The miR-27a/FOXJ3 Axis Dysregulates Mitochondrial Homeostasis in Colorectal Cancer Cells

Abstract

miR-27a plays a driver role in rewiring tumor cell metabolism. We searched for new miR-27a targets that could affect mitochondria and identified FOXJ3, an apical factor of mitochondrial biogenesis. We analyzed FOXJ3 levels in an in vitro cell model system that was genetically modified for miR-27a expression and validated it as an miR-27a target. We showed that the miR-27a/FOXJ3 axis down-modulates mitochondrial biogenesis and other key members of the pathway, implying multiple levels of control. As assessed by specific markers, the miR-27a/FOXJ3 axis also dysregulates mitochondrial dynamics, resulting in fewer, short, and punctate organelles. Consistently, in high miR-27a-/low FOXJ3-expressing cells, mitochondria are functionally characterized by lower superoxide production, respiration capacity, and membrane potential, as evaluated by OCR assays and confocal microscopy. The analysis of a mouse xenograft model confirmed FOXJ3 as a target and suggested that the miR-27a/FOXJ3 axis affects mitochondrial abundance in vivo. A survey of the TCGA-COADREAD dataset supported the inverse relationship of FOXJ3 with miR-27a and reinforced cellular component organization or biogenesis as the most affected pathway. The miR-27a/FOXJ3 axis acts as a central hub in regulating mitochondrial homeostasis. Its discovery paves the way for new therapeutic strategies aimed at restraining tumor growth by targeting mitochondrial activities.

Transcriptomic Analysis of Colorectal Cancer Cells Treated with Oil Production Waste Products (OPWPs) Reveals Enrichment of Pathways of Mitochondrial Functionality

“…To this purpose, we assessed the levels of the major proteins involved in these processes. FOXJ3 controls the very first step of the pathway by modulating PGC-1α either directly and through MEF2A and MEF2C, two members of the MEF2 family [ 27 , 35 , 36 ] ( , accessed on 20 October 2022). PGC-1α plays, in turn, a pivotal role in many mitochondrial activities, stimulating the expression of NRF1 and NRF2, two transcription factors which cooperate to activate nuclear and mitochondrial genes required for biogenesis and respiratory functions [ 17 , 37 ].…”

“…PGC-1α, in addition, stimulates the binding and activity of factors required for mitochondrial transcription factor A (TFAM) also involved in the replication and maintenance of mitochondrial DNA [ 20 , 38 ]. All together, these factors form a mutually self-regulatory and cross-regulatory network able to direct OXPHOS in muscle and in CRC cells [ 27 , 39 ]. In HCT116 cells, all treatments increased FOXJ3 protein, with TE and SE showing the strongest effect.…”

“…Next, we sought to unveil the mechanisms underlying the above-described effects of HTyr and OPWP extracts on mitochondrial structure and function. Data from the literature indicate that PPARGC1 is the master gene that orchestrates a myriad of mitochondrial functions such as OXPHOS, fatty acids synthesis and oxidation, tricarboxylic acid cycle, in addition to the biogenesis already described [ 20 , 21 , 27 ]. Among the transcription factors regulated by this coactivator, PPARγ is involved in lipidic and energetic control [ 23 , 40 ].…”

“…PPARGC1A is also likely to be regulated by PPARγ as PPREs have been reported in its promoter region ( , accessed on 20 October 2022). The resulting elevated PGC-1α protein, in turn, acts as a coactivator to the transcription factors NRF1 and NRF2 on the promoter of TFAM, stimulating its expression [ 27 , 37 ]. Further activation can be obtained by the direct binding of FOXJ3 to TFAM promoter as well as to that of TOMM20 ( , accessed on 20 October 2022, see Supplementary Figure S2 ).…”

“…Protein extracts were analyzed as previously reported [ 27 ]. Briefly, the cells were lysed by an ice-cold lysis buffer containing 25 mM Tris-HCl, pH 7.5, 150 mM NaCl, 2 mM EDTA, 1% Triton X-100, 1% sodium deoxycholate, 0.1% SDS, a cocktail of protease and phosphatase inhibitors (Roche, Basel, Switzerland).…”