Legionella pneumophila LidA Affects Nucleotide Binding and Activity of the Host GTPase Rab1

Abstract

bLegionella pneumophila, the causative agent of a severe pneumonia known as Legionnaires' disease, intercepts material from host cell membrane transport pathways to create a specialized vacuolar compartment that supports bacterial replication. Delivery of bacterial effector proteins into the host cell requires the Dot/Icm type IV secretion system. Several effectors, including SidM, SidD, and LepB, were shown to target the early secretory pathway by manipulating the activity of the host GTPase Rab1. While the function of these effectors has been well characterized, the role of another Rab1-interacting protein from L. pneumophila, the effector protein LidA, is poorly understood. Here, we show that LidA binding to Rab1 stabilized the Rab1-guanosine nucleotide complex, protecting it from inactivation by GTPase-activating proteins (GAPs) and from nucleotide extraction. The protective effect of LidA on the Rab1-guanine nucleotide complex was concentration dependent, consistent with a 1:1 stoichiometry of the LidA-Rab1 complex. The central coiled-coil region of LidA was sufficient for Rab1 binding and to prevent GAPmediated inactivation or nucleotide extraction from Rab1. In addition, the central region mediated binding to phosphatidylinositol 3-phosphate and other phosphoinositides. When bound to Rab1, LidA interfered with the covalent modification of Rab1 by phosphocholination or AMPylation, and it also blocked de-AMPylation of Rab1 by SidD and dephosphocholination by Lem3. Based on these findings, we propose a role for LidA in bridging the membrane of the Legionella-containing vacuole (LCV) with that of secretory transport vesicles surrounding the LCV.

Type II Secretion Is Necessary for Optimal Association of the Legionella-Containing Vacuole with Macrophage Rab1B but Enhances Intracellular Replication Mainly by Rab1B-Independent Mechanisms

“…S7). We surmise that in the case of the WT, other host GTPases and their associated vesicles compensate for the lack of Rab1B recruitment, as previously postulated (112). Since the growth of the sidM mutant was also unaffected by depletion of Rab1B in U937 cells (Fig.…”

“…Since the growth of the sidM mutant was also unaffected by depletion of Rab1B in U937 cells (Fig. 7B), we inferred that the sidM mutant can still recruit other vesicles from the human host cell in order to support its growth (112). Most importantly, in contrast to the WT and the sidM mutant, the lspF mutant showed a 21% reduction in growth in Rab1B-depleted U937 cells compared to scramble U937 cells ( Fig.…”

Creating a customized intracellular niche: subversion of host cell signaling byLegionellatype IV secretion system effectors

“…Moreover, during infection, the spatio-temporal control of translocation and localization of SidM and AnkX might predominantly determine which modification occurs at a given time. Interestingly, LidA-bound Rab1 was found to be less prone to modification by both SidM and AnkX, suggesting that other effectors also indirectly impact on the intricate regulation of the modification status of Rab1 (Neunuebel et al 2012). …”

LtpD Is a Novel Legionella pneumophila Effector That Binds Phosphatidylinositol 3-Phosphate and Inositol Monophosphatase IMPA1

“…L. pneumophila also recruits the inositol polyphosphate-5-phosphatase OCRL1 to the LCV, which dephosphorylates PtdIns(4,5)P 2 , yielding PtdIns(4)P (18). The accumulation of PtdIns(4)P on the LCV serves as a membrane anchor for SidC and SidM (19,20), whereas LidA, SetA, and LpnE are localized to the LCV membrane and bind PtdIns(3)P in vitro (18,21,22), suggesting that the LCV membrane also contains PtdIns(3)P. The aim of the present study was to investigate the effector LtpD (23), which we found to bind both PtdIns(3)P and IMPA1.…”