Identification of nuclear localization sequence of CXCR4 in renal cell carcinoma by constructing expression plasmids of different deletants.
We previously firstly discovered that CXCR4 nuclear localization may be responsible for metastasis of renal cell carcinoma (RCC), and that there was a nuclear localization sequence (NLS) that induced CXCR4 to transfer to the nucleus after combining with its ligand SDF-1. Using our previously constructed pEGFP-CXCR4 as the template, corresponding objective regions were amplified. The amplified PCR products were then digested and inserted into the pMD19-T simple vector and subcloned into the pEGFP-N1 vector. A recombinant expression vector containing different regions of CXCR4 was successfully constructed. After transfecting the recombinant expression vectors to RCC A498 cells, the intracellular locations of recombinant protein were examined by confocal microscopy. It was found that nuclear localization sequence of CXCR4 was located in amino acids 90-170, which accorded with the results of bioinformatics analysis software. The present study firstly discovered the NLS region of CXCR4, which may prove valuable for seeking new strategies to inhibit metastasis of RCC.
Wang
Lin-Hui
LH
Department of Urology, Changhai Hospital, 168 Changhai Road, Shanghai 200433, China.
Liu
Qian
Q
Xu
Bin
B
Chen
Wei
W
Yang
Qing
Q
Wang
Zhi-Xiang
ZX
Sun
Ying-Hao
YH
D002292:Carcinoma, Renal Cell / Q000378:metabolism*; D045744:Cell Line, Tumor; D005822:Genetic Vectors; D006801:Humans; D042541:Intracellular Space / Q000378:metabolism; D007680:Kidney Neoplasms / Q000378:metabolism*; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D019913:Nuclear Localization Signals / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics*; D016133:Polymerase Chain Reaction; D019718:Receptors, CXCR4 / Q000737:chemistry* / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D017384:Sequence Deletion
D050505:Mutant Proteins; D019913:Nuclear Localization Signals; D019718:Receptors, CXCR4; D011993:Recombinant Fusion Proteins