ABSTRACT
Legionella pneumophila actively modulates host vesicle trafficking pathways to facilitate its intracellular replication with effectors translocated by the Dot/Icm type IV secretion system (T4SS). The SidM/DrrA protein functions by locking the small GTPase Rab1 into an active form by its guanine nucleotide exchange factor (GEF) and AMPylation activity. Here we demonstrate that the L. pneumophila protein SidD preferably deAMPylates Rab1. We found that the deAMPylation activity of SidD could suppress the toxicity of SidM to yeast and is required to release Rab1 from bacterial phagosomes efficiently. A molecular mechanism for the temporal control of Rab1 activity in different phases of L. pneumophila infection is thus established. These observations indicate that AMPylation-mediated signal transduction is a reversible process regulated by specific enzymes.
Subject(s)
Bacterial Proteins/metabolism , Legionella pneumophila/enzymology , Legionnaires' Disease/enzymology , rab1 GTP-Binding Proteins/metabolism , Animals , Aspartic Acid/chemistry , Bacterial Proteins/chemistry , Bacterial Proteins/genetics , Bacterial Proteins/toxicity , Cells, Cultured , Female , Guanine Nucleotide Exchange Factors/genetics , Guanine Nucleotide Exchange Factors/metabolism , Guanine Nucleotide Exchange Factors/toxicity , Legionnaires' Disease/physiopathology , Macrophages/enzymology , Macrophages/pathology , Mice , Phenotype , Plasmids/genetics , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae/growth & development , Saccharomyces cerevisiae/metabolism , Time Factors , Transformation, GeneticABSTRACT
In the course of Legionnaires' disease, the bacterium Legionella pneumophila affects the intracellular vesicular trafficking of infected eukaryotic cells by recruiting the small guanosine triphosphatase (GTPase) Rab1 to the cytosolic face of the Legionella-containing vacuole. In order to accomplish this, the Legionella protein DrrA contains a specific guanine nucleotide exchange activity for Rab1 activation that exchanges guanosine triphosphate (GTP) for guanosine diphosphate on Rab1. We found that the amino-terminal domain of DrrA possesses adenosine monophosphorylation (AMPylation) activity toward the switch II region of Rab1b, leading to posttranslational covalent modification of tyrosine 77. AMPylation of switch II by DrrA restricts the access of GTPase activating proteins, thereby rendering Rab1b constitutively active.
