COX19 Is a New Target of MACC1 and Promotes Colorectal Cancer Progression by Regulating Copper Transport in Mitochondria.

BACKGROUND: Recent studies have demonstrated that copper (Cu) plays an important role in the progression of tumor diseases. Metastasis associated with colon cancer protein 1 (MACC1) promotes the transcription and expression of various tumor-related genes. Cytochrome c oxidase (COX) 19, present in the cytoplasm and intermembrane space of mitochondria, may transport Cu within the mitochondria. However, the mechanism through which MACC1 regulates the Cu homeostasis mediated by COX19 remains unclear. OBJECTIVES: The aim of this study was to elucidate the mechanism through which MACC1 initiates the transcription and expression of COX19, and promotes malignant behavior in tumor cells. METHODS: Immunohistochemistry, western blotting, and real-time polymerase chain reaction (PCR) analyses were conducted to analyze the expression of MACC1 and COX19 proteins and genes in tumor and normal tissues. RNA-chromatin immunoprecipitation was used to detect the transcriptional initiation of COX19 by MACC1. The effects of MACC1 and COX19 on mitochondrial activity were determined using an ATP assay kit and Cytochrome c Oxidase Assay Kit. A Cell Counting Kit-8 kit was used to detect the effect of high-dose Cu or overexpression of MACC1 and COX19 on tumor cell proliferation. A xenograft mouse model was used to analyze the effect of the COX19 overexpression on the malignant behavior of the tumors. RESULTS: Cu enhanced the proliferation, invasion, and migration and inhibited apoptosis of SW480 cells. MACC1 was highly expressed in colorectal cancer tissues and activated the expression of COX19 by binding to its promoter region of COX19. The overexpression of COX19 increased mitochondrial Cu content and enhanced the activity of mitochondrial COX and ATP content, and inhibited apoptosis, promoted tumor growth of mice. CONCLUSIONS: Our results indicate that COX19 functions as a target gene of MACC1 and regulates mitochondrial activity and promotes the progression of colorectal cancer. MACC1/COX19 may provide a novel therapeutic target for colorectal cancer.

Expression and copper binding studies of a Plasmodium falciparum protein with Cox19 copper binding motifs.

Copper can exist in an oxidized and a reduced form, which enables the metal to play essential roles as a catalytic co-factor in redox reactions in many organisms. Copper confers redox activity to the terminal electron transport chain cytochrome c oxidase protein. Cytochrome c oxidase in yeast obtains copper for the CuB site in the Cox1 subunit from Cox11 in association with Cox19. When copper is chelated in growth medium, Plasmodium falciparum parasite development in infected red blood cells is inhibited and excess copper is toxic for the parasite. The gene of a 26 kDa Plasmodium falciparum PfCox19 protein with two Cx9C Cox19 copper binding motifs, was cloned and expressed as a 66 kDa fusion protein with maltose binding protein and affinity purified (rMBP-PfCox19). rMBP-PfCox19 bound copper measured by: a bicinchoninic acid release assay; an in vivo bacterial host growth inhibition assay; ascorbate oxidation inhibition and differential scanning fluorimetry. The native protein was detected by antibodies raised against rMBP-PfCox19. PfCox19 binds copper and is predicted to associate with PfCox11 in the insertion of copper into the CuB site of Plasmodium cytochrome c oxidase. Characterisation of the proteins involved in Plasmodium spp. copper metabolism will help us understand the role of cytochrome c oxidase and this essential metal in Plasmodium homeostasis.

The mitochondrial copper chaperone COX19 influences copper and iron homeostasis in arabidopsis.

KEY MESSAGE: The mitochondrial metallochaperone COX19 influences iron and copper responses highlighting a role of mitochondria in modulating metal homeostasis in Arabidopsis. The mitochondrial copper chaperone COX19 participates in the biogenesis of cytochrome c oxidase (COX) in yeast and humans. In this work, we studied the function of COX19 in Arabidopsis thaliana, using plants with either decreased or increased COX19 levels. A fusion of COX19 to the red fluorescent protein localized to mitochondria in vivo, suggesting that Arabidopsis COX19 is a mitochondrial protein. Silencing of COX19 using an artificial miRNA did not cause changes in COX activity levels or respiration in plants grown under standard conditions. These amiCOX19 plants, however, showed decreased expression of the low-copper responsive miRNA gene MIR398b and an induction of the miR398 target CSD1 relative to wild-type plants. Plants with increased COX19 levels, instead, showed induction of MIR398b and other low-copper responsive genes. In addition, global transcriptional changes in rosettes of amiCOX19 plants resembled those observed under iron deficiency. Phenotypic analysis indicated that the roots of amiCOX19 plants show altered growth responses to copper excess and iron deficiency. COX activity levels and COX-dependent respiration were lower in amiCOX19 plants than in wild-type plants under iron deficiency conditions, suggesting that COX19 function is particularly important for COX assembly under iron deficiency. The results indicate that the mitochondrial copper chaperone COX19 has a role in regulating copper and iron homeostasis and responses in plants.

The mitochondrial intermembrane space oxireductase Mia40 funnels the oxidative folding pathway of the cytochrome c oxidase assembly protein Cox19.

Mia40-catalyzed disulfide formation drives the import of many proteins into the mitochondria. Here we characterize the oxidative folding of Cox19, a twin CX9C Mia40 substrate. Cox19 oxidation is extremely slow, explaining the persistence of import-competent reduced species in the cytosol. Mia40 accelerates Cox19 folding through the specific recognition of the third Cys in the second helical CX9C motif and the subsequent oxidation of the inner disulfide bond. This renders a native-like intermediate that oxidizes in a slow uncatalyzed reaction into native Cox19. The same intermediate dominates the pathway in the absence of Mia40, and chemical induction of an α-helical structure by trifluoroethanol suffices to accelerate productive folding and mimic the Mia40 folding template mechanism. The Mia40 role is to funnel a rough folding landscape, skipping the accumulation of kinetic traps, providing a rationale for the promiscuity of Mia40.

MicroRNA-21 regulates non-small cell lung cancer cell proliferation by affecting cell apoptosis via COX-19.

AIMS: This study is to investigate the regulatory effect of microRNA-21 (miR-21) on bone metastasis of non-small cell lung cancer (NSCLC). METHODS: In this study, 18 patients were diagnosed with vertebral column metastasis of NSCLC. MiR-21 or small interfering RNAs were transfected into H2170 cells using Lipofectamine 2000. Real-time PCR was performed to detect miR-21 expression. Western blotting was used to measure the expression of COX-19 protein. Enzymatic activity tests were performed to measure the activity of cytochrome C oxidase. Flow cytometry was used to monitor changes in cell apoptotic rate. MTT assay was used to determine the capability of cell proliferation. RESULTS: Bone metastasis of NSCLC enhanced the levels of miR-21 in NSCLC patients. Proliferation capability of cells with high expression of miR-21 was greater than that of cells with the inhibition of miR-21 expression. High expression of miR-21 promoted cell proliferation by inhibiting cell apoptosis. COX-19 was a key factor in the inhibition of apoptosis by miR-21. Inhibition of COX-19 expression reduced cell proliferation by enhancing cell apoptosis. CONCLUSIONS: This study demonstrates that inhibition of miRNA-21 suppresses NSCLC cell proliferation by promoting cell apoptosis via the decrease of COX-19 expression.