Cloning And Analysis Of Schistosoma Japanicum Wnt Signaling Pathway Receptors And Ligands

Schistosomiasis is one of the world wide spread parasitic diseases that causes serious healthy problems to human and other suspcetible mammals.Praziquantel is the only chemotherapy drug to treat this disease. therapeutic effect of drugs is satisfactory,but there is a risk of drug resistance. Therefore,the anti-schistosomiasis some new drugs developing is a promblem that we need to solve quickly.Many biological processes including growth development are cotrolled by diverse signaling network in cellular communication.Wnt signaling pathway is widespread muiticellular eukaryotes vivo conserved signaling pathways involved development procecess.In our previous work,we first cloned five potential signal proteins SjWnt1、SjWnt2、SjWnt4、SjWnt5、SjWnt11,two receptors SjFz5,SjFz9and some other elements of S.japonnicum wnt signaling.We declued that the Wnt signaling pathway is existance in Schistosoma japonicum through bioinformatic and experiment.In the view of the fact that binding of wnt ligands to Frizzled (Fz)receptor is the beginning of wnt signaling,It is crucial to elucidate the bingding mode of Wnt-Fz.We get the EST sequences of Schistosoma japonicum Frizzled4(SjFz4)by the NCBI BLAST.with RACE and ELECTRONIC assembling methods,We successfully choned the fully5041bp sequence of S.japonicum (SjFz4) cDNA,which contained a complete OFR with3426bp nucleotide,ecoding1142amino acid with relative molecular weight of131.14ku.theoretical isoelectric point is8.35.The SjFz4protein shared general feature of Frizzled family protein:N-termind cysteine rich domain(CRD)and seven transmenbrance spanning segments.Signal peptide software prediction is not the signal peptide stucture.We recombinded CRD domain of SjFz4,SjFz7,SjFz8with pET28a and sucessfully expressed the His-tagged recombinants in prokaryotic system.Expressed molecular weight is25ku,31.7ku,19.7ku.For the ani-serum of SjFzs and immunnohistochemistory experiment provide experimental basis.We investigated mRNA expression patterns of the SjFz8during different life stages in definitive hosts. Quantitative real-time PCR analysis revealed that SjFz8mRNA was highly expressed in schistosomulum stages and follow by egg during the13days after downreguiation of the expressed level and beyond is maintained at a minimum level. The expression profiles of these SjFz8gene suggest that they may participate in developmental processes of early stage schistosomulum and reproductive organ of schistosoma adult worms. Schistosoma japonicum SjWntl and SjWnt5gene structure is analyzed by Genome Walking technology.SjWnt1contain six introns and seven exons.SjWnt5contains four intros and five exons.In conclusion,we successfully obtain a new member of S.japonicum Frizzled receptor family,SjFz4cDNA and recombinded CRD of SjFz4,SjFz8with pET28a,All the data could provide basis for further studying on the detailed funtion of SjFZs gene and idetifying the Wnt-Fz interation mode of S.japonicum wnt signaling.