title: RalF-Mediated Activation of Arf6 Controls Rickettsia typhi Invasion by Co-Opting Phosphoinositol Metabolism.
abstract: Rickettsiae are obligate intracellular pathogens that induce their uptake into nonphagocytic cells; however, the events instigating this process are incompletely understood. Importantly, diverse Rickettsia species are predicted to utilize divergent mechanisms to colonize host cells, as nearly all adhesins and effectors involved in host cell entry are differentially encoded in diverse Rickettsia species. One particular effector, RalF, a Sec7 domain-containing protein that functions as a guanine nucleotide exchange factor of ADP-ribosylation factors (Arfs), is critical for Rickettsia typhi (typhus group rickettsiae) entry but pseudogenized or absent from spotted fever group rickettsiae. Secreted early during R. typhi infection, RalF localizes to the host plasma membrane and interacts with host ADP-ribosylation factor 6 (Arf6). Herein, we demonstrate that RalF activates Arf6, a process reliant on a conserved Glu within the RalF Sec7 domain. Furthermore, Arf6 is activated early during infection, with GTP-bound Arf6 localized to the R. typhi entry foci. The regulation of phosphatidylinositol 4-phosphate 5-kinase (PIP5K), which generates PI(4,5)P2, by activated Arf6 is instrumental for bacterial entry, corresponding to the requirement of PI(4,5)P2for R. typhi entry. PI(3,4,5)P3is then synthesized at the entry foci, followed by the accumulation of PI(3)P on the short-lived vacuole. Inhibition of phosphoinositide 3-kinases, responsible for the synthesis of PI(3,4,5)P3and PI(3)P, negatively affects R. typhi infection. Collectively, these results identify RalF as the first bacterial effector to directly activate Arf6, a process that initiates alterations in phosphoinositol metabolism critical for a lineage-specific Rickettsia entry mechanism.
authors.affiliation: Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, Maryland, USA.
authors.forename: Kristen E
authors.initials: KE
authors.lastname: Rennoll-Bankert
authors.affiliation: Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, Maryland, USA.
authors.forename: M Sayeedur
authors.initials: MS
authors.lastname: Rahman
authors.affiliation: Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, Maryland, USA.
authors.forename: Mark L
authors.initials: ML
authors.lastname: Guillotte
authors.affiliation: Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, Maryland, USA.
authors.forename: Stephanie S
authors.initials: SS
authors.lastname: Lehman
authors.affiliation: Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, Maryland, USA.
authors.forename: Magda
authors.initials: M
authors.lastname: Beier-Sexton
authors.affiliation: Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, Maryland, USA.
authors.forename: Joseph J
authors.initials: JJ
authors.lastname: Gillespie
authors.affiliation: Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore, Maryland, USA aazad@som.umaryland.edu.
authors.forename: Abdu F
authors.initials: AF
authors.lastname: Azad
mesh_terms: D000089982:ADP-Ribosylation Factor 6; D020727:ADP-Ribosylation Factors / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D002522:Chlorocebus aethiops; D005786:Gene Expression Regulation / Q000502:physiology; D006160:Guanosine Triphosphate; D006367:HeLa Cells; D006801:Humans; D010716:Phosphatidylinositols / Q000378:metabolism*; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics / Q000378:metabolism; D010957:Plasmids; D011485:Protein Binding; D012286:Rickettsia typhi / Q000235:genetics / Q000378:metabolism*; D014709:Vero Cells
chemical_list: D000089982:ADP-Ribosylation Factor 6; D001426:Bacterial Proteins; D010716:Phosphatidylinositols; D006160:Guanosine Triphosphate; D017853:Phosphotransferases (Alcohol Group Acceptor); D020727:ADP-Ribosylation Factors; C000715348:ARF6 protein, human