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Saposin-like proteins from the intestine of the blood-feeding hookworm, Ancylostoma caninum

Published online by Cambridge University Press:  17 November 2006

T. A. DON ,
Y. OKSOV ,
S. LUSTIGMAN  and
A. LOUKAS
T. A. DON
Affiliation:
Helminth Biology Laboratory, Division of Infectious Diseases and Immunology, Queensland Institute of Medical Research, Queensland 4006, Australia School of Molecular and Microbial Sciences, The University of Queensland, Queensland 4072, Australia
Y. OKSOV
Affiliation:
Lindsley F. Kimball Research Institute, The New York Blood Center, NY 10021, USA
S. LUSTIGMAN
Affiliation:
Lindsley F. Kimball Research Institute, The New York Blood Center, NY 10021, USA
A. LOUKAS
Affiliation:
Helminth Biology Laboratory, Division of Infectious Diseases and Immunology, Queensland Institute of Medical Research, Queensland 4006, Australia
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Abstract

Hookworms feed on blood, utilizing haemoglobin for nutrition, growth and reproduction. The haemoglobin digestion cascade has been partially elucidated, but the process immediately preceding this event, haemolysis, has received considerably less attention. We have cloned and expressed Ancylostoma caninum mRNAs encoding 2 proteins belonging to the saposin-like protein (SAPLIP) family, termed Ac-slp-1 and Ac-slp-2. The open reading frames of SLP-1 and SLP-2 were used to identify expressed sequence tags encoding SAPLIPs from the 4 major clades of animal parasitic nematodes. Both Ac-slp-1 and slp-2 mRNAs were shown to be expressed in all life stages assessed, with slp-1 predominantly being expressed in third-stage larvae (L3) before and after activation with dog serum. Recombinant SLP-1 and SLP-2 were expressed in insect cells and used to raise specific antisera in mice. These antisera were used as probes in fluorescence microscopy to localize the anatomic expression sites of both proteins to small, punctate organelles or vesicles within the intestinal cells of adult worms; weak staining was detected on the microvillar brush border of the intestine. Using transmission electron microscopy, both proteins were localized to similar vesicles in the intestinal cells of the L3. Recombinant proteins contained C-terminal purification tags that potentially precluded dimerization and possibly interfered with the subsequent detection of haemolytic activity. Their expression in the gut of the L3 and adult stages suggests a role for these hookworm SAPLIPs in the lysis of host cells during tissue migration and/or feeding.

Keywords

Ancylostoma caninumhookwormSAPLIPsaposin
Type
Research Article
Information
Parasitology , Volume 134 , Issue 3 , March 2007 , pp. 427 - 436
Copyright
© 2006 Cambridge University Press

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