Effect of MALAT1 in the crosstalk between nucleus and mitochondria on mitochondrial reprogramming in hepatocellular carcinoma cells.

Abstract

e14711 Background: Mitochondria-nuclear crosstalk is a bidirectional pathway of communication between mitochondria and nucleus that influences many cellular and organismal activities. This crosstalk can regulate several oncogenic pathways involved in tumorigenesis. MALAT1 (metastasis-associated lung adenocarcinoma transcript 1), a nucleus-encoded lncRNA, dysregulated in multiple human malignancies is recently found to drive mitochondria dysfunction in Mitochondria-nuclear crosstalk. Methods: RNA sequencing, “RNA reverse transcription-associated trap sequencing” (RAT-seq), RNA immunoprecipitation(RIP), fluorescence in situ hybridization (FISH), were performed to detect the position of the MALAT1, and its interaction protein and DNAs in HepG2 cell line. After silencing MALAT1 by shRNAs, Seahorse, ATP production, lysotracker staining, Western blot, and electronic microscope were used to measure metabolism, ROS amount, mitophagy, apoptosis and mitochondrial morphology of the silencing cell lines. Results: By combining mitochondrial RNA-Seq with FISH, it is surprised to discover that MALAT1 was enriched in the mitochondria of HepG2 cells. Using RAT-seq approach, MALAT1 was found to utilized it 3’-fragment to interact with multiple loci of mitochondrial DNA( D-loop, COX2, ND3, and CYTB ). The RIP and affinity RNA pulldown assays suggested that the RNA-binding protein HuR mediated the transportation of MALAT1 to the mitochondria. Also, mitochondria transmembrane protein mitochondrial carrier 2(MTCH2) is found to interacted MALAT1, suggesting that MALAT1 may through the MTCH2 to get into the inside of the mitochondria. Knockdown of MALAT1 induced multiple abnormalities in mitochondrial functions, including low OXPHOS, low ATP production, reduced mitophagy, declined mtDNA copy number, and activation of the mitochondrial apoptosis pathway. Conclusions: Together, this study greatly expands our knowledge of the nucleus-encoded lncRNA MALAT1 driving the mitochondria dysfunction. Our study establishes MALAT1 as a regulator through interacting with MT-DNA, HuR, MTCH2 protein, revealing a new regulatory mechanism of mitochondria. Many novel nucleus-encoded RNAs existing in mitochondria were identified at the first time, suggesting novel biological functions of lncRNAs, laying the foundation for further clarifying their roles.