Functional Phosphoproteomics in Cancer Chemoresistance Using CRISPR‐Mediated Base Editors
Abstract Selective inhibition of targeted protein kinases is an effective therapeutic approach for treatment of human malignancies, which interferes phosphorylation of cellular substrates. However, a drug‐imposed selection creates pressures for tumor cells to acquire chemoresistance‐conferring mutat...
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| Format: | Article |
| Language: | English |
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Wiley
2022-10-01
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| Series: | Advanced Science |
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| Online Access: | https://doi.org/10.1002/advs.202200717 |
| _version_ | 1811235068490809344 |
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| author | Jianan Li Jianxiang Lin Shisheng Huang Min Li Wenxia Yu Yuting Zhao Junfan Guo Pumin Zhang Xingxu Huang Yunbo Qiao |
| author_facet | Jianan Li Jianxiang Lin Shisheng Huang Min Li Wenxia Yu Yuting Zhao Junfan Guo Pumin Zhang Xingxu Huang Yunbo Qiao |
| author_sort | Jianan Li |
| collection | DOAJ |
| description | Abstract Selective inhibition of targeted protein kinases is an effective therapeutic approach for treatment of human malignancies, which interferes phosphorylation of cellular substrates. However, a drug‐imposed selection creates pressures for tumor cells to acquire chemoresistance‐conferring mutations or activating alternative pathways, which can bypass the inhibitory effects of kinase inhibitors. Thus, identifying downstream phospho‐substrates conferring drug resistance is of great importance for developing poly‐pharmacological and targeted therapies. To identify functional phosphorylation sites involved in 5‐fluorouracil (5‐FU) resistance during its treatment of colorectal cancer cells, CRISPR‐mediated cytosine base editor (CBE) and adenine base editor (ABE) are utilized for functional screens by mutating phosphorylated amino acids with two libraries specifically targeting 7779 and 10 149 phosphorylation sites. Among the top enriched gRNAs‐induced gain‐of‐function mutants, the target genes are involved in cell cycle and post‐translational covalent modifications. Moreover, several substrates of RSK2 and PAK4 kinases are discovered as main effectors in responding to 5‐FU chemotherapy, and combinational treatment of colorectal cancer cells with 5‐FU and RSK2 inhibitor or PAK4 inhibitor can largely inhibit cell growth and enhance cell apoptosis through a RSK2/TP53BP1/γ‐H2AX phosphorylation signaling axis. It is proposed that this screen approach can be used for functional phosphoproteomics in chemotherapy of various human diseases. |
| first_indexed | 2024-04-12T11:45:56Z |
| format | Article |
| id | doaj.art-3138ac8b45f34d1c87bd25c3ed52f91a |
| institution | Directory Open Access Journal |
| issn | 2198-3844 |
| language | English |
| last_indexed | 2024-04-12T11:45:56Z |
| publishDate | 2022-10-01 |
| publisher | Wiley |
| record_format | Article |
| series | Advanced Science |
| spelling | doaj.art-3138ac8b45f34d1c87bd25c3ed52f91a2022-12-22T03:34:22ZengWileyAdvanced Science2198-38442022-10-01930n/an/a10.1002/advs.202200717Functional Phosphoproteomics in Cancer Chemoresistance Using CRISPR‐Mediated Base EditorsJianan Li0Jianxiang Lin1Shisheng Huang2Min Li3Wenxia Yu4Yuting Zhao5Junfan Guo6Pumin Zhang7Xingxu Huang8Yunbo Qiao9School of Life Science and Technology ShanghaiTech University Shanghai 201210 ChinaNinth People's Hospital Shanghai Jiao Tong University School of Medicine Shanghai 200125 ChinaZhejiang Lab Hangzhou Zhejiang 311121 ChinaPrecise Genome Engineering Center School of Life Sciences Guangzhou University Guangzhou 510006 ChinaSchool of Life Science and Technology ShanghaiTech University Shanghai 201210 ChinaPrecise Genome Engineering Center School of Life Sciences Guangzhou University Guangzhou 510006 ChinaSchool of Life Science and Technology ShanghaiTech University Shanghai 201210 ChinaZhejiang Provincial Key Laboratory of Pancreatic Disease The First Affiliated Hospital and Institute of Translational Medicine Zhejiang University School of Medicine Hangzhou 310029 ChinaSchool of Life Science and Technology ShanghaiTech University Shanghai 201210 ChinaNinth People's Hospital Shanghai Jiao Tong University School of Medicine Shanghai 200125 ChinaAbstract Selective inhibition of targeted protein kinases is an effective therapeutic approach for treatment of human malignancies, which interferes phosphorylation of cellular substrates. However, a drug‐imposed selection creates pressures for tumor cells to acquire chemoresistance‐conferring mutations or activating alternative pathways, which can bypass the inhibitory effects of kinase inhibitors. Thus, identifying downstream phospho‐substrates conferring drug resistance is of great importance for developing poly‐pharmacological and targeted therapies. To identify functional phosphorylation sites involved in 5‐fluorouracil (5‐FU) resistance during its treatment of colorectal cancer cells, CRISPR‐mediated cytosine base editor (CBE) and adenine base editor (ABE) are utilized for functional screens by mutating phosphorylated amino acids with two libraries specifically targeting 7779 and 10 149 phosphorylation sites. Among the top enriched gRNAs‐induced gain‐of‐function mutants, the target genes are involved in cell cycle and post‐translational covalent modifications. Moreover, several substrates of RSK2 and PAK4 kinases are discovered as main effectors in responding to 5‐FU chemotherapy, and combinational treatment of colorectal cancer cells with 5‐FU and RSK2 inhibitor or PAK4 inhibitor can largely inhibit cell growth and enhance cell apoptosis through a RSK2/TP53BP1/γ‐H2AX phosphorylation signaling axis. It is proposed that this screen approach can be used for functional phosphoproteomics in chemotherapy of various human diseases.https://doi.org/10.1002/advs.2022007175‐fluorouracil (5‐FU)base editorsfunctional phosphoproteomicsRSK2screen |
| spellingShingle | Jianan Li Jianxiang Lin Shisheng Huang Min Li Wenxia Yu Yuting Zhao Junfan Guo Pumin Zhang Xingxu Huang Yunbo Qiao Functional Phosphoproteomics in Cancer Chemoresistance Using CRISPR‐Mediated Base Editors Advanced Science 5‐fluorouracil (5‐FU) base editors functional phosphoproteomics RSK2 screen |
| title | Functional Phosphoproteomics in Cancer Chemoresistance Using CRISPR‐Mediated Base Editors |
| title_full | Functional Phosphoproteomics in Cancer Chemoresistance Using CRISPR‐Mediated Base Editors |
| title_fullStr | Functional Phosphoproteomics in Cancer Chemoresistance Using CRISPR‐Mediated Base Editors |
| title_full_unstemmed | Functional Phosphoproteomics in Cancer Chemoresistance Using CRISPR‐Mediated Base Editors |
| title_short | Functional Phosphoproteomics in Cancer Chemoresistance Using CRISPR‐Mediated Base Editors |
| title_sort | functional phosphoproteomics in cancer chemoresistance using crispr mediated base editors |
| topic | 5‐fluorouracil (5‐FU) base editors functional phosphoproteomics RSK2 screen |
| url | https://doi.org/10.1002/advs.202200717 |
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