title: Repression of rac2 mRNA expression by Anaplasma phagocytophila is essential to the inhibition of superoxide production and bacterial proliferation.
abstract: Anaplasma phagocytophila, the etiologic agent of human granulocytic ehrlichiosis, is an emerging bacterial pathogen that invades neutrophils and can be cultivated in HL-60 cells. Infected neutrophils and HL-60 cells fail to produce superoxide anion (O(2)(-)), which is partially attributable to the fact that A. phagocytophila inhibits transcription of gp91(phox), an integral component of NADPH oxidase. cDNA microarray and RT-PCR analyses demonstrated that transcription of the gene encoding Rac2, a key component in NADPH oxidase activation, was down-regulated in infected HL-60 cells. Quantitative RT-PCR demonstrated that rac2 mRNA expression was reduced 7-fold in retinoic acid-differentiated HL-60 cells and 50-fold in neutrophils following A. phagocytophila infection. Rac2 protein expression was absent in infected HL-60 cells. Rac1 and Rac2 are interchangeable in their abilities to activate NADPH oxidase. HL-60 cells transfected to express myc-tagged rac1 and gp91(phox) from the CMV immediate early promoter maintained the ability to generate O(2)(-) 120 h postinfection. A. phagocytophila proliferation was severely inhibited in these cells. These results directly attribute the inhibition of rac2 and gp91(phox) transcription to the loss of NADPH oxidase activity in A. phagocytophila-infected cells and demonstrate its importance to bacterial intracellular survival.
authors.affiliation: Section of Rheumatology, Department of Internal Medicine, Yale University School of Medicine, New Haven, CT 06520-8031, USA.
authors.forename: Jason A
authors.initials: JA
authors.lastname: Carlyon
authors.forename: Wai-Tsing
authors.initials: WT
authors.lastname: Chan
authors.forename: Jorge
authors.initials: J
authors.lastname: Galán
authors.forename: Dirk
authors.initials: D
authors.lastname: Roos
authors.forename: Erol
authors.initials: E
authors.lastname: Fikrig
mesh_terms: D041081:Anaplasma phagocytophilum / Q000166:cytology / Q000235:genetics / Q000254:growth & development* / Q000502:physiology*; D002455:Cell Division / Q000235:genetics; D015536:Down-Regulation / Q000502:physiology*; D016873:Ehrlichiosis / Q000378:metabolism / Q000382:microbiology; D020869:Gene Expression Profiling; D018922:HL-60 Cells / Q000187:drug effects / Q000378:metabolism / Q000382:microbiology; D006801:Humans; D008562:Membrane Glycoproteins / Q000096:biosynthesis / Q000235:genetics; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases; D009504:Neutrophils / Q000187:drug effects / Q000378:metabolism / Q000382:microbiology; D012333:RNA, Messenger / Q000032:analysis / Q000037:antagonists & inhibitors* / Q000096:biosynthesis*; D013481:Superoxides / Q000037:antagonists & inhibitors* / Q000378:metabolism; D014162:Transfection; D014212:Tretinoin / Q000494:pharmacology; D020744:rac GTP-Binding Proteins / Q000037:antagonists & inhibitors* / Q000096:biosynthesis* / Q000235:genetics; D020830:rac1 GTP-Binding Protein / Q000235:genetics / Q000378:metabolism; D000097766:RAC2 GTP-Binding Protein
chemical_list: D008562:Membrane Glycoproteins; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases; D012333:RNA, Messenger; D013481:Superoxides; D014212:Tretinoin; D020744:rac GTP-Binding Proteins; D020830:rac1 GTP-Binding Protein; D000097766:RAC2 GTP-Binding Protein; C056558:CYBB protein, human