Long Noncoding RNA Metastasis-Associated Lung Adenocarcinoma Transcript 1 (MALAT1) Promotes Renal Cell Carcinoma Progression via Sponging miRNA-429

Abstract

Background: It is well known that long noncoding RNA (lncRNA) metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) is closely correlated with the tumorigenesis of multiple cancers, including renal cell carcinoma (RCC). However, the potential functional mechanism is still elusive.

Material/Methods: In our present research, quantitative real-time polymerase chain reaction (qRT-PCR) was performed for the measurement of MALAT1 and miR-429. CCK-8 assay and Transwell assay were performed for the proliferation, migration, and invasion abilities of RCC cells. Dual-luciferase reporter assay was performed to validate the interaction within MALAT1 and miR-429.

Results: Data found that MALAT1 was overexpressed in RCC clinical samples and cell lines. Moreover, loss-of-functional experiments showed that MALAT1 knockdown suppress the proliferation, migration, and invasion abilities of RCC cells. RT-PCR showed that miR-429 expression was downregulated in RCC cell lines, which was negatively correlated with that of MALAT1. Bioinformatics analysis suggested that miR-429 had complementary binding sequences with MALAT1, which was confirmed by dual-luciferase reporter assay.

Conclusions: In summary, our results concluded that MALAT1 functioned as an oncogene in RCC by sponging miR-429, acting as its competing endogenous RNA (ceRNA).

Keywords: Carcinoma, Renal Cell; Nephrology; RNA, Long Noncoding.

Figure 1

MALAT1 was upregulated in renal cell carcinoma (RCC) patient tissues and cell lines. (A) Expression of MALAT1 was significantly higher in RCC tissues than normal tissues. (B) Expression level of MALAT1 was significantly higher in RCC cell lines, 786-O and ACHN, than HEK 293-T cells. Data was presented as mean ±SD, ** P<0.01, *** P<0.001 calculated with Student’s t-test.

Figure 2

Downregulation of MALAT1 inhibited proliferation, migration, and invasion of renal cell carcinoma (RCC) cells. (A) Expression level of MALAT1 is significantly decreased after MALAT1 siRNA transfection. (B) Proliferation of RCC cells was determined at indicated time point after transfection using CCK-8 assay. (C) Migration ability of RCC cells was determined after transfection using Transwell assay. Random fields are shown below. (D) Invasion ability of RCC cells was determined after transfection using Transwell assay. Random fields are shown below. Data was presented as mean ±SD, * P<0.05, ** P<0.01, calculated with Student’s t-test.

Figure 3

MiR-429 is a direct target of MALAT1. (A) MiRNA expression profiles in renal cell carcinoma (RCC) tissue was detected by miRNA microarray. (B) Expression of miR-429 was significantly higher in RCC tissues than normal tissues. (C) Expression level of miR-429 was significantly higher in RCC cell lines, 786-O and ACHN, than HEK 293-T cells. (D) Pearson’s correlation analyzed that expression of miR-429 was negative related to expression of MALAT1 (R2=0.766). (E) A presumptive complementary alignment between MALAT1 and miR-429. (F) RCC 786-O cells were co-transfected with WT/Mut- MALAT1 vectors and miR-429/NC. Dual-luciferase reporter assay indicated that there was certain possibility of 3′-UTR of MALAT1 (WT) and miR-429 sharing a complementary binding site (firefly luciferase activity in overexpressed miR-429 786-O cells was decreased by 61% comparing to 786-O cells transfected with NC). Data was presented as mean ±SD, ** P<0.01, calculated with Student’s t-test.

Figure 4

Functions of MALAT1 and miR-429 on RCC 786-O cells. (A) Expression of miR-429 in RCC 786-O cells after transfections. (B) Cell viability of RCC 786-O cells determined by CCK-8 assay. (C, D) Cell migration and invasion abilities of RCC 786-O cells were determined by Transwell assays. Data was presented as mean ±SD, ** P<0.01, calculated with Student’s t-test.