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Fig. 5 | Molecular Cancer

Fig. 5

From: LncRNA TROJAN promotes proliferation and resistance to CDK4/6 inhibitor via CDK2 transcriptional activation in ER+ breast cancer

Fig. 5

TROJAN promoted the transcription of CDK2 via NKRF. a Schematic diagram of the screening of the eight potential target genes (CCNA2, MCM7, CDC45, CDK2, MCM3, PCNA, BUB1 and MCM6). First, 694 genes down regulated by TROJAN knockdown (according to transcriptome analysis of TROJAN knockdown MCF7 cells) were used for screening. After the overlapping with pearson correlation analysis of genes that were positively correlated with TROJAN (R > 0.2, P < 0.05) in FUSCC and TCGA cohort, as well as genes from the KEGG cell cycle pathways, the potential eight target genes were identified. b qRT-PCR detecting the expression of CDK2 during TROJAN or NKRF knockdown. n = 3 independent experiments. Unpaired t test was used. c qRT-PCR detecting the expression of CDK2 in MCF7 cells with TROJAN or TROJAN/NKRF knockdown. n = 3 independent experiments. Unpaired t test was used. d Luciferase reporter assay detecting the activity of the CDK2 promoter during TROJAN or NKRF knockdown. n = 3 independent experiments. Unpaired t test was used. e Western blot images of p-CDK2 Thr160 and total CDK2 in MCF7 cells expressing TROJAN shRNAs. n = 3 independent experiments. f Western blot images of CDK2 in MCF7 cells ectopically expressing TROJAN. n = 3 independent experiments. g Immunohistochemical detection of CDK2 in a mammary fat pad xenograft model generated by injecting MCF7 cells with TROJAN knockdown. The H-score is shown. h RELA ChIP-Seq signals in lymphocyte at CDK2 nearby genomic location. i qRT-PCR analysis of NKRF at the CDK2 promoter after ChIP assays in MCF7 cells expressing control or TROJAN shRNA. n = 3 independent experiments. Unpaired t test was used. j Western blot images of MCF7 and PDR cells treated for 24 and 48 h with 0.1 μM PD and blotted for phospho-CDK2 (p-CDK2) Thr160, total CDK2, phospho-RB1 (p-RB1) S807/811, and total RB1. n = 3 independent experiments. k Western blot images of MCF7, PDR and PDR with TROJAN knockdown cells treated for 24 h with 0.1 μM PD and blotted for p-CDK2 Thr160, total CDK2, p-RB1 S807/811, and total RB1. n = 3 independent experiments. l In vitro growth curves of MCF7 cells with ± CDK2 knockdown, ± TROJAN knockdown and ± CDK2 over expression. Two-way ANOVA analysis was used. **p < 0.01, ***p < 0.001 and NS: not significant

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