SidD, a novel deAMPlyase from L. pneumophila

Wednesday, October 26, 2011 — Poster Session IV
2:00 p.m. – 4:00 p.m.	Natcher Conference Center	NICHD	VIROL/MICRO-5

* FARE Award Winner

Authors

• Y Chen
• M Neunuebel
• M Machner

Abstract

Legionella pneumophila is a Gram-negative bacterium that can cause Legionnaires' disease, a severe pneumonia in humans. Following infection, the bacterium interferes with host cell early secretory route by exploiting the activity of the small GTPase Rab1, the regulator of the host cell secretory pathway. The bacterial protein SidM modifies Rab1 through AMPylation, the covalent addition of adenosine monophosphate (AMP). Overproduction of SidM in tissue culture cells causes cytotoxicity due to SidM’s AMPylation activity. Here we characterized effector protein SidD from L. pneumophila that catalyzed AMP removal from Rab1, a process named DeAMPylation. Using SidD variants in invitro assay we mapped the activity region to the central part of SidD. Fluorescently labeled SidD showed colocalization with the Golgi when overproduced in COS1 cells, consistent with the primary localization of Rab1. All fragments containing the C-terminus of SidD localized to the Golgi, indicating a role of the C-terminus for SidD localization. Cytotoxicity studies revealed that simultaneous production of SidD led to a significant reduction of cytotoxicity in cells producing SidM. Moreover, both the de-AMPylation activity and the C-terminus localization signal are required to rescue the cytotoxicity. Our studies provided new insights on how L. pneumophila effectors modulate the host cell GTPase Rab1.

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