Discussion
Our results demonstrated the mechanism by which the LINC00504-miR-140-5p-VEGFA axis participates in breast cancer cell proliferation and invasion and may lead to new lncRNA-based diagnostic or therapeutic strategies for breast cancer.
Methods
qPCR assay was used to detect the expression of LINC00504 in tissue and cell lines. The online database and chromatin immunoprecipitation assay (ChIP) were employed to confirm the transcription factor of LINC00504. Cell function assays including cell proliferation, migration and invasion were designed to detect the function of LINC00504 in vitro and in vivo. Luciferase reporter assay and RNA immunoprecipitation (RIP) assay were used to confirm the relationship between LINC00504 and miR-140-5p. And Western blot assay was employed for testing the key protein.
Results
We found that LINC00504 is upregulated in breast cancer. In addition, we found that the transcription factor regulatory factor X5 (RFX5) can strongly bind to the LINC00504 promoter region and subsequently increase its transcriptional activity. We also found that the manipulation of RFX5 expression can significantly affect LINC00504 expression, which suggested that RFX5 can transcriptionally activate LINC00504 in breast cancer (BC). Knockdown of LINC00504 inhibits cell proliferation, migration and invasion in vitro and in vivo. We further found that LINCOO504 inhibits miR-140-5p, which decreases the levels of VEGFA. The further results showed that miR-140-5p was one of the target gene of LINC00504. The WB assay demonstrated that the E-cadherin was increased and Vimentin was decreased when knocking down of LINC00504 and they can be rescued while adding the inhibitors of miR-140-5p.