Multiple host proteins that function in phosphatidylinositol-4-phosphate metabolism are recruited to the chlamydial inclusion.
Chlamydiae replicate within a nonacidified vacuole, termed an inclusion. As obligate intracellular bacteria, chlamydiae actively modify their vacuole to exploit host signaling and trafficking pathways. Recently, we demonstrated that several Rab GTPases are actively targeted to the inclusion. To define the biological roles of inclusion localized Rab GTPases, we have begun to identify inclusion-localized Rab effectors. Here we demonstrate that oculocerebrorenal syndrome of Lowe protein 1 (OCRL1), a Golgi complex-localized phosphatidylinositol (PI)-5-phosphatase that binds to multiple Rab GTPases, localizes to chlamydial inclusions. By examining the intracellular localization of green fluorescent protein (GFP) fusion proteins that bind to unique phosphoinositide species, we also demonstrate that phosphatidylinositol-4-phosphate (PI4P), the product of OCRL1, is present at the inclusion membrane. Furthermore, two additional host proteins, Arf1, which together with PI4P mediates the recruitment of PI4P-binding proteins to the Golgi complex, and PI4KII alpha, a major producer of Golgi complex-localized PI4P, also localize to chlamydial inclusions. Depletion of OCRL1, Arf1, or PI4KII alpha by small interfering RNA (siRNA) decreases inclusion formation and the production of infectious progeny. Infectivity is further decreased in cells simultaneously depleted for all three host proteins, suggesting partially overlapping functions in infected cells. Collectively, these data demonstrate that Chlamydia species create a unique replication-competent vacuolar environment by modulating both the Rab GTPase and the PI composition of the chlamydial inclusion.
Department of Microbiology and Immunology, College of Veterinary Medicine, Cornell University, Ithaca, NY 14853, USA.
Andrew M
AM
Moorhead
Joo-Yong
JY
Jung
Asya
A
Smirnov
Susanne
S
Kaufer
Marci A
MA
Scidmore
D020823:ADP-Ribosylation Factor 1 / Q000032:analysis; D001419:Bacteria; D002462:Cell Membrane / Q000737:chemistry*; D002689:Chlamydia / Q000472:pathogenicity*; D006367:HeLa Cells; D006801:Humans; D015778:Minor Histocompatibility Antigens; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism*; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism*; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000032:analysis; D014617:Vacuoles / Q000201:enzymology* / Q000382:microbiology*; D020691:rab GTP-Binding Proteins / Q000378:metabolism*
D015778:Minor Histocompatibility Antigens; D018129:Phosphatidylinositol Phosphates; C037178:phosphatidylinositol 4-phosphate; D017853:Phosphotransferases (Alcohol Group Acceptor); C402182:phosphatidylinositol phosphate 4-kinase; D010744:Phosphoric Monoester Hydrolases; C075695:OCRL protein, human; D020823:ADP-Ribosylation Factor 1; D020691:rab GTP-Binding Proteins