Gene expression profile of the SmRad23 and SmRad4, from the nucleotide excision repair, in Schistosoma mansoni adult worms after exposition to different DNA-damaging agents
Silva, C.S. 1,2; Silva, S.H. 1; Pereira Júnior, O.S. 1; Macedo, E. 1; Brigatto, O.M. 1; Momo, P.B. 1; Costa-Cruz, J.M. 2; Rodrigues, V.1
1 Department of Biochemistry and Imunnology, School of Medicine, University of São Paulo, Ribeirão Preto, SP; 2 Department of Imunnology, Microbiology and Parasitology, Federal University of Uberlândia, MG.
DNA is often damaged by environmental agents, which lead to the up-regulation of several genes involved in different repair pathways. Schistosoma mansoni is a parasite that undergoes several modifications in its complex life cycle, being exposed to a subset of DNA-damaging agents, and therefore it is likely to be provided for efficient repair mechanisms. Recently, studies have shown that Nucleotide Excision Repair consists in an indispensable mechanism for removing a broad spectrum of DNA lesions. Objectives: To analyze the gene expression of Nucleotide Excision Repair Factor 2 - SmRad23 and SmRad4, in S. mansoni life cycle, as well as the expression level of these genes in S. mansoni adult worms treated with DNA-damaging agents. Methodology: The comet assay was used in order to verify the DNA damaged. RT-PCR was performed using total RNA (adult worms, eggs, hepatopancreas of infected and non-infected Biomphalaria snails, cercariae, schistosomulae) for qualitative PCR, and (cultivated adult worms: colchicin, TMA, H2O2, control) for the semi quantitative PCR. The transcript was amplified using the primers for Rad23 and Rad4, designed from the S. mansoni transcriptome project, ONSA-Fapesp. The PCR product was cloned, and sequenced in ABI-3100. The homology search was developed using BLAST and the alignment of the predict aminoacids sequence using the ClustalW. It was performed a densitometry of the semi quantitative PCR. Results: It was confirmed the expression of SmRad23 and SmRad4 in all of the developmental stages studied of the parasite, and shown a differential expression after treatment with the chemical agents. Furthermore, it was revealed the correlation of these genes with their orthologues. Conclusion: The presence of SmRad23 and SmRad4 in all of the developmental stages of S. mansoni, as well as their differential expression following exposition to DNA-damaging agents, suggest that NER is an important repair pathway during the complex life cycle of this parasite. Financial Support: CAPES, FAPESP, CNPq, FAEPA.
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