The Legionella pneumophila effector protein DrrA is a Rab1 guanine nucleotide-exchange factor

Abstract

The intracellular pathogen Legionella pneumophila avoids fusion with lysosomes and subverts membrane transport from the endoplasmic reticulum to create an organelle that supports bacterial replication. Transport of endoplasmic reticulum-derived vesicles to the Legionella-containing vacuole (LCV) requires bacterial proteins that are translocated into host cells by a type IV secretion apparatus called Dot/Icm. Recent observations have revealed recruitment of the host GTPase Rab1 to the LCV by a process requiring the Dot/Icm system. Here, a visual screen was used to identify L. pneumophila mutants with defects in Rab1 recruitment. One of the factors identified in this screen was DrrA, a new Dot/Icm substrate protein translocated into host cells. We show that DrrA is a potent and highly specific Rab1 guanine nucleotide-exchange factor (GEF). DrrA can disrupt Rab1-mediated secretory transport to the Golgi apparatus by competing with endogenous exchange factors to recruit and activate Rab1 on plasma membrane-derived organelles. These data establish that intracellular pathogens have the capacity to directly modulate the activation state of a specific member of the Rab family of GTPases and thus further our understanding of the mechanisms used by bacterial pathogens to manipulate host vesicular transport.

The Response Regulator CpxR Directly Regulates Expression of SeveralLegionella pneumophila icm/dotComponents as Well as New Translocated Substrates

“…A second explanation might be related to the very large number of different protozoans that were shown to serve as hosts for L. pneumophila. It might be that replication in different hosts requires different sets of IDTS (overlapping or nonoverlapping), and it is possible that the set of IDTS which is regulated by CpxR is less critical for intracellular growth in the two hosts examined; however, the cpxR mutant might have Even though no intracellular growth defect was observed for the cpxR deletion mutant, the CpxR regulator was found to affect several pathogenesis-related phenotypes: (i) the CpxR regulator was found in a screen aimed at identification of suppressors of the IcmO/DotL lethality phenotype (57); (ii) the CpxR regulator was identified in a screen that was conducted to identify L. pneumophila mutants with defects in Rab1 recruitment (37); and (iii) the cpxR deletion mutation was found to strongly affect IDTS translocation (this study). All these results indicate that CpxR is a central regulator of L. pneumophila pathogenesis.…”

“…1A), including (i) the icmV and icmW genes, both of which are part of an operon along with one additional gene (icmV-dotA and icmW-icmX) and which were found to contain the CpxR regulatory element on the complementary strand (the CpxR regulatory element that was characterized previously in the icmR regulatory region was located on the direct strand [ Fig. 1A]); (ii) the lvgA gene, whose product was shown previously to interact with the IcmS protein (58) and which was also found to contain the CpxR regulatory element on the complementary strand; and (iii) the sidH and sidM/drrA genes, whose products were shown previously to be translocated into host cells in an Icm/Dot-dependent manner (31,37,43) and which were found to contain the CpxR regulatory element on the direct strand.…”

“…The icm/dot type IV secretion system is the major virulence system known in L. pneumophila; this system consists of 26 Icm/Dot proteins that probably constitute the secretion complex itself, as well as accessory proteins, such as chaperones (for a review, see reference 49). In the last 5 years, a growing number of protein substrates (RalF, LidA, Lep, Sid, Vip, Wip, Ylf, Leg, Vpd, Drr, and Ceg) that are translocated into host cells via the icm/dot secretion system were identified (for a review, see reference 42); in most cases the function of these proteins is not known, but two of them (SidF and SdhA) were shown to be required for inhibition of host cell death (5,27) and four others (RalF, LidA, DrrA/SidM, and SidJ) were shown to be required for manipulation of host cell vesicular trafficking (28,31,37,38), as well as other functions (11). Even though many icm/dot genes and many more icm/dot translocated substrates (IDTS), as well as potential IDTS, have been identified, there is only a limited amount of information about direct regulators that control the levels of expression of these genes.…”

“…In addition, after the discovery that CpxR is a direct regulator of icmR (18), this protein was identified in two additional screens related to pathogenesis. The CpxR regulator was found in a screen aimed at identification of suppressors of the IcmO/DotL lethality phenotype (57), as well as in a screen that was conducted to identify L. pneumophila mutants with defects in Rab1 recruitment (37). However, no additional genes directly regulated by CpxR were identified in these two screens.…”

Legionella pneumophila inhibits macrophage apoptosis by targeting pro-death members of the Bcl2 protein family

“…By intercepting vesicles originating from the endoplasmic reticulum (ER), the L. pneumophila-containing vacuole eventually is transformed into a compartment characteristic of rough ER (5). Consistent with this notion, L. pneumophila modulates the activities of vesicle trafficking regulatory molecules such as small GTPases Arf1 and Rab1 by translocated guanine nucleotide exchange factors specific for these proteins, thus facilitating the acquisition of ER materials and the subsequent intracellular bacterial multiplication (6)(7)(8).…”