Transduction of Schistosoma japonicum schistosomules with vesicular stomatitis virus glycoprotein pseudotyped murine leukemia retrovirus and expression of reporter human telomerase reverse transcriptase in the transgenic schistosomes.
Although draft genome sequences of two of the major human schistosomes, Schistosoma japonicum and Schistosoma mansoni are available, the structures and characteristics of most genes and the influence of exogenous genes on the metabolism of schistosomes remain uncharacterized. Furthermore, which functional genomics approaches will be tractable for schistosomes are not yet apparent. Here, the vesicular stomatitis virus glycoprotein (VSVG)-pseudotyped pantropic retroviral vector pBABE-puro was modified to incorporate the human telomerase reverse transcriptase gene (hTERT) as a reporter, under the control of the retroviral long terminal repeat (LTR). Pseudotyped virions were employed to transduce S. japonicum to investigate the utility of retrovirus-mediated transgenesis of S. japonicum and the activity of human telomerase reverse transcriptase as a reporter transgene in schistosomes. Schistosomules perfused from experimentally infected rabbits were cultured for 6 days after exposure to the virions after which genomic DNAs from virus exposed and control worms were extracted. Analysis of RNA from transduced parasites and immunohistochemistry of thin parasite sections revealed expression of hTERT in the transduced worms. Expression of hTERT was also confirmed by immunoblot analysis. These findings indicated that S. japonicum could be effectively transduced by VSVG-pseudotyped retrovirus carrying the hTERT gene. Given the potential of hTERT to aid in derivation of immortalized cells, these findings suggest that this pantropic retroviral approach can be employed to transduce cells from specific tissues and organs of schistosomes to investigate the influence of transgene hTERT on growth and proliferation of schistosome cells.
Yang
Shenghui
S
Centre of Cell and Molecular Biology Experiment, Xiangya School of Medicine, Central South University, Changsha, Hunan province, China.
Brindley
Paul J
PJ
Zeng
Qingren
Q
Li
Yuesheng
Y
Zhou
Jun
J
Liu
Yan
Y
Liu
Biyuan
B
Cai
Liting
L
Zeng
Tiebing
T
Wei
Qi
Q
Lan
Lingmei
L
McManus
Donald P
DP
D000818:Animals; D030801:Animals, Genetically Modified / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D017930:Genes, Reporter; D006801:Humans; D015151:Immunoblotting; D007150:Immunohistochemistry; D009052:Leukemia Virus, Murine / Q000235:genetics* / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D041681:NIH 3T3 Cells; D016133:Polymerase Chain Reaction; D011817:Rabbits; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012549:Schistosoma japonicum / Q000235:genetics / Q000254:growth & development / Q000378:metabolism / Q000821:virology*; D019098:Telomerase / Q000235:genetics / Q000378:metabolism*; D014161:Transduction, Genetic; D019076:Transgenes; D014759:Viral Envelope Proteins / Q000235:genetics* / Q000378:metabolism
C028391:G protein, vesicular stomatitis virus; D008562:Membrane Glycoproteins; D014759:Viral Envelope Proteins; C509186:TERT protein, human; D019098:Telomerase