Global expression analysis revealed novel gender-specific gene expression features in the blood fluke parasite Schistosoma japonicum

Abstract

Background: Schistosoma japonicum is one of the remarkable Platyhelminths that are endemic in China and Southeast Asian countries. The parasite is dioecious and can reside inside the host for many years. Rapid reproduction by producing large number of eggs and count-react host anti-parasite responses are the strategies that benefit long term survival of the parasite. Praziquantel is currently the only drug that is effective against the worms. Development of novel antiparasite reagents and immune-prevention measures rely on the deciphering of parasite biology. The decoding of the genomic sequence of the parasite has made it possible to dissect the functions of genes that govern the development of the parasite. In this study, the polyadenylated transcripts from male and female S. japonicum were isolated for deep sequencing and the sequences were systematically analysed.

Results: First, the number of genes actively expressed in the two sexes of S. japonicum was similar, but around 50% of genes were biased to either male or female in expression. Secondly, it was, at the first time, found that more than 50% of the coding region of the genome was transcribed from both strands. Among them, 65% of the genes had sense and their cognate antisense transcripts co-expressed, whereas 35% had inverse relationship between sense and antisense transcript abundance. Further, based on gene ontological analysis, more than 2,000 genes were functionally categorized and biological pathways that are differentially functional in male or female parasites were elucidated.

Conclusions: Male and female schistosomal parasites differ in gene expression patterns, many metabolic and biological pathways have been identified in this study and genes differentially expressed in gender specific manner were presented. Importantly, more than 50% of the coding regions of the S. japonicum genome transcribed from both strands, antisense RNA-mediated gene regulation might play a critical role in the parasite biology.

Figure 1. Schematic illustration of the principle and procedure of Tag preparation.

Biotin-conjugated Oligo-dT was used to enrich mRNA and cDNA synthesis. The double strand cDNA was first digested with the 4 base (GTAC) recognition enzyme NlaIII, and Illumina adapter 1 was linked afterwards. Mmel was used to digest at 17 bp downstream of CATG site which was ligated with Illumina adapter 2 at the 3′ end. Sequencing anchor primers were added to the end of each fragment by PCR and the PCR product were purified and followed by Solexa sequencing.

Figure 2. Percentage of tags in copy number identified in the two libraries (A Male worm, B Female worm).

More than 60% of the tags identified in the two libraries are single copies.

Figure 3. Tags represented differential expression in male and female parasites.

A Distribution by Scatter plotting of expressed sequence tags identified in male and female parasites. Tags biased towards male parasite were in red color, while tags biased towards female parasite were labeled in green color. B Verification of gender-biased expression of 6 genes by real-time RT PCR. The differences in copy numbers of transcripts relative to that of α-tubulin were presented in log 10 scale.

Figure 4. Sequence tags identified from both sense and antisense strands of the genome.

A Gene numbers that with differential transcription patterns of the two DNA strands. Genes with more transcription from the sense strand were dominant. B Tags differentially expressed in male and female parasites.

Figure 5. Functional categorization of genes identified in male and female parasites.

A Number of genes that can be categorized into four main functional groups (Metabolic pathway, genetic information processing, environmental information processing and cellular processing). B Number of genes within the four functional categories that showed up- or down-regulation in male parasite compared to female counterpart.