m6A 调控网络分析与验证：乳腺癌进展中的新型 circBACH2/has-miR-944/HNRNPC 轴
Analysis and validation of m6A regulatory network: a novel circBACH2/has-miR-944/HNRNPC axis in breast cancer progression.
机构信息
Department of Plastic and Cosmetic Surgery, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, 1095 Jiefang Avenue, Wuhan, 430030, Hubei, China.
出版信息
J Transl Med. 2021 Dec 24;19(1):527. doi: 10.1186/s12967-021-03196-4.
BACKGROUND
N6-methyladenosine (m6A), the most abundant and reversible modification of mRNAs in eukaryotes, plays pivotal role in breast cancer (BC) tumorigenesis and progression. Circular RNAs (circRNAs) can act as tumor promoters or suppressors by microRNA (miRNA) sponges in BC. However, the underlying mechanism of circRNAs in BC progression via regulating m6A modulators remains unclear.
METHODS
Prognostic m6A RNA methylation regulators were identified in 1065 BC patients from The Cancer Genome Atlas (TCGA) project. Differentially expressed (DE) miRNAs and DE circRNAs were identified between BC and normal samples in TCGA and GSE101123, respectively. MiRNA-mRNA interactive pairs and circRNA-miRNA interactive pairs were verified by MiRDIP and Circular RNA Interactome. GSEA, KEGG, and ssGSEA were executed to explore the potential biological and immune functions between HNRNPC-high and HNRNPC-low expression groups. qRT-PCR and Western blot were used to quantify the expression of HNRNPC and circBACH2 in MCF-7 and MDA-MB-231 cells. The proliferation of BC cells was assessed by CCK-8 and EdU assay.
RESULTS
2 m6A RNA methylation regulators with prognostic value, including HNRNPC and YTHDF3, were identified in BC patients. Then, the regulatory network of circRNA-miRNA-m6A modulators was constructed, which consisted of 2 DE m6A modulators (HNRNPC and YTHDF3), 12 DE miRNAs, and 11 DE circRNAs. Notably, BC patients with high expression of HNRNPC and low expression of hsa-miR-944 were correlated with late clinical stages and shorter survival times. Besides, the results from the KEGG inferred that the DE HNRNPC was associated with the MAPK signaling pathway in BC. Moreover, the circBACH2 (hsa_circ_0001625) was confirmed to act as hsa-miR-944 sponge to stimulate HNRNPC expression to promote BC cell proliferation via MAPK signaling pathway, thus constructing a circBACH2/hsa-miR-944/HNRNPC axis in BC.
CONCLUSIONS
Our findings decipher a novel circRNA-based m6A regulatory mechanism involved in BC progression, thus providing attractive diagnostic and therapeutic strategies for combating BC.
背景
N6-甲基腺苷（m6A）是真核生物中 mRNA 最丰富和最可逆转的修饰，在乳腺癌（BC）的发生和发展中起着关键作用。环状 RNA（circRNA）可以通过 miRNA（miRNA）海绵在 BC 中作为肿瘤促进剂或抑制剂发挥作用。然而，circRNA 通过调节 m6A 调节剂在 BC 进展中的潜在机制尚不清楚。
方法
从癌症基因组图谱（TCGA）项目中鉴定了 1065 名 BC 患者的预后 m6A RNA 甲基化调节剂。在 TCGA 和 GSE101123 中分别鉴定了 BC 和正常样本之间的差异表达（DE）miRNA 和 DE circRNA。通过 MiRDIP 和 Circular RNA Interactome 验证了 miRNA-mRNA 相互作用对和 circRNA-miRNA 相互作用对。执行 GSEA、KEGG 和 ssGSEA 以探索 HNRNPC-高和 HNRNPC-低表达组之间的潜在生物学和免疫功能。使用 qRT-PCR 和 Western blot 定量 MCF-7 和 MDA-MB-231 细胞中 HNRNPC 和 circBACH2 的表达。通过 CCK-8 和 EdU 测定评估 BC 细胞的增殖。
结果
在 BC 患者中鉴定出 2 种具有预后价值的 m6A RNA 甲基化调节剂，包括 HNRNPC 和 YTHDF3。然后，构建了 circRNA-miRNA-m6A 调节剂的调控网络，该网络由 2 个 DE m6A 调节剂（HNRNPC 和 YTHDF3）、12 个 DE miRNA 和 11 个 DE circRNA 组成。值得注意的是，HNRNPC 高表达和 hsa-miR-944 低表达的 BC 患者与晚期临床分期和较短的生存时间相关。此外，KEGG 的结果表明，DE HNRNPC 与 BC 中的 MAPK 信号通路有关。此外，circBACH2（hsa_circ_0001625）被证实可作为 hsa-miR-944 海绵，通过 MAPK 信号通路刺激 HNRNPC 表达，从而促进 BC 细胞增殖，从而在 BC 中构建了 circBACH2/hsa-miR-944/HNRNPC 轴。
结论
我们的研究结果揭示了一种新的涉及 BC 进展的基于 circRNA 的 m6A 调控机制，为对抗 BC 提供了有吸引力的诊断和治疗策略。
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