[Tissular and subcellular localization of the ATP synthase B subunit in Clonorchis sinensis].
OBJECTIVE
To illustrate the distribution of ATP synthase b subunit in the tissue of Clonorchis sinensis adult and its subcellular mimical localization in HeLa cells.


METHODS
With the antiserum against recombinant CsATP-synt_B protein raised from SD rats as primary antibody, paraffin sections of the adult of C. sinensis were processed by the method of fluorescent immunohistochemistry to observe the distribution of CsATP-synt_B protein in adult worm. According to the prediction by bioinformatics of the definite mitochondrial targeting sequence (MTS) and probable Bipartite nuclear localization signals (NLS_BP)in CsATP-syntB sequence, recombinant pEGFP-N1 plasmids containing the intact and three defective CsATP-synt_B sequence with single defect of MTS or NLS_BP or double defect respectively were constructed. The recombinant plasmids and the control plasmid-pEGFP-N1, pEYFP-Mito and H2B-CFP, were transfected into the HeLa cells by Lipofectamine 2000 reagent and the subcellular location of the GFP fusion protein was observed with confocal microscopy.


RESULTS
The CsATP-synt_B protein appeared to distribute all over the adult worm, especially abundant on the acetabulum, ovary, vitellarium and tegument. The intact CsATP-synt_B was definitely expressed in mitochondria and/or nucleus of infected HeLa cells, whereas the MTS-deleted mutant only in cytoplasma and nucleus, the NLS_BP-deleted mutant in mitochondria and cytoplasm, and the double defect mutant only in cytoplasm.


CONCLUSIONS
The distribution of CsATP-synt_B in adult is accord with that of mitochondria, and mainly exits in the organs and the tissues of active energy metabolism. This study first predicted and confirmed that CsATP-synt_B can be expressed in the nucleus.
Hu
Xu-Chu
XC
Department of Parasitology, Zhongshan College of Medicine, Sun Yat-sen University, Guangzhou 510080, China.
Zhou
Hong-Juan
HJ
Hu
Feng-Yu
FY
Ma
Chang-Ling
CL
Zhao
Jun-Hong
JH
Huang
Can
C
Zheng
Xiao-Ling
XL
Xu
Jin
J
Yu
Xin-Bing
XB
D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism; D003004:Clonorchis sinensis / Q000201:enzymology* / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism; D006367:HeLa Cells / Q000469:parasitology; D006801:Humans; D008928:Mitochondria / Q000378:metabolism; D006180:Proton-Translocating ATPases / Q000378:metabolism*
D006180:Proton-Translocating ATPases