Up-regulation of NKG2F receptor, a functionally unknown killer receptor, of human natural killer cells by interleukin-2 and interleukin-15.
The NKG2 family receptors are C-type lectin, type II transmembrane molecules, and play important roles in regulation of natural killer (NK) cell functions against tumor and virus. NKG2F is a new member of NKG2 family, and may possibly associate with DAP 12 to activate NK cells. Since lacking available antibody against human NKG2F, the features of NKG2F expression on NK cells remains unclear. In this study, human NKG2F recombinant expression in E. coli was carried out by using pET-28a with a hexahistidine (6x His) tag and a thrombin digestion sequence to the N-terminus of the recombinant protein NKG2F. IPTG (isopropyl-beta-d-thio-galactoside) induction resulted in high expression of recombinant NKG2F protein, which was then purified and identified by anti-His western blotting and LC-MS/MS. Polyclonal antibody was produced by immunization of BALB/c mice with recombinant NKG2F, and then used to detect NKG2F in western blotting and flow cytometry. Our results demonstrated that NKG2F was expressed only by PBMCs but not by human NK cell lines such as NKL and YT at mRNA level. It was observed that NKG2F was expressed on surface of human blood NK cells, and may be up-regulated at mRNA level and protein level after IL-2 or IL-15 stimulation.
Huang
Haihua
H
Institute of Immunology, Hefei National Laboratory for Physical Sciences at Microscale and School of Life Sciences, University of Science and Technology of China, Hefei, P.R. China.
Wang
Xin
X
Zhang
Yukun
Y
Zheng
Xiaodong
X
Wei
Haiming
H
Sun
Rui
R
D000818:Animals; D015153:Blotting, Western; D002469:Cell Separation; D002853:Chromatography, Liquid; D005434:Flow Cytometry; D006801:Humans; D019409:Interleukin-15 / Q000276:immunology*; D007376:Interleukin-2 / Q000276:immunology*; D007694:Killer Cells, Natural / Q000378:metabolism*; D008297:Male; D013058:Mass Spectrometry; D051379:Mice; D008807:Mice, Inbred BALB C; D055654:NK Cell Lectin-Like Receptor Subfamily C / Q000096:biosynthesis* / Q000276:immunology / Q000302:isolation & purification*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015854:Up-Regulation
D019409:Interleukin-15; D007376:Interleukin-2; C527066:KLRC4 protein, human; D055654:NK Cell Lectin-Like Receptor Subfamily C