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Journal of Experimental & Clinical Cancer Research

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Open Access 01-12-2023 | Sarcoma | Research

CRISPR-Cas9 knockout screening identifies KIAA1429 as an essential gene in Ewing sarcoma

Authors: Kezhe Tan, Wenjie Lu, Feng Chen, Hao Shi, Yingxuan Ma, Zhou Chen, Wei Wu, Zhibao Lv, Jialin Mo

Published in: Journal of Experimental & Clinical Cancer Research | Issue 1/2023

Abstract

Background

Ewing sarcoma (ES) is an aggressive childhood bone and soft tissue cancer. KIAA1429 is one type of N6-methyladenosine (m6A) writer that plays a tumor-progressive role in various cancers, but the role of KIAA1429 in ES remains to be elucidated. The aim of the study was to investigate the role of KIAA1429 in ES.

Methods

We performed a multi-omic screen including CRISPR-Cas9 functional genomic and transcriptomic approaches, and identified that KIAA1429 played a significant role in ES progression. Gene knockdown, quantitative real-time PCR (Q-RT-PCR), immunoblotting, CellTiter-Glo assays, clonogenic assays, a subcutaneous xenograft model and immunohistochemistry were used to assess the functional role of KIAA1429 in ES. We mainly conducted RNA sequencing (RNA-seq) in ES cells to analyze the downstream regulatory mechanism of KIAA1429. An integrative analysis of chromatin immunoprecipitation sequencing (ChIP-seq) and RNA-seq indicated the upstream regulatory mechanism of KIAA1429.

Results

In vitro and in vivo CRISPR-Cas9 knockout screening identified KIAA1429 as an ES-dependent gene. Genetic suppression of KIAA1429 inhibited ES cell proliferation and tumorigenicity both in vitro and in vivo. Further studies revealed that KIAA1429 promotes ES tumorigenesis by regulating the ribosome-associated cell cycle and cancer-related inflammation. Interestingly, we found that STAT3 was a target of KIAA1429 and that a STAT3 inhibitor reduced KIAA1429 transcript levels, indicating positive feedback between KIAA1429 and STAT3. Finally, we found that NKX2-2 bound to the KIAA1429 promoter and transactivated KIAA1429.

Conclusion

Our study systematically analyzed ES-dependent epigenetic/transcriptional regulatory genes and identified KIAA1429 as a biomarker of tumor progression in ES, providing a potential therapeutic target for treating ES.

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Title: CRISPR-Cas9 knockout screening identifies KIAA1429 as an essential gene in Ewing sarcoma
Authors: Kezhe Tan
Wenjie Lu
Feng Chen
Hao Shi
Yingxuan Ma
Zhou Chen
Wei Wu
Zhibao Lv
Jialin Mo
Publication date: 01-12-2023
Publisher: BioMed Central
Keyword: Sarcoma
Published in: Journal of Experimental & Clinical Cancer Research / Issue 1/2023
Electronic ISSN: 1756-9966
DOI: https://doi.org/10.1186/s13046-023-02828-5

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