Molecular and functional characterisation of the heat shock protein 10 of Strongyloides ratti.
Strongyloides stercoralis and S. ratti are intestinal parasitic nematodes infecting rats and humans, respectively. Both present extraordinary life cycles comprising a free-living generation in addition to parasitic stages. In search of molecules possibly involved in parasite-host interaction, we performed mass spectrometry to identify excretory/secretory products of S. ratti. Amongst others we detected homologs of the heat shock proteins HSP10 and HSP60 (Sr-HSP10 and Sr-HSP60). HSPs are well known as chaperones involved in stress responses of cells, but recent studies suggest additional roles of small HSPs for parasite biology including immune modulation. To characterise Sr-HSP10, we cloned its full-length cDNA, analysed the genomic organisation, tested its presumptive role as an interaction partner of Sr-HSP60, studied its transcription in the parasite, and expressed the protein to test its immune responses. The cDNA contains an open reading frame of 330bp encoding a polypeptide of 110 amino acids with an approximate molecular weight of 10kDa. The Sr-HSP10 protein is highly homologous to that of the human pathogen S. stercoralis with only eight amino acid substitutions. Analysis of the genomic organisation of the Sr-HSP10 locus revealed that the gene is linked head-to-head to the gene encoding Sr-HSP60, and both share a bidirectional promoter. RT-PCR experiments indicated potential independent expression of the Sr-HSPs genes. In situ hybridisation results demonstrate Sr-HSP10 transcription in the gut area. Mammalian and yeast two-hybrid assays show dimerisation of Sr-HSP10, but no binding to recombinant Sr-HSP60. Immunisation experiments finally revealed a strong immunogenicity of Sr-HSP10 and provided evidence for a role in regulating the host-parasite interaction.
Tazir
Yasmina
Y
Bernhard Nocht Institute for Tropical Medicine, Bernhard-Nocht-Strasse 74, 20359 Hamburg, Germany.
Steisslinger
Vera
V
Soblik
Hanns
H
Younis
Abuelhassan Elshazly
AE
Beckmann
Svenja
S
Grevelding
Christoph G
CG
Steen
Hanno
H
Brattig
Norbert W
NW
Erttmann
Klaus D
KD
D000818:Animals; D000909:Antibodies, Helminth / Q000097:blood; D000947:Antigens, Helminth / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D001483:Base Sequence; D018835:Chaperonin 10 / Q000737:chemistry / Q000235:genetics* / Q000276:immunology / Q000378:metabolism*; D018834:Chaperonin 60 / Q000378:metabolism; D018509:DNA, Helminth / Q000737:chemistry / Q000235:genetics; D019281:Dimerization; D020869:Gene Expression Profiling; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011485:Protein Binding; D025941:Protein Interaction Mapping; D051381:Rats; D017208:Rats, Wistar; D017422:Sequence Analysis, DNA; D017170:Strongyloides ratti / Q000502:physiology*; D020798:Two-Hybrid System Techniques
D000909:Antibodies, Helminth; D000947:Antigens, Helminth; D018835:Chaperonin 10; D018834:Chaperonin 60; D018509:DNA, Helminth