Rab1 Guanine Nucleotide Exchange Factor SidM Is a Major Phosphatidylinositol 4-Phosphate-binding Effector Protein of Legionella pneumophila

被引:197
作者
Brombacher, Eva [1 ]
Urwyler, Simon [1 ]
Ragaz, Curdin [1 ]
Weber, Stefan S. [1 ]
Kami, Keiichiro [2 ]
Overduin, Michael [2 ]
Hilbi, Hubert [1 ]
机构
[1] ETH, Inst Microbiol, CH-8093 Zurich, Switzerland
[2] Univ Birmingham, Canc Res UK Inst Canc Studies, Birmingham B15 2TT, W Midlands, England
基金
瑞士国家科学基金会;
关键词
CIRCULAR-DICHROISM SPECTRA; TRANSLOCATED SUBSTRATE; LEGIONNAIRES-DISEASE; ENDOPLASMIC-RETICULUM; HOMOLOGY-DOMAIN; HUMAN-MONOCYTES; EXIT SITES; BINDING; SYSTEM; GENES;
D O I
10.1074/jbc.M807505200
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The causative agent o f Legionnaires disease, Legionella pneumophila, forms a replicative vacuole in phagocytes by means of the intracellular multiplication/defective organelle trafficking (Icm/Dot) type IV secretion system and translocated effector proteins, some of which subvert host GTP and phosphoinositide (PI) metabolism. The Icm/Dot substrate SidC anchors to the membrane of Legionella-containing vacuoles (LCVs) by specifically binding to phosphatidylinositol 4-phosphate (PtdIns(4) P). Using a nonbiased screen for novel L. pneumophila PI-binding proteins, we identified the Rab1 guanine nucleotide exchange factor (GEF) SidM/DrrA as the predominant PtdIns(4) P-binding protein. Purified SidM specifically and directly bound to PtdIns(4) P, whereas the SidM-interacting Icm/Dot substrate LidA preferentially bound PtdIns(3) P but also PtdIns(4) P, and the L. pneumophila Arf1 GEF RalF did not bind to any PIs. The PtdIns(4) P-binding domain of SidM was mapped to the 12-kDa C-terminal sequence, termed "P4M" (PtdIns4P binding of SidM/DrrA). The isolated P4M domain is largely helical and displayed higher PtdIns(4) P binding activity in the context of the alpha-helical, monomeric full-length protein. SidM constructs containing P4M were translocated by Icm/Dot-proficient L. pneumophila and localized to the LCV membrane, indicating that SidM anchors to PtdIns(4) P on LCVs via its P4M domain. An L. pneumophila Delta sidM mutant strain displayed significantly higher amounts of SidC on LCVs, suggesting that SidM and SidC compete for limiting amounts of PtdIns(4) P on the vacuole. Finally, RNA interference revealed that PtdIns(4) P on LCVs is specifically formed by host PtdIns 4-kinase III beta. Thus, L. pneumophila exploits PtdIns(4) P produced by PtdIns 4-kinase III beta to anchor the effectors SidC and SidM to LCVs.
引用
收藏
页码:4846 / 4856
页数:11
相关论文
共 62 条
[41]   A C-terminal translocation signal required for Dot/lcm-dependent delivery of the Legionella RalF protein to host cells [J].
Nagai, H ;
Cambronne, ED ;
Kagan, JC ;
Amor, JC ;
Kahn, RA ;
Roy, CR .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2005, 102 (03) :826-831
[42]   A bacterial guanine nucleotide exchange factor activates ARF on Legionella phagosomes [J].
Nagai, H ;
Kagan, JC ;
Zhu, XJ ;
Kahn, RA ;
Roy, CR .
SCIENCE, 2002, 295 (5555) :679-682
[43]   Sell repeat protein LpnE is a Legionella pneumophila virulence determinant that influences vacuolar trafficking [J].
Newton, Hayley J. ;
Sansom, Fiona M. ;
Dao, Jenny ;
McAlister, Adrian D. ;
Sloan, Joan ;
Cianciotto, Nicholas P. ;
Hartland, Elizabeth L. .
INFECTION AND IMMUNITY, 2007, 75 (12) :5575-5585
[44]   Identification of Legionella pneumophila-specific genes by genomic subtractive hybridization with Legionella micdadei and identification of lpnE, a gene required for efficient host cell entry [J].
Newton, HJ ;
Sansom, FM ;
Bennett-Wood, V ;
Hartland, EL .
INFECTION AND IMMUNITY, 2006, 74 (03) :1683-1691
[45]   The Legionella IcmS-IcmW protein complex is important for Dot/Icm-mediated protein translocation [J].
Ninio, S ;
Zuckman-Cholon, DM ;
Cambronne, ED ;
Roy, CR .
MOLECULAR MICROBIOLOGY, 2005, 55 (03) :912-926
[46]   Effector proteins translocated by Legionella pneumophila:: strength in numbers [J].
Ninio, Shira ;
Roy, Craig R. .
TRENDS IN MICROBIOLOGY, 2007, 15 (08) :372-380
[47]   RAB-GDP DISSOCIATION INHIBITOR - PUTTING RAB-GTPASES IN THE RIGHT PLACE [J].
PFEFFER, SR ;
DIRACSVEJSTRUP, AB ;
SOLDATI, T .
JOURNAL OF BIOLOGICAL CHEMISTRY, 1995, 270 (29) :17057-17059
[48]   The Legionella pneumophila phosphatidylinositol-4 phosphate-binding type IV substrate SidC recruits endoplasmic reticulum vesicles to a replication-permissive vacuole [J].
Ragaz, Curdin ;
Pietsch, Holger ;
Urwyler, Simon ;
Tiaden, Andre ;
Weber, Stefan S. ;
Hilbi, Hubert .
CELLULAR MICROBIOLOGY, 2008, 10 (12) :2416-2433
[49]   Attachment and fusion of endoplasmic reticulum with vacuoles containing Legionella pneumophila [J].
Robinson, CG ;
Roy, CR .
CELLULAR MICROBIOLOGY, 2006, 8 (05) :793-805
[50]   Host cell killing and bacterial conjugation require overlapping sets of genes within a 22-kb region of the Legionella pneumophila genome [J].
Segal, G ;
Purcell, M ;
Shuman, HA .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 1998, 95 (04) :1669-1674