Cloning and sequence analysis of a highly polymorphic Cryptosporidium parvum gene encoding a 60-kilodalton glycoprotein and characterization of its 15- and 45-kilodalton zoite surface antigen products.
The apicomplexan parasite Cryptosporidium parvum is a major cause of serious diarrheal disease in both humans and animals. No efficacious chemo- or immunotherapies have been identified for cryptosporidiosis, but certain antibodies directed against zoite surface antigens and/or proteins shed by gliding zoites have been shown to neutralize infectivity in vitro and/or to passively protect against, or ameliorate, disease in vivo. We previously used monoclonal antibody 11A5 to identify a 15-kDa surface glycoprotein that was shed behind motile sporozoites and was recognized by several lectins that neutralized parasite infectivity for cultured epithelial cells. Here we report the cloning and sequence analysis of the gene encoding this 11A5 antigen. Surprisingly, the gene encoded a 330-amino-acid, mucin-like glycoprotein that was predicted to contain an N-terminal signal peptide, a homopolymeric tract of serine residues, 36 sites of O-linked glycosylation, and a hydrophobic C-terminal peptide specifying attachment of a glycosylphosphatidylinositol anchor. The single-copy gene lacked introns and was expressed during merogony to produce a 60-kDa precursor which was proteolytically cleaved to 15- and 45-kDa glycoprotein products that both localized to the surface of sporozoites and merozoites. The gp15/45/60 gene displayed a very high degree of sequence diversity among C. parvum isolates, and the numerous single-nucleotide and single-amino-acid polymorphisms defined five to six allelic classes, each characterized by additional intra-allelic sequence variation. The gp15/45/60 single-nucleotide polymorphisms will prove useful for haplotyping and fingerprinting isolates and for establishing meaningful relationships between C. parvum genotype and phenotype.
Strong
W B
WB
Division of Infectious Diseases, San Francisco General Hospital, University of California, San Francisco, San Francisco, California 94143-0811, USA.
Gut
J
J
Nelson
R G
RG
D000483:Alleles; D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D000953:Antigens, Protozoan / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D000954:Antigens, Surface / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D003001:Cloning, Molecular; D003457:Cryptosporidiosis / Q000469:parasitology; D016785:Cryptosporidium parvum / Q000235:genetics* / Q000276:immunology* / Q000472:pathogenicity; D017931:DNA Primers / Q000235:genetics; D016054:DNA, Protozoan / Q000235:genetics; D018507:Gene Expression Regulation, Developmental; D017125:Genes, Protozoan; D005838:Genotype; D006801:Humans; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011110:Polymorphism, Genetic; D011499:Protein Processing, Post-Translational; D015800:Protozoan Proteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology*; D017386:Sequence Homology, Amino Acid
D000911:Antibodies, Monoclonal; D000953:Antigens, Protozoan; D000954:Antigens, Surface; D017931:DNA Primers; D016054:DNA, Protozoan; D015800:Protozoan Proteins