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, 52 (2), 139-144

SETDB1 Regulates SMAD7 Expression for Breast Cancer Metastasis

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SETDB1 Regulates SMAD7 Expression for Breast Cancer Metastasis

Tae Young Ryu et al. BMB Rep.

Abstract

Breast cancer (BRC) is the most invasive cancer in women. Although the survival rate of BRC is gradually increasing due to improved screening systems, development of novel therapeutic targets for inhibition of BRC proliferation, metastasis and recurrence have been constantly needed. Thus, in this study, we identified overexpression of SETDB1 (SET Domain Bifurcated 1), a histone methyltransferase, in RNA-seq data of BRC derived from TCGA portal. In Gene Ontology (GO) analysis, cell migration-related GO terms were enriched, and we confirmed down-regulation of cell migration/invasion and alteration of EMT /MET markers after knockdown of SETDB1. Moreover, gene network analysis showed that SMAD7 expression is regulated by SETDB1 levels, indicating that up-regulation of SMAD7 by SETDB1 knockdown inhibited BRC metastasis. Therefore, development of SETDB1 inhibitors and functional studies may help develop more effective clinical guidelines for BRC treatment. [BMB Reports 2019; 52(2): 139-144].

Conflict of interest statement

CONFLICTS OF INTEREST

The authors have no conflicting interests.

Figures

Fig. 1
Fig. 1
RNA-seq analysis of SETDB1 knockdown MB-231 cells. (A) DAVID-based gene ontology analysis of RNA-seq results from the siSETDB1 and siCont groups. Enriched terms are shown. Raw transcriptome sequence data (RNA-seq) were deposited in the Short Read Archive (SRA) of NCBI under the accession number PRJNA506342. (B) GO pathway term enrichment networks. GO pathway term networks in the SETDB1 knockdown and control groups were functionally grouped by ClueGO. (C) KEGG pathway analysis of RNA-seq results from the siSETDB1 and siCont groups. (D) RNA-seq results from the siSETDB1 and siCont groups. Up-regulation of epithelial cell markers (CDH1 and Claudin 1) and down-regulation of mesenchymal cell markers (CDH2 and Vimentin).
Fig. 2
Fig. 2
SETDB1 knockdown changed EMT/MET markers. (A, B) Migration and invasion assays after SETDB1 knockdown. Cell migration and invasion assays were performed after 36 h. Migrated/invaded cells were stained with crystal violet (left); P values were calculated using Student’s t-tests (***P < 0.001, *P < 0.05) (right). (C) Wound healing assay. After 24 h of SETDB1 knockdown, scratch assays were performed. After another 24 h, wound closure was measured. (D, E) RT-PCR (D) and qRT-PCR (E) analysis of EMT markers (CDH1, CDH2, Claudin 1, and Vimentin). Actin (ACTB) was used as an internal control.
Fig. 3
Fig. 3
Up-regulation of SMAD7 by SETDB1 knockdown. (A) Heat map of gene expression related to SETDB1 knockdown, sorted by fold change in the siCont/siSETDB1 FPKM value. In the heat map, red indicates up-regulation, while green indicates down-regulation. (B) Gene-to-gene networks of SETDB1 and SMAD7. Up- and down-regulated genes are indicated in red and green, respectively. Each line and arrow represents functional and physical interactions between the genes and the direction of regulation reported in the literature. (C, D) Up-regulation of SMAD7 after the knockdown of SETDB1. RT-PCR (C) and qRT-PCR analysis (D), P values were calculated using Student’s t-tests (**P < 0.01).
Fig. 4
Fig. 4
Down-regulation of SMAD7 recovered SETDB1-induced migration and invasion. (A) Wound healing assay. After 24 h of SMAD7 knockdown, scratch assays were performed. After another 48 h, wound closure was measured. (B) qRT-PCR analysis of EMT markers (CDH1, CDH2, Claudin 1, and Vimentin). P values were calculated using Student’s t-tests (***P < 0.001, **P < 0.01, *P < 0.05). (C) Wound healing assay. After treatment with siCont, siSETDB1, siSMAD7 and siSETDB1/siSMAD7 for 24 h, wound closure was measured after another 24 h. (D) qRT-PCR analysis of EMT markers (CDH1 and Claudin 1) after treatment with siCont, siSETDB1, siSMAD7 and siSETDB1/siSMAD7 for 48 h. (E) Schematic summary of the effects of SETDB1 on BRC metastasis.

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