Figure 4

MSP analysis in whole nematode extracts. A: MSP is not detectable by Western blot in lysates of the parthenogenetic nematodes A. nanus and D. coronatus. Lysates of nematodes with various reproductive modes were, in roughly equal amounts, subjected to SDS-PAGE and Western blotting. One membrane of a duplicate set was incubated with monoclonal anti-MSP antibody, the other as a control with anti-α-tubulin antibody. While MSP is recognised in C. elegans and hermaphroditic or bisexual representatives of the distant Acrobeloides genus, none of the parthenogenetic species (violet) displays a signal. Ce: C. elegans; Abi: Acrobeloides (bisex); Ah: Acrobeloides (herma); An: A. nanus; Dc: D. coronatus. MW: molecular weight [kDa]. B: MSP is not detectable in additional parthenogenetic nematode species. As above, roughly equal amounts of nematode lysate were subjected to SDS-PAGE and Western blotting. While MSP is recognised in C. elegans and the sperm producing Acrobeloides (bisex), two further parthenogenetic species (violet) do not reveal a signal. Ce: C. elegans; Abi: Acrobeloides (bisex); An: A. nanus; Ax: A. maximus; Zp: Z. punctata. MW: molecular weight [kDa]. C: MSP is detectable over a wide range of concentrations. Serial dilutions of C. elegans lysates were subjected to SDS-PAGE and Western blotting using the monoclonal anti-MSP antibody. Numbers above the lanes indicate the relative amount of lysate used. 100% approximately correspond to 8 mg of worms, the amount used in experiments 4A and 4B. D: The C-terminal epitope recognised by the MSP antibody is widely conserved throughout the Nematode phylum. An alignment of C. elegans and other MSP sequences, derived from nematodes of clades 8-12 (Figure 1A), demonstrates the almost perfect conservation of MSP C-termini in a wide phylogenetic range. Accession numbers: Ce (C. elegans, AAC26926.1); Ov (Onchocerca volvulus, J04662.1); As (Ascaris suum, P27440.3); Gr (Globodera rostochiensis, AAA29146.1).