虫草素抗三阴性乳腺癌的转录组学分析

doi:10.13560/j.cnki.biotech.bull.1985.2021-0444

摘要/Abstract

摘要：

前期研究表明,蛹虫草活性成分虫草素能有效抑制乳腺癌移植瘤的生长和转移,但其作用机制未清楚。本研究取三阴性乳腺癌移植瘤样品,通过转录组测序技术分析虫草素抗乳腺癌移植瘤的分子作用机制。结果表明,以P<0.05、log₂|FC|>1为标准,筛选获得584个差异表达基因,其中上调基因69个,下调基因515个;KEGG分析表明,虫草素对乳腺癌移植瘤的分子调控涉及Hippo signaling pathway、Hedgehog signaling pathway、ECM-receptor interction、TGF-beta signaling pathway等多条癌症相关信号通路;GO分析表明,这些差异基因富集在biological adhesion、growth、metabolic process、locomotion和biological process等16个生物学过程中,提示虫草素对乳腺癌移植瘤生长与转移有极显著的调节作用;进一步对生长和转移过程的基因进行PPI互作分析,提示Hedgehog signaling pathway、Hippo signaling pathway是虫草素抗三阴性乳腺癌的重要潜在作用通路。研究结果提示Hippo-Yap/TAZ信号通路和Hedgehog-Gli信号通路为潜在作用通路,为进一步理解虫草素治疗三阴性乳腺癌的分子机制奠定了基础。

关键词: 虫草素, 转录组, 三阴性乳腺癌, Hedgehog, Hippo

Abstract:

Previous studies have shown that cordycepin,the active ingredient of Cordyceps militaris,can effectively inhibit the growth and metastasis of triple-negative breast cancer xenografts tumor,however its mechanism is not clear yet,thus the purpose of this paper is to predict its mechanism. Taking samples of triple-negative breast cancer xenografts tumor,the molecular mechanism of cordycepin against breast cancer xenografts tumor was analyzed through transcriptome sequencing technology. The results showed that 584 differentially expressed genes were screened using P<0.05,log₂|FC|>1 as the standard,including 69 up-regulated genes and 515 down-regulated genes. KEGG analysis demonstrated that cordycepin had molecular regulation on breast cancer xenografts tumors including Hippo signaling pathway,Hedgehog signaling pathway,ECM-receptor interaction,TGF-beta signaling pathway and other cancer-related signaling pathways. GO analysis revealed that these differential genes were enriched in 16 biological processes,such as biological adhesion,growth,metabolic process,and locomotion,etc. It is suggested that cordycepin had significant regulatory effect on the growth and metastasis of breast cancer xenografts tumors. Further protein-protein interaction analysis of these genes in the growth and metastasis process indicated that Hedgehog signaling pathway and Hippo signaling pathway were important potential ones for cordycepin against triple-negative breast cancer. The research results suggest that the Hippo-Yap/TAZ signaling pathway and the Hedgehog-Gli signaling pathway are potential ones,laying the foundation for exploring the molecular mechanism of cordycepin in the treatment of triple-negative breast cancer.

Key words: cordycepin, RNA-Seq, triple negative breast cancer, Hedgehog, Hippo

黄海辰, 吴文雅, 戚梦, 薛帆正, 吴小平, 张君丽, 傅俊生. 虫草素抗三阴性乳腺癌的转录组学分析[J]. 生物技术通报, 2021, 37(11): 72-80.

HUANG Hai-chen, WU Wen-ya, QI Meng, XUE Fan-zheng, WU Xiao-ping, ZHANG Jun-li, FU Jun-sheng. RNA-Seq Analysis of Cordycepin Against Triple-negative Breast Cancer[J]. Biotechnology Bulletin, 2021, 37(11): 72-80.

图/表 15

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基因Gene	引物序列Primer sequence（5'-3'）
PDK4	F:5'-GAGGATTACTGACCGCCTCTTTAG-3' R:5'-TTCCGGGAATTGTCCATCAC-3'
CD24	F:5'-CCCACGCAGATTTATTCCAG-3' R:5'-GACTTCCAGACGCCATTTG-3'
MMP7	F:5'-AGCCAAACTCAAGGAGATGC-3' R:5'-GCCAATCATGATGTCAGCAG-3'
SHH	F:5'-AGGGCACCATTCTCATCAAC-3' R:5'-GGAGCGGTTAGGGCTACTCT-3'
GLI2	F:5'-GCCCTT CCTGAAAAGAAGAC-3' R:5'-CATTGGAGA AACAGGATTGG-3'
GADPH	F:5'-GAAGGACTCATGACCACAG-3' R:5'-CTTCACCACCTTCTTGATG-3'

基因Gene	引物序列Primer sequence（5'-3'）
PDK4	F:5'-GAGGATTACTGACCGCCTCTTTAG-3' R:5'-TTCCGGGAATTGTCCATCAC-3'
CD24	F:5'-CCCACGCAGATTTATTCCAG-3' R:5'-GACTTCCAGACGCCATTTG-3'
MMP7	F:5'-AGCCAAACTCAAGGAGATGC-3' R:5'-GCCAATCATGATGTCAGCAG-3'
SHH	F:5'-AGGGCACCATTCTCATCAAC-3' R:5'-GGAGCGGTTAGGGCTACTCT-3'
GLI2	F:5'-GCCCTT CCTGAAAAGAAGAC-3' R:5'-CATTGGAGA AACAGGATTGG-3'
GADPH	F:5'-GAAGGACTCATGACCACAG-3' R:5'-CTTCACCACCTTCTTGATG-3'

样品 Sample	浓度 Concentration/（ng·μL^-1）	体积 Volume /μL	总量 Total /μg	RIN值 RIN value	文库类型 Library type	结论 Conclusion
C0R1	295	35	10.33	6.8	RNAseq	A
C0R2	528	35	18.46	7.9	RNAseq	A
C0R3	761	35	26.67	9.3	RNAseq	A
C0N1	652	76	49.55	8.3	RNAseq	A
C0N2	695	76	52.78	9.3	RNAseq	A
C0N3	602	74	44.51	9.3	RNAseq	A

样品 Sample	浓度 Concentration/（ng·μL^-1）	体积 Volume /μL	总量 Total /μg	RIN值 RIN value	文库类型 Library type	结论 Conclusion
C0R1	295	35	10.33	6.8	RNAseq	A
C0R2	528	35	18.46	7.9	RNAseq	A
C0R3	761	35	26.67	9.3	RNAseq	A
C0N1	652	76	49.55	8.3	RNAseq	A
C0N2	695	76	52.78	9.3	RNAseq	A
C0N3	602	74	44.51	9.3	RNAseq	A

样品名 Sample	总reads数量 Number of total reads	比对上rRNA的 reads 数及占总数的比例 Mapped reads（ratio in total）	未比对上rRNA的 reads 数及占总数的比例 Unmapped reads（ratio in total）
CON1	53195320	410412（0.77%）	52784908（99.23%）
CON2	69510386	493598（0.71%）	69016788（99.29%）
CON3	65958874	550350（0.83%）	65408524（99.17%）
COR1	90497008	812122（0.90%）	89684886（99.10%）
COR2	78992736	387314（0.49%）	78605422（99.51%）
COR3	56738684	310270（0.55%）	56428414（99.45%）