Nayoung Kim1, Miju Kim2, Sohyun Yun1, Junsang Doh2, Philip D Greenberg3, Tae-Don Kim4, Inpyo Choi5. 1. Immunotherapy Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, Korea. 2. School of Interdisciplinary Bioscience and Bioengineering (I-Bio), POSTECH, Pohang, Korea; Department of Mechanical Engineering, POSTECH, Pohang, Korea. 3. Departments of Immunology and Medicine, University of Washington School of Medicine, and Fred Hutchinson Cancer Research Center, Seattle, Wash. 4. Immunotherapy Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, Korea; Department of Functional Genomics, University of Science and Technology (UST), KRIBB, Daejeon, Korea. Electronic address: tdkim@kribb.re.kr. 5. Immunotherapy Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, Korea; Department of Functional Genomics, University of Science and Technology (UST), KRIBB, Daejeon, Korea. Electronic address: ipchoi@kribb.re.kr.
Abstract
BACKGROUND: Perforin-1 (Prf1) is the predominant cytolytic protein secreted by natural killer (NK) cells. For a rapid immune response, resting NK cells contain high Prf1 mRNA concentrations while exhibiting minimal cytotoxicity caused by a blockage of Prf1 protein synthesis, implying that an unknown posttranscriptional regulatory mechanism exists. OBJECTIVE: We sought to determine whether microRNA-150 (miR-150) posttranscriptionally regulates Prf1 translation in both mouse and human NK cells at rest and at various time points after activation. METHODS: Mouse NK cells with a targeted deletion of miR-150 (miR-150(-/-) NK cells), primary human NK cells, and NK92 MI cells were used to investigate the role of miR-150 in NK cells. NK cell cytotoxicity assays and Western blotting proved that activated miR-150(-/-) NK cells expressed upregulated Prf1, augmenting NK cell cytotoxicity. When immunodeficient mice were injected with miR-150(-/-) NK cells, there was a significant reduction in tumor growth and metastasis of B16F10 melanoma. RESULTS: We report that miR-150 binds to 3' untranslated regions of mouse and human Prf1, posttranscriptionally downregulating its expression. Mouse wild-type NK cells displayed downregulated miR-150 expression in response to IL-15, which led to corresponding repression and induction of Prf1 during rest and after IL-15 activation, respectively. CONCLUSION: Our results indicate that miR-150 is a common posttranscriptional regulator for Prf1 in mouse and human NK cells that represses NK cell lytic activity. Thus the therapeutic control of miR-150 in NK cells could enhance NK cell-based immunotherapy against cancer, providing a better clinical outcome.
BACKGROUND:Perforin-1 (Prf1) is the predominant cytolytic protein secreted by natural killer (NK) cells. For a rapid immune response, resting NK cells contain high Prf1 mRNA concentrations while exhibiting minimal cytotoxicity caused by a blockage of Prf1 protein synthesis, implying that an unknown posttranscriptional regulatory mechanism exists. OBJECTIVE: We sought to determine whether microRNA-150 (miR-150) posttranscriptionally regulates Prf1 translation in both mouse and humanNK cells at rest and at various time points after activation. METHODS:MouseNK cells with a targeted deletion of miR-150 (miR-150(-/-) NK cells), primary humanNK cells, and NK92 MI cells were used to investigate the role of miR-150 in NK cells. NK cell cytotoxicity assays and Western blotting proved that activated miR-150(-/-) NK cells expressed upregulated Prf1, augmenting NK cell cytotoxicity. When immunodeficientmice were injected with miR-150(-/-) NK cells, there was a significant reduction in tumor growth and metastasis of B16F10 melanoma. RESULTS: We report that miR-150 binds to 3' untranslated regions of mouse and humanPrf1, posttranscriptionally downregulating its expression. Mouse wild-type NK cells displayed downregulated miR-150 expression in response to IL-15, which led to corresponding repression and induction of Prf1 during rest and after IL-15 activation, respectively. CONCLUSION: Our results indicate that miR-150 is a common posttranscriptional regulator for Prf1 in mouse and humanNK cells that represses NK cell lytic activity. Thus the therapeutic control of miR-150 in NK cells could enhance NK cell-based immunotherapy against cancer, providing a better clinical outcome.
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