: 24668264

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Project description:N-Methyl-d-aspartate receptors (NMDARs) are ionotropic ligand-gated glutamate receptors that mediate fast excitatory synaptic transmission in the central nervous system (CNS). Several neurological disorders may involve NMDAR hypofunction, which has driven therapeutic interest in positive allosteric modulators (PAMs) of NMDAR function. Here we describe modest changes to the tetrahydroisoquinoline scaffold of GluN2C/GluN2D-selective PAMs that expands activity to include GluN2A- and GluN2B-containing recombinant and synaptic NMDARs. These new analogues are distinct from GluN2C/GluN2D-selective compounds like (+)-(3-chlorophenyl)(6,7-dimethoxy-1-((4-methoxyphenoxy)methyl)-3,4-dihydroisoquinolin-2(1H)-yl)methanone (CIQ) by virtue of their subunit selectivity, molecular determinants of action, and allosteric regulation of agonist potency. The (S)-enantiomers of two analogues (EU1180-55, EU1180-154) showed activity at NMDARs containing all subunits (GluN2A, GluN2B, GluN2C, GluN2D), whereas the (R)-enantiomers were primarily active at GluN2C- and GluN2D-containing NMDARs. Determination of the actions of enantiomers on triheteromeric receptors confirms their unique pharmacology, with greater activity of (S) enantiomers at GluN2A/GluN2D and GluN2B/GluN2D subunit combinations than (R) enantiomers. Evaluation of the (S)-EU1180-55 and EU1180-154 response of chimeric kainate/NMDA receptors revealed structural determinants of action within the pore-forming region and associated linkers. Scanning mutagenesis identified structural determinants within the GluN1 pre-M1 and M1 regions that alter the activity of (S)-EU1180-55 but not (R)-EU1180-55. By contrast, mutations in pre-M1 and M1 regions of GluN2D perturb the actions of only the (R)-EU1180-55 but not the (S) enantiomer. Molecular modeling supports the idea that the (S) and (R) enantiomers interact distinctly with GluN1 and GluN2 pre-M1 regions, suggesting that two distinct sites exist for these NMDAR PAMs, each of which has different functional effects.

Dataset Information

Structural determinants in GABARAP required for the selective binding and recruitment of ALFY to LC3B-positive structures.

Publications

Structural determinants in GABARAP required for the selective binding and recruitment of ALFY to LC3B-positive structures.

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