Abstract
This paper describes the purification of yolk proteins, which are important for the reproduction of egg-laying animals, and the structural characterization of two vitellogenins, VT1 and OTI-VIT-6, of the nematode Oscheius tipulae. O. tipulae is an alternative model organism to its relative, the widely used Caenorhabditis elegans, and is a good model to understand reproduction in insect parasitic nematodes of the genus Heterorhabditis. The native purified O. tipulae vitellogenin is composed of three polypeptides (VT1, VT2 and VT3), whereas in C. elegans, vitellogenin is composed of four polypeptides. The gene (Oti-vit-1) encoding yolk polypeptide VT1 has been recently identified in the genome of O. tipulae. Immunoblotting and N-terminal sequencing confirmed that VT1 is indeed coded by Oti-vit-1. Utilizing the same experimental approaches, we showed that the polypeptides VT2 and VT3 are derived from the proteolytic processing of the C- and N-terminal portions of the precursor OTI-VIT-6, respectively. We also showed that the recombinant polypeptide (P40), corresponding to the N-terminal sequence of OTI-VIT-6, preferentially interacts with a 100-kDa polypeptide found in adult worm extracts, as we have previously shown for the native vitellins of O. tipulae. Using the putative nematode vitellogenin amino acid sequences available in the UniProtKB database, we constructed a phylogenetic tree and showed that the O. tipulae vitellogenins characterized in this study are orthologous to those of the Caenorhabditis spp. Together, these results represent the first structural and functional comparative study of nematode yolk proteins outside the Caenorhabditis genus and provide insight into the evolution of these lipoproteins within the Nematode Phylum.
Highlights
Nematode egg production is a key strategy for the colonization of new habitats and hosts
We showed the position of the vitellins VT2 and VT3 in the precursor polypeptide (OTI-VIT-6) using Western Blotting with antisera raised against recombinant polypeptides of the N-terminal sequence of VT3 and C-terminal sequence of VT2
The N-terminal sequencing of VT1 resulted in clear nonambiguous signals that corresponded to the amino acid sequence
Summary
Nematode egg production is a key strategy for the colonization of new habitats and hosts. The primary stored proteins are the vitellins, which are lipoproteins that share a common ancestor that predates the origin of the bilateral metazoans [1,2,3,4,5,6]. Vitellins originate from an extra-ovarian protein, named vitellogenin, utilized by the growing oocyte [7]. Vitellogenin is processed in endosomal compartments and stored in the yolk granules until further use by the developing embryo or recently emerged L1 larvae [10]. Yolk proteins and vitellogenins have been studied to understand the structure and function of other lipoproteins, including human apolipoprotein B [11] or in environmental research [12]
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