De-AMPylation of the Small GTPase Rab1 by the Pathogen Legionella pneumophila

Abstract

The bacterial pathogen Legionella pneumophila exploits host cell vesicle transport by transiently manipulating the activity of the small guanosine triphosphatase (GTPase) Rab1. The effector protein SidM recruits Rab1 to the Legionella-containing vacuole (LCV), where it activates Rab1 and then AMPylates it by covalently adding adenosine monophosphate (AMP). L. pneumophila GTPase-activating protein LepB inactivates Rab1 before its removal from LCVs. Because LepB cannot bind AMPylated Rab1, the molecular events leading to Rab1 inactivation are unknown. We found that the effector protein SidD from L. pneumophila catalyzed AMP release from Rab1, generating de-AMPylated Rab1 accessible for inactivation by LepB. L. pneumophila mutants lacking SidD were defective for Rab1 removal from LCVs, identifying SidD as the missing link connecting the processes of early Rab1 accumulation and subsequent Rab1 removal during infection.

Characterization of Enzymes from Legionella pneumophila Involved in Reversible Adenylylation of Rab1 Protein

“…S4), confirming the presence of the previously identified deadenylylase SidD (21,22) and indicating the necessity of this specific enzyme for removal of the covalent modification. To obtain insights into the competition of adenylylation by DrrA and deadenylylation by SidD, we quantified the catalytic activity of recombinantly produced and purified SidD.…”

“…The necessity of a protein catalyzing the demodification of Rab1 has been proposed after identification of the adenylyltransferase domain of DrrA (18). Confirming this hypothesis, recently a protein named SidD from Legionella was demonstrated to catalyze the hydrolysis of AMP-Rab1 (21,22).…”

“…Very recently, it has also been demonstrated for DrrA-catalyzed adenylylation that the modification can be reversed by the Legionella protein SidD (21,22), similarly to the antagonistic proteins AnkX and Lem3 (35,51).…”

Posttranslational modifications of Rab GTPases help their insertion into membranes

“…The active GTP-bound form of Rab1 is a preferable substrate for DrrA/SidM-catalyzed adenylylation, suggesting that Rab1 activation is coupled to its modification. The recently reported identification of SidD with deadenylylation activity toward Rab1-AMP (13) confirms that AMPylation is a regulated reversible modification like many other posttranslational modifications. The other Legionella effector studied by Oesterlin et al (2) is AnkX/ Lpg0695.…”

A New Chemical Handle for Protein AMPylation at the Host–Pathogen Interface

“…In 2011, the first de-AMPylating enzyme, or 'protein AMPylase', was discovered in L. pneumophila, confirming the importance and the dynamic nature of posttranslational AMPylation. [13,14] Apart from its role in pathogenicity, there is growing evidence that AMPylation may operate as a general intracellular signaling mechanism in normal cell function. Bacterial Fido domain proteins are involved in cell division, whilst HYPE, a human Fido motif protein, may regulate interactions of small GTPases with their binding partners.…”