Downregulation of mitochondrial complex I induces ROS production in colorectal cancer subtypes that differently controls migration

Abstract

Background Colorectal cancer (CRC) can be classified into four molecular subtypes (CMS) among which CMS1 is associated with the best prognosis, while CMS4, the mesenchymal subtype, has the worst outcome. Although mitochondria are considered to be hubs of numerous signaling pathways, the study of mitochondrial metabolism has been neglected for many years. Mitochondrial Complex I (CI) plays a dual role, both in energy and reactive oxygen species (ROS) production. However, the possible contribution of CI to tumorigenesis in cancer remains unclear. The purpose of this study was to investigate the CI under the prism of the CMS classification of CRC in ex vivo models. Methods Biochemical dosages, bioenergetics analysis and western-blot were used to characterize CI expression, function and redox balance in LoVo and MDST8 cell lines, belonging to CMS1 and CMS4 subgroups, respectively. Cell proliferation and migration were assessed by xCELLigence technology. Overproduction or scavenging of mitochondrial ROS (mtROS) were performed to analyze the effect of mtROS on proliferation, migration, and mesenchymal markers. Focal adhesion kinase (FAK) and its activation were analyzed by immunofluorescence. We assessed the distribution of two CI scores in CRC cohorts according to CMS classification and their relevance for patient survival. Results We found that CI is downregulated in CMS4 cells and is associated with elevated mtROS. We establish for the first time that in these migrating cells, mtROS production is maintained at optimal levels not only through changes in CI activity but also by inactivation/acetylation of superoxide dismutase 2 (SOD2), a major mitochondrial antioxidant enzyme. We show that promoting or scavenging mtROS both mitigate CMS4 cells’ migration. Our results also point to a mtROS-mediated focal adhesion kinase (FAK) activation, which likely sustains their migratory phenotype. Using cohorts of CRC patients, we document that the expression of CI is downregulated in the CMS4 subgroup, and that low CI expression is associated with poor prognosis. Patients’ datasets reveal an inverse correlation between CI and the epithelial-mesenchymal transition (EMT) pathway. Conclusion We showed that inhibition of CI contributes to heighten mtROS, which likely foster MDST8 migration and might account for the specific EMT signature of CMS4 tumors. These data reveal a novel role of mitochondrial CI in CRC, with biological consequences that may be targeted with anti- or pro-oxidant drugs in clinical practice.

Unlocking Mitochondrial Dysfunction-Associated Senescence (MiDAS) with NAD+– a Boolean Model of Mitochondrial Dynamics and Cell Cycle Control

“…While PGC1α can induce SIRT3 [16], its activity depends on NAD + , which requires a functioning ETC. Our model assumes that the combination of low ΔΨ M and mitochondrial ROS – which damages ETC components and mDNA – hinder the regeneration of the mitochondrial NAD + pool [96,97].…”

Clinical Challenges of Consensus Molecular Subtype CMS4 Colon Cancer in the Era of Precision Medicine

“…Another effector is the integrin linked kinase (ILK), which we found to be overexpressed in CMS4 colon cancer and to act as a relay downstream from PrP C to activate YAP/TAZ signaling ( 39 ). Among other biological hallmarks of this subtype shown to contribute to CMS4 traits feature elevated AKT3 ( 42 ) and KIT ( 43 ), reduced mitochondrial respiratory complex I activity ( 44 )—possibly resulting from KIT overexpression ( 43 )—or modified alternative splicing landscape ( 45 ). Relevant signatures further include β3 integrin activation ( 1 ), which could in turn foster ILK activation ( 46 ), hypoxia, and DNA damage ( 47 ), potentially accounting for elevated PrP C expression (reviewed in ref.…”

“…Overexpressed/overactivated genes, proteins, or pathways are highlighted in red, and underexpressed/underactivated genes, proteins, or pathways are highlighted in green. Established links are indicated with a plain line: reduced atypical PKC expression and associated increased hyaluronic acid (HA) synthesis ( 50 ), AKT-dependent activation of NOTCH ( 49 ), PrP C -dependent control on soluble TGFβ levels, ILK and YAP/TAZ activation ( 35, 39 ), reduced oxidative phosphorylation and increased ZEB1 levels ( 44 ), ZEB1-dependent negative regulation of the ESPR1 splicing regulator expression ( 45 ), phosphorylation-dependent STAT3 activation and downstream silencing of miR200, itself promoting enhanced expression of ZEB2 ( 21 ), decreased levels of cyclin A2 (CCNA2), fostering DNA damage ( 51 ). Dashed lines represent potential links: β3 integrin-dependent activation of ILK ( 46 ) and KIT-dependent reduction of oxidative phosphorylation ( 43 ).…”

Pitfalls of Mitochondrial Redox Signaling Research

“…Thus, mt redox signals are implicated in aging [ 116 , 117 , 118 ], as well as in the development of cancer and metastases [ 119 , 120 , 121 , 122 , 123 ]. Cytosolic redox signals/gene regulations are hypothetically involved in type 2 diabetes changes in pancreatic β-cells.…”

“…Also, a few examples are listed for cytosolic redox signals, such as those initiated via NADPH oxidases. Thus, mt redox signals are implicated in aging [116][117][118], as well as in the development of cancer and metastases [119][120][121][122][123]. Cytosolic redox signals/gene regulations are hypothetically involved in type 2 diabetes changes in pancreatic β-cells.…”