Identification of <i>NRAS</i> Downstream Genes with CRISPR Activation Screening
Mutations in <i>NRAS</i> constitutively activate cell proliferation signaling in malignant neoplasms, such as leukemia and melanoma, and the clarification of comprehensive downstream genes of <i>NRAS</i> might lead to the control of cell-proliferative signals of <i>NRAS...
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MDPI AG
2022-10-01
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| author | Akiya Tatsumi Haruka Hirakochi Satomi Inoue Yosuke Tanaka Hidehiro Furuno Masumi Ikeda Sachiko Ishibashi Towako Taguchi Kouhei Yamamoto Iichiroh Onishi Zohar Sachs David A. Largaespada Masanobu Kitagawa Morito Kurata |
| author_facet | Akiya Tatsumi Haruka Hirakochi Satomi Inoue Yosuke Tanaka Hidehiro Furuno Masumi Ikeda Sachiko Ishibashi Towako Taguchi Kouhei Yamamoto Iichiroh Onishi Zohar Sachs David A. Largaespada Masanobu Kitagawa Morito Kurata |
| author_sort | Akiya Tatsumi |
| collection | DOAJ |
| description | Mutations in <i>NRAS</i> constitutively activate cell proliferation signaling in malignant neoplasms, such as leukemia and melanoma, and the clarification of comprehensive downstream genes of <i>NRAS</i> might lead to the control of cell-proliferative signals of <i>NRAS</i>-driven cancers. We previously established that <i>NRAS</i> expression and proliferative activity can be controlled with doxycycline and named as THP-1 B11. Using a CRISPR activation library on THP-1 B11 cells with the <i>NRAS</i>-off state, survival clones were harvested, and 21 candidate genes were identified. By inducting each candidate guide RNA with the CRISPR activation system, <i>DOHH</i>, <i>HIST1H2AC</i>, <i>KRT32</i>, and <i>TAF6</i> showed higher cell-proliferative activity. The expression of <i>DOHH</i>, <i>HIST1H2AC,</i> and <i>TAF6</i> was definitely upregulated with <i>NRAS</i> expression. Furthermore, MEK inhibitors resulted in the decreased expression of DOHH, HIST1H2AC, and TAF6 proteins in parental THP-1 cells. The knockdown of <i>DOHH</i>, <i>HIST1H2AC</i>, and <i>TAF6</i> was found to reduce proliferation in THP-1 cells, indicating that they are involved in the downstream proliferation of <i>NRAS</i>. These molecules are expected to be new therapeutic targets for <i>NRAS</i>-mutant leukemia cells. |
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| institution | Directory Open Access Journal |
| issn | 2079-7737 |
| language | English |
| last_indexed | 2024-03-09T19:16:31Z |
| publishDate | 2022-10-01 |
| publisher | MDPI AG |
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| spelling | doaj.art-c9febd99cf9d4275864311a1c8b8ea412023-11-24T03:47:18ZengMDPI AGBiology2079-77372022-10-011111155110.3390/biology11111551Identification of <i>NRAS</i> Downstream Genes with CRISPR Activation ScreeningAkiya Tatsumi0Haruka Hirakochi1Satomi Inoue2Yosuke Tanaka3Hidehiro Furuno4Masumi Ikeda5Sachiko Ishibashi6Towako Taguchi7Kouhei Yamamoto8Iichiroh Onishi9Zohar Sachs10David A. Largaespada11Masanobu Kitagawa12Morito Kurata13Department of Comprehensive Pathology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Bunkyo-ku, Tokyo 113-8510, JapanDepartment of Comprehensive Pathology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Bunkyo-ku, Tokyo 113-8510, JapanDepartment of Comprehensive Pathology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Bunkyo-ku, Tokyo 113-8510, JapanDepartment of Comprehensive Pathology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Bunkyo-ku, Tokyo 113-8510, JapanDepartment of Comprehensive Pathology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Bunkyo-ku, Tokyo 113-8510, JapanDepartment of Comprehensive Pathology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Bunkyo-ku, Tokyo 113-8510, JapanDepartment of Comprehensive Pathology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Bunkyo-ku, Tokyo 113-8510, JapanDepartment of Comprehensive Pathology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Bunkyo-ku, Tokyo 113-8510, JapanDepartment of Comprehensive Pathology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Bunkyo-ku, Tokyo 113-8510, JapanDepartment of Comprehensive Pathology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Bunkyo-ku, Tokyo 113-8510, JapanMasonic Cancer Center, University of Minnesota, Minneapolis, MN 55455, USAMasonic Cancer Center, University of Minnesota, Minneapolis, MN 55455, USADepartment of Comprehensive Pathology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Bunkyo-ku, Tokyo 113-8510, JapanDepartment of Comprehensive Pathology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Bunkyo-ku, Tokyo 113-8510, JapanMutations in <i>NRAS</i> constitutively activate cell proliferation signaling in malignant neoplasms, such as leukemia and melanoma, and the clarification of comprehensive downstream genes of <i>NRAS</i> might lead to the control of cell-proliferative signals of <i>NRAS</i>-driven cancers. We previously established that <i>NRAS</i> expression and proliferative activity can be controlled with doxycycline and named as THP-1 B11. Using a CRISPR activation library on THP-1 B11 cells with the <i>NRAS</i>-off state, survival clones were harvested, and 21 candidate genes were identified. By inducting each candidate guide RNA with the CRISPR activation system, <i>DOHH</i>, <i>HIST1H2AC</i>, <i>KRT32</i>, and <i>TAF6</i> showed higher cell-proliferative activity. The expression of <i>DOHH</i>, <i>HIST1H2AC,</i> and <i>TAF6</i> was definitely upregulated with <i>NRAS</i> expression. Furthermore, MEK inhibitors resulted in the decreased expression of DOHH, HIST1H2AC, and TAF6 proteins in parental THP-1 cells. The knockdown of <i>DOHH</i>, <i>HIST1H2AC</i>, and <i>TAF6</i> was found to reduce proliferation in THP-1 cells, indicating that they are involved in the downstream proliferation of <i>NRAS</i>. These molecules are expected to be new therapeutic targets for <i>NRAS</i>-mutant leukemia cells.https://www.mdpi.com/2079-7737/11/11/1551CRISPR screeningNRAS |
| spellingShingle | Akiya Tatsumi Haruka Hirakochi Satomi Inoue Yosuke Tanaka Hidehiro Furuno Masumi Ikeda Sachiko Ishibashi Towako Taguchi Kouhei Yamamoto Iichiroh Onishi Zohar Sachs David A. Largaespada Masanobu Kitagawa Morito Kurata Identification of <i>NRAS</i> Downstream Genes with CRISPR Activation Screening Biology CRISPR screening NRAS |
| title | Identification of <i>NRAS</i> Downstream Genes with CRISPR Activation Screening |
| title_full | Identification of <i>NRAS</i> Downstream Genes with CRISPR Activation Screening |
| title_fullStr | Identification of <i>NRAS</i> Downstream Genes with CRISPR Activation Screening |
| title_full_unstemmed | Identification of <i>NRAS</i> Downstream Genes with CRISPR Activation Screening |
| title_short | Identification of <i>NRAS</i> Downstream Genes with CRISPR Activation Screening |
| title_sort | identification of i nras i downstream genes with crispr activation screening |
| topic | CRISPR screening NRAS |
| url | https://www.mdpi.com/2079-7737/11/11/1551 |
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