title: Enhanced intracellular replication of Salmonella enteritidis in HLA-B27-expressing human monocytic cells: dependency on glutamic acid at position 45 in the B pocket of HLA-B27.
abstract: OBJECTIVE
  To reveal the cause of the impaired elimination of Salmonella enteritidis in HLA-B27-transfected human monocytic cells and to study whether the B pocket of HLA-B27 contributes to these modulatory effects.
  
  
  METHODS
  Stable U937 cell transfectants expressing HLA-A2, B27, or different forms of B27 with amino acid substitutions in the B pocket were prepared. Mock-transfected cells were prepared using the antibiotic resistance vector (pSV2neo) alone. Cells were differentiated, infected with S enteritidis, and the number of live intracellular S enteritidis organisms was determined using the colony-forming unit method. To visualize intracellular S enteritidis, the bacteria were transformed with green fluorescent protein (GFP), and studied by confocal microscopy.
  
  
  RESULTS
  Cells expressing wild-type HLA-B27 were more permissive of intracellular replication of S enteritidis compared with mock-transfected or A2-transfected controls. Cells expressing B27 with an altered B pocket composition having either 6 amino acid substitutions (B27.A2B; substitutions H9F, T24A, E45M, I66K, C67V, and K70H) or a single substitution (B27.E45M) were no longer permissive of S enteritidis replication. In contrast, cells expressing B27 with the single substitution of F for H at position 9 (B27.H9F) retained their permissiveness. Studies using GFP-transformed S enteritidis confirmed that the increase in the amount of intracellular bacteria in B27-expressing cells was due to replication of the bacteria.
  
  
  CONCLUSIONS
  Our data indicate that HLA-B27 expression modulates the host-microbe interaction that results in an impaired capacity of monocytes to resist intracellular replication of S enteritidis. The phenotype is dependent on glutamic acid at position 45 in the B pocket and, thus, may be due to properties of the B27 heavy chain that are related to this residue. The ability of HLA-B27 to confer susceptibility to Salmonella-triggered reactive arthritis may occur, at least in part, through these modulatory effects.
authors.affiliation: Department of Human Microbial Ecology and Inflammation, National Public Health Institute, Turku, Finland. markus.penttinen@utu.fi
authors.forename: Markus A
authors.initials: MA
authors.lastname: Penttinen
authors.forename: Kaisa M
authors.initials: KM
authors.lastname: Heiskanen
authors.forename: Rajashree
authors.initials: R
authors.lastname: Mohapatra
authors.forename: Monica L
authors.initials: ML
authors.lastname: DeLay
authors.forename: Robert A
authors.initials: RA
authors.lastname: Colbert
authors.forename: Lea
authors.initials: L
authors.lastname: Sistonen
authors.forename: Kaisa
authors.initials: K
authors.lastname: Granfors
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism; D018014:Gene Transfer Techniques; D049452:Green Fluorescent Proteins; D015796:HLA-B27 Antigen / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015395:Histocompatibility Antigens Class I / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007425:Intracellular Membranes / Q000382:microbiology*; D008164:Luminescent Proteins / Q000235:genetics; D008715:Methionine; D009000:Monocytes / Q000276:immunology* / Q000382:microbiology*; D010641:Phenotype; D017510:Protein Folding; D012480:Salmonella Infections / Q000382:microbiology; D012477:Salmonella enteritidis / Q000254:growth & development*; D013997:Time Factors; D014162:Transfection
chemical_list: D015796:HLA-B27 Antigen; D015395:Histocompatibility Antigens Class I; D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins; D008715:Methionine