Role of carbohydrate recognition domains of pertussis toxin in adherence of Bordetella pertussis to human macrophages.
Pertussis toxin (PT) and filamentous hemagglutinin can each mediate the association of Bordetella pertussis with human macrophages. Adherence via filamentous hemagglutinin leads to integrin-mediated entry and survival of the bacteria within the human cell. We determined the contribution of PT to bacterial adherence to human macrophages. Plating macrophages on wells coated with recombinant PT subunit 2 (S2) or S3 decreased PT-dependent bacterial binding by greater than 60%; S1, S4, and S5 were ineffective. S3-dependent adherence was reduced 63% +/- 8% by sialic acid, while S2-dependent adherence was reduced 53% +/- 11% by galactose. Loss of the carbohydrate recognition properties of S2 by deletion of residues 40 to 54 or site-specific mutations at Asn-93, His-47, or Arg-50 eliminated the ability of the subunit protein to competitively inhibit bacterial binding. Peptides corresponding to residues 28 to 45 of S2 and S3 competitively inhibited adherence. Treatment of macrophages with antibodies to Le(a) or Le(x) but not CD14, CD15, CD18, or HLA interfered with PT-mediated binding. Exposure of the macrophages to the B oligomer, S2, or S3 increased binding to the CD11b/CD18 integrin. These results indicate that the carbohydrate recognition domains of both S2 and S3 participate in adherence of B. pertussis to human macrophages. The PT receptor(s), as yet unidentified, appears to carry the Le(a) or Le(x) determinants and is functionally capable of modulating integrin-mediated binding to the macrophage.
Rockefeller University, New York, New York 10021.
J
J
van't Wout
W N
WN
Burnette
V L
VL
Mar
E
E
Rozdzinski
S D
SD
Wright
E I
EI
Tuomanen
D000595:Amino Acid Sequence; D015703:Antigens, CD / Q000378:metabolism; D001422:Bacterial Adhesion; D001886:Bordetella pertussis / Q000472:pathogenicity / Q000502:physiology*; D018821:CD18 Antigens; D050260:Carbohydrate Metabolism; D006801:Humans; D016023:Integrins / Q000502:physiology; D008264:Macrophages / Q000382:microbiology*; D008969:Molecular Sequence Data; D037342:Pertussis Toxin; D010566:Virulence Factors, Bordetella / Q000378:metabolism*
D015703:Antigens, CD; D018821:CD18 Antigens; D016023:Integrins; D010566:Virulence Factors, Bordetella; D037342:Pertussis Toxin