Profiling Schistosoma mansoni development using serial analysis of gene expression (SAGE)

Abstract

Despite the widespread use of chemotherapy and other control strategies over the past 50years, transmission rates for schistosomiasis have changed little. Regardless of the approach used, future control efforts will require a more complete understanding of fundamental parasite biology. Schistosomes undergo complex development involving an alteration of parasite generations within a mammalian and freshwater molluscan host in the completion of its lifecycle. Little is known about factors controlling schistosome development, but understanding these processes may facilitate the discovery of new control methods. Therefore, our goal in this study is to determine global developmentally regulated and stage-specific gene expression in Schistosoma mansoni using serial analysis of gene expression (SAGE). We present a preliminary analysis of genes expressed during development and sexual differentiation in the mammalian host and during early larval development in the snail host. A number of novel, differentially expressed genes have been identified, both within and between the different developmental stages found in the mammalian and snail hosts.

Fig. 1

Hierarchical clustering of 502, differentially expressed (R > 4) PS tags from all stages profiled was done using Cluster 3.00 and Treeview 1.0.13 programs (Stanford MicroArray Database). Red indicates up regulation and green indicates down regulation relative to the median abundance (black) for each tag. Abbreviations used for stage SAGE libraries are sub-adult liver, SA_Liver; adult female and male from single-sexed infections, AF_SS and AM_SS; adult male and female from mixed sex infections, AM_MS and AF_MS; miracidia, MIRA; 6-day mother sporocysts without or with Bge-conditioned media, 6D_S_UN and 6D_S_COND; and 20-day sporocyst without or with Bge-conditioned media 20D_S_UN and 20D_S_COND.

Fig. 2

SAGE profiling of Hox genes during S. mansoni lifecycle. A. SAGE tags from all 10 profiled stages for S. mansoni Fushi Tarazu-Factor 1 interacting protein (SmFIP-1), labial (SmHox1), deformed (SmHox4), and abdominal A (SmHox8) are shown. In this figure, and in Figures 4 and 5, for each gene examined the stages are shown in the following order: 1–5, mammalian host stages (closed bars), 1: sub-adult liver; 2: adult males from single-sex infections; 3: adult females from single-sex infections; 4: adult males from mixed sex infections; 5 adult females from mixed sex infections; 6: miracidia (open bar); 7–10 invertebrate host stages (gray bars), 7: 6-day mother sporocysts without Bge-conditioned media; 8: 6-day mother sporocysts with Bge-conditioned media; 9: 20-day sporocysts without Bge-conditioned media; and 10: 20-day sporocysts with Bge-conditioned media. B. SAGE tags for two constitutive, highly expressed genes, beta-tubulin and glyceraldehyde 3-phosphate dehydrogenase (GAPDH), are shown. Tag libraries as in A.

Fig. 3

Semi-quantitative reverse-transcription – PCR analysis of SAGE tags. Identity of the SAGE tags and the number of cycles used in the PCR are: 2, GAPDH, 21 cycles; 13441, high affinity cationic amino acid transporter, 36 cycles; 15463, purine nucleoside phosphorylase, 35 cycles; 25004, lamin B receptor, 36 cycles; 10596, histone H3.3, 33 cycles; 4, fatty acid binding protein, 21 cycles; 10, 28 kDa glutathione S-transferase, 23 cycles; 1885, similar to nuclear ribonucleoprotein K, 36 cycles; and 684, similar to nucleoside diphosphate kinase, 30 cycles.

Fig. 4

SAGE profiling energy metabolism during S. mansoni development. Stage-specific SAGE-tag libraries are shown as indicated in Figure 2. GAPDH, glyceraldehyde 3-phosphate dehydrogenase.

Fig. 5

SAGE profiling redox enzymes during S. mansoni development. Stage-specific SAGE-tag libraries are shown as indicated in Figure 2. Abbreviations used: Prx, peroxiredoxin; GPx, glutathione peroxidase; SOD, superoxide dismutase; Sec-SOD, secretory SOD; GST, glutathione S-transferase. The bar for adult female worms from mixed sex infections does not accurately indicate the SAGE tag percent, which is 0.71%, or 20 times the level found in adult male worms from mixed sex infections.