Dataset Information

Structural insight into the membrane targeting domain of the Legionella deAMPylase SidD.

ABSTRACT: AMPylation, the post-translational modification with adenosine monophosphate (AMP), is catalyzed by effector proteins from a variety of pathogens. Legionella pneumophila is thus far the only known pathogen that, in addition to encoding an AMPylase (SidM/DrrA), also encodes a deAMPylase, called SidD, that reverses SidM-mediated AMPylation of the vesicle transport GTPase Rab1. DeAMPylation is catalyzed by the N-terminal phosphatase-like domain of SidD. Here, we determined the crystal structure of full length SidD including the uncharacterized C-terminal domain (CTD). A flexible loop rich in aromatic residues within the CTD was required to target SidD to model membranes in vitro and to the Golgi apparatus within mammalian cells. Deletion of the loop (?loop) or substitution of its aromatic phenylalanine residues rendered SidD cytosolic, showing that the hydrophobic loop is the primary membrane-targeting determinant of SidD. Notably, deletion of the two terminal alpha helices resulted in a CTD variant incapable of discriminating between membranes of different composition. Moreover, a L. pneumophila strain producing SidD?loop phenocopied a L. pneumophila ?sidD strain during growth in mouse macrophages and displayed prolonged co-localization of AMPylated Rab1 with LCVs, thus revealing that membrane targeting of SidD via its CTD is a critical prerequisite for its ability to catalyze Rab1 deAMPylation during L. pneumophila infection.

SUBMITTER: Tascon I

PROVIDER: S-EPMC7480848 | biostudies-literature | 2020 Aug

REPOSITORIES: biostudies-literature

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Structural insight into the membrane targeting domain of the Legionella deAMPylase SidD.

Tascón Igor I Li Xiao X Lucas María M Nelson D'anna D Vidaurrazaga Ander A Lin Yi-Han YH Rojas Adriana L AL Hierro Aitor A Machner Matthias P MP

PLoS pathogens 20200827 8

AMPylation, the post-translational modification with adenosine monophosphate (AMP), is catalyzed by effector proteins from a variety of pathogens. Legionella pneumophila is thus far the only known pathogen that, in addition to encoding an AMPylase (SidM/DrrA), also encodes a deAMPylase, called SidD, that reverses SidM-mediated AMPylation of the vesicle transport GTPase Rab1. DeAMPylation is catalyzed by the N-terminal phosphatase-like domain of SidD. Here, we determined the crystal structure of ...[more]

PMID: 32853279

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Project description:Cardiac membrane excitability is tightly regulated by an integrated network of membrane-associated ion channels, transporters, receptors, and signaling molecules. Membrane protein dynamics in health and disease are maintained by a complex ensemble of intracellular targeting, scaffolding, recycling, and degradation pathways. Surprisingly, despite decades of research linking dysfunction in membrane protein trafficking with human cardiovascular disease, essentially nothing is known regarding the molecular identity or function of these intracellular targeting pathways in excitable cardiomyocytes.We sought to discover novel pathways for membrane protein targeting in primary cardiomyocytes.We report the initial characterization of a large family of membrane trafficking proteins in human heart. We used a tissue-wide screen for novel ankyrin-associated trafficking proteins and identified 4 members of a unique Eps15 homology (EH) domain-containing protein family (EHD1, EHD2, EHD3, EHD4) that serve critical roles in endosome-based membrane protein targeting in other cell types. We show that EHD1-4 directly associate with ankyrin, provide the first information on the expression and localization of these molecules in primary cardiomyocytes, and demonstrate that EHD1-4 are coexpressed with ankyrin-B in the myocyte perinuclear region. Notably, the expression of multiple EHD proteins is increased in animal models lacking ankyrin-B, and EHD3-deficient cardiomyocytes display aberrant ankyrin-B localization and selective loss of Na/Ca exchanger expression and function. Finally, we report significant modulation of EHD expression following myocardial infarction, suggesting that these proteins may play a key role in regulating membrane excitability in normal and diseased heart.Our findings identify and characterize a new class of cardiac trafficking proteins, define the first group of proteins associated with the ankyrin-based targeting network, and identify potential new targets to modulate membrane excitability in disease. Notably, these data provide the first link between EHD proteins and a human disease model.

Dataset Information

Structural insight into the membrane targeting domain of the Legionella deAMPylase SidD.

Publications

Structural insight into the membrane targeting domain of the Legionella deAMPylase SidD.

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