title: Anaplasma phagocytophilum MSP2(P44)-18 predominates and is modified into multiple isoforms in human myeloid cells.
abstract: Anaplasma phagocytophilum is the etiologic agent of human granulocytic anaplasmosis. MSP2(P44), the bacterium's major surface protein, is encoded by a paralogous gene family and has been implicated in a variety of pathobiological processes, including antigenic variation, host adaptation, adhesion, porin activity, and structural integrity. The consensus among several studies performed at the DNA and RNA levels is that a heterogeneous mix of a limited number of msp2(p44) transcripts is expressed by A. phagocytophilum during in vitro cultivation. Such analyses have yet to be extended to the protein level. In this study, we used proteomic and molecular approaches to determine that MSP2(P44)-18 is the predominant if not the only paralog expressed and is modified into multiple 42- to 44-kDa isoforms by A. phagocytophilum strain HGE1 during infection of HL-60 cells. The msp2(p44) expression profile was homogeneous for msp2(p44)-18. Thus, MSP2(P44)-18 may have a fitness advantage in HL-60 cell culture in the absence of selective immune pressure. Several novel 22- to 27-kDa MSP2 isoforms lacking most of the N-terminal conserved region were also identified. A. phagocytophilum MSP2(P44) orthologs expressed by other pathogens in the family Anaplasmataceae are glycosylated. Gas chromatography revealed that recombinant MSP2(P44)-18 is modified by glucose, galactose, xylose, mannose, and trace amounts of other glycosyl residues. These data are the first to confirm differential modification of any A. phagocytophilum MSP2(P44) paralog and the first to provide evidence for expression of truncated versions of such proteins.
authors.affiliation: Department of Microbiology, Immunology, and Molecular Genetics, University of Kentucky College of Medicine, Lexington, Kentucky 40504, USA.
authors.forename: Madhubanti
authors.initials: M
authors.lastname: Sarkar
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Troese
authors.forename: Sarah A
authors.initials: SA
authors.lastname: Kearns
authors.forename: Tian
authors.initials: T
authors.lastname: Yang
authors.forename: Dexter V
authors.initials: DV
authors.lastname: Reneer
authors.forename: Jason A
authors.initials: JA
authors.lastname: Carlyon
mesh_terms: D000595:Amino Acid Sequence; D041081:Anaplasma phagocytophilum / Q000378:metabolism*; D001425:Bacterial Outer Membrane Proteins / Q000096:biosynthesis* / Q000737:chemistry; D015153:Blotting, Western; D002241:Carbohydrates / Q000032:analysis; D002460:Cell Line; D002849:Chromatography, Gas; D015180:Electrophoresis, Gel, Two-Dimensional; D006031:Glycosylation; D006801:Humans; D008969:Molecular Sequence Data; D022423:Myeloid Cells / Q000382:microbiology*; D020033:Protein Isoforms / Q000096:biosynthesis / Q000737:chemistry
chemical_list: D001425:Bacterial Outer Membrane Proteins; D002241:Carbohydrates; D020033:Protein Isoforms; C488283:msp2 protein, Anaplasma phagocytophilum