MIEF1/2 orchestrate mitochondrial dynamics through direct engagement with both the fission and fusion machineries.

BACKGROUND: Mitochondrial dynamics is the result of a dynamic balance between fusion and fission events, which are driven via a set of mitochondria-shaping proteins. These proteins are generally considered to be binary components of either the fission or fusion machinery, but potential crosstalk between the fission and fusion machineries remains less explored. In the present work, we analyzed the roles of mitochondrial elongation factors 1 and 2 (MIEF1/2), core components of the fission machinery in mammals. RESULTS: We show that MIEFs (MIEF1/2), besides their action in the fission machinery, regulate mitochondrial fusion through direct interaction with the fusion proteins Mfn1 and Mfn2, suggesting that MIEFs participate in not only fission but also fusion. Elevated levels of MIEFs enhance mitochondrial fusion in an Mfn1/2- and OPA1-dependent but Drp1-independent manner. Moreover, mitochondrial localization and self-association of MIEFs are crucial for their fusion-promoting ability. In addition, we show that MIEF1/2 can competitively decrease the interaction of hFis1 with Mfn1 and Mfn2, alleviating hFis1-induced mitochondrial fragmentation and contributing to mitochondrial fusion. CONCLUSIONS: Our study suggests that MIEFs serve as a central hub that interacts with and regulates both the fission and fusion machineries, which uncovers a novel mechanism for balancing these opposing forces of mitochondrial dynamics in mammals.

The phosphorylation status of Ser-637 in dynamin-related protein 1 (Drp1) does not determine Drp1 recruitment to mitochondria.

Recruitment of the GTPase dynamin-related protein 1 (Drp1) to mitochondria is a central step required for mitochondrial fission. Reversible Drp1 phosphorylation has been implicated in the regulation of this process, but whether Drp1 phosphorylation at Ser-637 determines its subcellular localization and fission activity remains to be fully elucidated. Here, using HEK 293T cells and immunofluorescence, immunoblotting, RNAi, subcellular fractionation, co-immunoprecipitation assays, and CRISPR/Cas9 genome editing, we show that Drp1 phosphorylated at Ser-637 (Drp1pS637) resides both in the cytosol and on mitochondria. We found that the receptors mitochondrial fission factor (Mff) and mitochondrial elongation factor 1/2 (MIEF1/2) interact with and recruit Drp1pS637 to mitochondria and that elevated Mff or MIEF levels promote Drp1pS637 accumulation on mitochondria. We also noted that protein kinase A (PKA), which mediates phosphorylation of Drp1 on Ser-637, is partially present on mitochondria and interacts with both MIEFs and Mff. PKA knockdown did not affect the Drp1-Mff interaction, but slightly enhanced the interaction between Drp1 and MIEFs. In Drp1-deficient HEK 293T cells, both phosphomimetic Drp1-S637D and phospho-deficient Drp1-S637A variants, like wild-type Drp1, located to the cytosol and to mitochondria and rescued a Drp1 deficiency-induced mitochondrial hyperfusion phenotype. However, Drp1-S637D was less efficient than Drp1-WT and Drp1-S637A in inducing mitochondrial fission. In conclusion, the Ser-637 phosphorylation status in Drp1 is not a determinant that controls Drp1 recruitment to mitochondria.

Mief1 augments thyroid cell dysfunction and apoptosis through inhibiting AMPK-PTEN signaling pathway.

Objective: Inflammation-mediated thyroid cell dysfunction and apoptosis increases the like-hood of hypothyroidism.Aim: Our aim in the present study is to explore the role of mitochondrial elongation factor 1 (Mief1) in thyroid cell dysfunction induced by TNFα.Materials and methods: Different doses of TNFα were used to incubate with thyroid cells in vitro. The survival rate, apoptotic index and proliferation capacity of thyroid cells were measured. Cellular energy metabolism and endoplasmic reticulum function related to protein synthesis were detected.Results: In response to TNFα treatment, the levels of Mief1 were increased, coinciding with a drop in the viability of thyroid cells in vitro. Loss of Mief1 attenuates TNFα-induced cell death through reducing the ratio of cell apoptosis. Further, we found that Mief1 deletion reversed cell energy metabolism and this effect was attributable to mitochondrial protection. Mief1 knockdown sustained mitochondrial membrane potential and reduced mitochondrial ROS overproduction. In addition, Mief1 knockdown also reduced endoplasmic reticulum stress, as evidenced by decreased levels of Chop and Caspase-12. Finally, our data verified that TNFα treatment inhibited the activity of AMPK-PTEN pathway whereas Mief1 deletion reversed the activity of AMPK and thus promoted the upregulation of PTEN. However, inhibition of AMPK-PTEN pathways could abolish the beneficial effects exerted by Mief1 deletion on thyroid cells damage and dysfunction.Conclusions: Altogether, our data indicate that immune abnormality-mediated thyroid cell dysfunction and death are alleviated by Mief1 deletion possible driven through reversing the activity of AMPK-PTEN pathways.

Melatonin attenuates inflammation-related venous endothelial cells apoptosis through modulating the MST1-MIEF1 pathway.

Chronic venous disease (CVD) is a prevalent and potentially debilitating condition that affects millions of individuals. An excessive endothelial inflammatory response is reportedly involved in the development of CVD. In this study, we explored the effect and mechanism of melatonin on venous endothelial damage induced by tumor necrosis factor α (TNF-α). Our data demonstrated that inflammation injury triggered mitochondrial dysfunction, activated reactive oxygen species-related oxidative damage, inhibited mitochondrial potential and ultimately initiated caspase-involved cellular death. Interestingly, melatonin preserved inflammation-attacked mitochondrial performance and thus increased cell survival under TNF-α. Cellular experiments illustrated that inflammation injury promoted the levels of mammalian sterile 20-like kinase 1 (MST1) and mitochondrial elongation factor 1 (MIEF1); active MST1-MIEF1 pathway disturbed mitochondria-related energy production, leading to mitochondria-induced cell damage. Interestingly, melatonin effectively suppressed MST1-MIEF1 axis and thus improved cell survival ratio under TNF-α-mediated inflammation injury. Reactivation of MST1-MIEF1 pathway attenuated melatonin-related endothelial protective actions. Herein, our results illuminate that melatonin is an effective approach to attenuate inflammation-related venous endothelial cell damage through handling the MST1-MIEF1 signaling pathway.

Matrine promotes apoptosis in SW480 colorectal cancer cells via elevating MIEF1-related mitochondrial division in a manner dependent on LATS2-Hippo pathway.

Matrine, an alkaloid compound isolated from Sophora flavescens Ait, has been shown to exert cancer-killing actions in a variety of tumors; however, its anticancer mechanism in colorectal cancer (CRC) is not clear. The goal of our study was to characterize the anticancer effects and molecular mechanisms of matrine in SW480 CRC cells in vitro. Matrine treatment reduced mitochondrial metabolic function and ATP levels, repressed mitochondrial membrane potential, evoked mitochondrial reactive oxygen species accumulation, and promoted cyt-c-related mitochondrial apoptosis activation. In addition, we found that matrine treatment activated mitochondrial fission through upregulating mitochondrial elongation factor 1 (MIEF1); silencing of MIEF1 prevented matrine-mediated mitochondrial damage and reversed the decrease in SW480 cell viability. Moreover, matrine treatment affected MIEF1 expression via the large tumor suppressor-2 (LATS2)-Hippo axis, and LATS2 deficiency suppressed the anticancer actions exerted by matrine on SW480 cancer cells. In summary, we show for the first time that matrine inhibits SW480 cell survival by activating MIEF1-related mitochondrial division via the LATS2-Hippo pathway. These findings explain the anticancer mechanisms of matrine in CRC and also identify the LATS2-MIEF1 signaling pathway as an effective target for the treatment of CRC.

MKP1 overexpression reduces TNF-α-induced cardiac injury via suppressing mitochondrial fragmentation and inhibiting the JNK-MIEF1 pathways.

Mitochondrial stress has been acknowledged as the pathogenesis for tumor necrosis factor-α (TNF-α)-induced septic cardiomyopathy. Recently, MAP kinase phosphatase 1 (MKP1) downregulation and mitochondrial fragmentation modulate the mitochondrial stress via multiple molecular mechanisms. Thereby, the goal of our current work is to figure out the functional role of mitochondrial fragmentation in TNF-α-induced septic cardiomyopathy. Our results exhibited that MKP1 expression was significantly repressed in hearts treated by TNF-α. Overexpression of MKP1 sustained cardiac function and attenuated cardiomyocytes death in TNF-α-treated hearts. At the molecular levels, decreased MKP1 induced mitochondrial stress, as indicated by mitochondrial calcium overloading, mitochondrial oxidative stress, mitochondrial antioxidant downregulation, mitochondrial membrane potential reduction, mitochondrial bioenergetics suppression, mitochondrial proapoptotic factors liberation, and caspase-9 apoptotic pathway activation. To the end, we illustrated that MKP1-modulated mitochondrial stress via mitochondrial fragmentation; reactivation of mitochondrial fragmentation abolished the protective effect of MKP1 overexpression on mitochondrial function. Further, MKP1 affected mitochondrial division in a mechanism through the JNK-MIEF1 axis. Blockade of JNK pathway abolished the regulatory actions of MKP1 on mitochondrial division. Altogether, our results identify MKP1 as a novel cardioprotective factor in TNF-α-related septic cardiomyopathy via affecting mitochondrial division by the way of JNK-MIEF1 signaling pathway. Therefore, MKP1 expression, mitochondrial fragmentation modification, and JNK-MIEF1 pathway modulation may be considered as potential therapeutic targets for the treatment of cardiac injury induced by sepsis.

Large tumor suppressor kinase 2 overexpression attenuates 5-FU-resistance in colorectal cancer via activating the JNK-MIEF1-mitochondrial division pathway.

BACKGROUND: 5-Fluorouracil (5-FU) is a standard treatment for colorectal cancer, but most patients develop 5-FU resistance. Here, we conducted experiments to identify an effective approach to augment 5-FU-based treatment in colorectal cancer in vitro. METHODS: SW480 cells were in the present study and treated with 5-FU. Besides, LATS2 adenovirus vectors were infected into SW480 cells. Western blotting, immunofluorescence and ELISA were used to evaluate cell death and mitochondrial function. Pathway blocker was used to verify the role of MAPK-JNK pathway in SW480 cell death. RESULTS: An obvious drop in large tumor suppressor kinase 2 (LATS2) expression was observed in SW480 cells after treatment with 5-FU. In addition, upregulation of LATS2 expression through infection with LATS2 adenovirus further increased the reduction of SW480 cell viability induced by 5-FU. Functional exploration showed that 5-FU treatment suppressed mitochondrial membrane potential, enhanced cyt-c release into the nucleus, induced an oxidative injury environment by promoting ROS production, and eventually upregulated Bax-related mitochondrial apoptosis. Besides, LATS2 overexpression in combination with 5-FU treatment further perturbed mitochondrial homeostasis, and this effect was achieved by elevating mitochondrial division. Mechanistically, LATS2 overexpression and 5-FU co-treatment amplified mitochondrial division by upregulating MIEF1 expression in a manner dependent on MAPK-JNK axis. Knockdown of MIEF1 using an siRNA-mediated loss of function assay and/or inhibition of the MAPK-JNK pathway using the specific inhibitor SP600125 abolished LATS2/5-FU-mediated deleterious effects on mitochondrial performance and SW480 cell viability. CONCLUSIONS: In light of the above findings, LATS2 downregulation could be a potential mechanism of low response to 5-FU treatment. Overexpression of LATS2 to further disrupt mitochondrial function via the JNK-MIEF1 signalling pathway might be a method to optimize 5-FU-based chemotherapy.

Mst1 overexpression combined with Yap knockdown augments thyroid carcinoma apoptosis via promoting MIEF1-related mitochondrial fission and activating the JNK pathway.

BACKGROUND: Cancer cell viability is strongly modulated by the Hippo pathway, which includes mammalian STE20-like protein kinase 1 (Mst1) and yes-associated protein (Yap). Although the roles of Mst1 and Yap in thyroid carcinoma cell death have been fully addressed, no study has determined whether differential modification of Mst1 and Yap could further suppress thyroid carcinoma progression. The aim of our study was to explore the antiapoptotic effects exerted by combined Mst1 overexpression and Yap knockdown in thyroid carcinoma MDA-T32 cells in vitro. METHODS: Mst1 adenovirus and Yap shRNA were transfected into MDA-T32 cells to overexpress Mst1 and inhibit Yap, respectively. Cell viability and death were determined via an MTT assay, a TUNEL assay and western blotting. Mitochondrial function, mitochondrial fission and pathway studies were performed via western blotting and immunofluorescence. RESULTS: The results of our study showed that combined Mst1 overexpression and Yap knockdown further augmented MDA-T32 cell death by mediating mitochondrial damage. In addition, cancer cell migration and proliferation were suppressed by combined Mst1 overexpression and Yap knockdown. At the molecular level, mitochondrial membrane potential, ATP production, respiratory function, and caspase-9-related apoptosis were activated by combined Mst1 overexpression and Yap knockdown. Further, we found that fatal mitochondrial fission was augmented by combined Mst1 overexpression and Yap knockdown in a manner dependent on the JNK-MIEF1 pathway. Inhibition of JNK-MIEF1 pathway activity abolished the proapoptotic effects exerted by Mst1/Yap on MDA-T32 cells. CONCLUSIONS: Taken together, our data suggest that Mst1 activation and Yap inhibition coordinate to augment thyroid cancer cell death by controlling the JNK-MIEF1-mitochondria pathway, suggesting that differential regulation of the core Hippo pathway components is potentially a novel therapeutic tool for the treatment of thyroid cancer.

Drp1, Mff, Fis1, and MiD51 are coordinated to mediate mitochondrial fission during UV irradiation-induced apoptosis.

Mitochondrial fission and proteins vital to this process play essential roles in apoptosis. Several mitochondrial outer membrane proteins, including mitochondrial fission protein 1 (Fis1), mitochondrial fission factor (Mff) and mitochondrial dynamics of 51 kDa protein (MiD51), also known as mitochondrial elongation factor 1 (MEIF1), have been reported to promote mitochondrial fission by recruiting the GTPase dynamin-related protein 1 (Drp1). However, it remains unclear how these fission factors coordinate to control apoptotic mitochondrial fission. Molecular studies have suggested the existence of interaction between Mff and Drp1, but fundamental questions remain concerning their function. In the present study, we reported that the phosphorylation status of Drp1-Ser(637) was essential for its interaction with Mff. UV stimulation induced a decrease in cytoplasmic and mitochondrial Drp1 phosphorylation on Ser(637) and enhanced the interaction between Drp1 and Mff, resulting in mitochondrial fragmentation. Simultaneously, the interaction increased markedly between Fis1 and MiD51/MIEF1, whereas the interaction between Drp1 and MiD51/MIEF1 decreased significantly after UV irradiation, which suggests that Fis1 competitively binds to MiD51/MIEF1 to activate Drp1 indirectly. Moreover, Mff-Drp1 binding and Mff-mediated recruitment of Drp1 to mitochondria did not require Bax during UV stimulation. Our study revealed a novel role of Mff in regulation of mitochondrial fission and showed how the fission proteins are orchestrated to mediate the fission process during apoptosis.

The effect of mitochondrial calcium uniporter on mitochondrial fission in hippocampus cells ischemia/reperfusion injury.

The mitochondrial calcium uniporter (MCU) transports free Ca(2+) into the mitochondrial matrix, maintaining Ca(2+) homeostasis, thus regulates the mitochondrial morphology. Previous studies have indicated that there was closely crosstalk between MCU and mitochondrial fission during the process of ischemia/reperfusion injury. This study constructed a hypoxia reoxygenation model using primary hippocampus neurons to mimic the cerebral ischemia/reperfusion injury and aims to explore the exactly effect of MCU on the mitochondrial fission during the process of ischemia/reperfusion injury and so as the mechanisms. Our results found that the inhibitor of the MCU, Ru360, decreased mitochondrial Ca(2+) concentration, suppressed the expression of mitochondrial fission protein Drp1, MIEF1 and Fis1, and thus improved mitochondrial morphology significantly. Whereas spermine, the agonist of the MCU, had no significant impact compared to the I/R group. This study demonstrated that the MCU regulates the process of mitochondrial fission by controlling the Ca(2+) transport, directly upregulating mitochondrial fission proteins Drp1, Fis1 and indirectly reversing the MIEF1-induced mitochondrial fusion. It also provides new targets for brain protection during ischemia/reperfusion injury.

Valproic acid regulates MIEF1 through MST2-HIPPO to suppress breast cancer growth.

AIMS: To determine the effects of valproic acid (VPA) on anti-proliferative effects and mitochondrial function in breast cancer cells and the underlying mechanisms involved in the effects, with a focus on signal transduction. MAIN METHODS: The inhibitory effect of valproic acid on breast cancer in vivo and in vitro was evaluated by cellular and animal experiments. Mitochondria-related proteins as well as hippo pathway were monitored by western blotting. The effects of VPA on mitochondrial membrane potential, reactive oxygen species, and apoptosis were confirmed by flow cytometry. In addition, the involvement of hippo pathway in the regulation of mitochondrial function by VPA was verified by XMU-MP-1 (MST2 inhibitor). KEY FINDINGS: In this study, we highlight that VPA significantly attenuates mitochondrial function, leading to inhibited cell proliferation and reduced colony formation in MCF-7 and MDA-MB-231 breast cancer cells. Mechanistically, VPA-induced suppression of mitochondrial aerobic respiration was mediated by decreased expression of mitochondrial elongation factor 1 through activation of the hippo pathway, resulting in impaired breast cancer growth. In summary, we uncover a novel mechanism of VPA in regulating mitochondrial aerobic respiration, which is essential for developing an effective approach in breast cancer therapy. SIGNIFICANCE: Mitochondrial aerobic respiration and its products are the main sources of energy for tumors; therefore, studying the role of mitochondrial function in tumor cells is significant. VPA has been used as a therapeutic agent for cancer. However, the detail mechanism underlying the effects of VPA on mitochondrial function in breast cancer remains unclear.

The Molecular Assembly State of Drp1 Controls its Association With the Mitochondrial Recruitment Receptors Mff and MIEF1/2.

Drp1 is a central player in mitochondrial fission and is recruited to mitochondria by Mff and MIEFs (MIEF1 and MIEF2), but little is known about how its assembly state affects Drp1 mitochondrial recruitment and fission. Here, we used in vivo chemical crosslinking to explore the self-assembly state of Drp1 and how it regulates the association of Drp1 with MIEFs and Mff. We show that in intact mammalian cells Drp1 exists as a mixture of multiple self-assembly forms ranging from the minimal, probably tetrameric, self-assembly subunit to several higher order oligomers. Precluding mitochondria-bound Drp1 in Mff/MIEF1/2-deficient cells does not affect the oligomerization state of Drp1, while conversely forced recruitment of Drp1 to mitochondria by MIEFs or Mff facilitates Drp1 oligomerization. Mff preferentially binds to higher order oligomers of Drp1, whereas MIEFs bind to a wider-range of Drp1 assembly subunits, including both lower and higher oligomeric states. Mff only recruits active forms of Drp1, while MIEFs are less selective and recruit both active and inactive Drp1 as well as oligomerization- or GTPase-deficient Drp1 mutants to mitochondria. Moreover, all the fission-incompetent Drp1 mutants tested (except the monomeric mutant K668E) affect Drp1-driven mitochondrial dynamics via incorporation of the mutants into the native oligomers to form function-deficient Drp1 assemblies. We here confirm that MIEFs also serve as a platform facilitating the binding of Drp1 to Mff and loss of MIEFs severely impairs the interaction between Drp1 and Mff. Collectively, our findings suggest that Mff and MIEFs respond differently to the molecular assembly state of Drp1 and that the extent of Drp1 oligomerization regulates mitochondrial dynamics.

Dominant mutations in MIEF1 affect mitochondrial dynamics and cause a singular late onset optic neuropathy.

Inherited optic neuropathies are the most common mitochondrial diseases, leading to neurodegeneration involving the irreversible loss of retinal ganglion cells, optic nerve degeneration and central visual loss. Importantly, properly regulated mitochondrial dynamics are critical for maintaining cellular homeostasis, and are further regulated by MIEF1 (mitochondrial elongation factor 1) which encodes for MID51 (mitochondrial dynamics protein 51), an outer mitochondrial membrane protein that acts as an adaptor protein to regulate mitochondrial fission. However, dominant mutations in MIEF1 have not been previously linked to any human disease. Using targeted sequencing of genes involved in mitochondrial dynamics, we report the first heterozygous variants in MIEF1 linked to disease, which cause an unusual form of late-onset progressive optic neuropathy characterized by the initial loss of peripheral visual fields. Pathogenic MIEF1 variants linked to optic neuropathy do not disrupt MID51's localization to the outer mitochondrial membrane or its oligomerization, but rather, significantly disrupt mitochondrial network dynamics compared to wild-type MID51 in high spatial and temporal resolution confocal microscopy live imaging studies. Together, our study identifies dominant MIEF1 mutations as a cause for optic neuropathy and further highlights the important role of properly regulated mitochondrial dynamics in neurodegeneration.

Loss of MIEF1/MiD51 confers susceptibility to BAX-mediated cell death and PINK1-PRKN-dependent mitophagy.

Mitochondrial dynamics is highly implicated in a plethora of cellular processes including apoptosis and mitophagy. However, little is known about the scope and precise functions of mitochondrial dynamics proteins for mitochondrial quality control and cellular homeostasis. Whether mitochondrial dynamics proteins serve in cellular processes reliant on mitochondrial fission-fusion is still not fully explored. MIEF1/MiD51 (mitochondrial elongation factor 1) is known to promote mitochondrial fission via the recruitment of GTPase protein DNM1L/DRP1 (dynamin 1 like), but the fundamental understandings of MIEF1 for mitochondrial-dependent cellular processes are largely elusive. Here, we report novel roles of MIEF1 in responding to apoptotic stimuli and mitochondrial damage. Given our result that staurosporine (STS) treatment induced the degradation of MIEF1 via the ubiquitin-proteasome system (UPS), we are motivated to explore the role of MIEF1 in apoptosis. MIEF1 loss triggered the imbalance of BCL2 family members on the mitochondria, consequently initiating the translocation of BAX onto the mitochondria, catalyzing the decrease of mitochondrial membrane potential and promoting the release of DIABLO/SMAC (diablo IAP-binding mitochondrial protein) and CYCS (cytochrome c, somatic). We further demonstrate that MIEF1 deficiency impaired mitochondrial respiration and induced mitochondrial oxidative stress, sensitizing cells to PINK1-PRKN-mediated mitophagy. The recruitment of PRKN to depolarized mitochondria modulated the UPS-dependent degradation of MFN2 (mitofusin 2) and FIS1 (fission, mitochondrial 1) specifically, to further promote mitophagy. Our findings uncover a bridging role of MIEF1 integrating cell death and mitophagy, unlikely dependent on mitochondrial dynamics, implying new insights to mechanisms determining cellular fate.Abbreviations: ActD: actinomycin D; BAX: BCL2 associated X, apoptosis regulator; BAK1: BCL2 antagonist/killer 1; BCL2L1: BCL2 like 1; BMH: 1,6-bismaleimidohexane; CCCP: carbonyl cyanide 3-chlorophenylhydrazone; CHX: cycloheximide; CQ: chloroquine; CYCS: cytochrome c, somatic; DIABLO: diablo IAP-binding mitochondrial protein; DKO: double knockout; DNM1L/DRP1: dynamin 1 like; FIS1: fission, mitochondrial 1; GFP: green fluorescent protein; IP: immunoprecipitation; MFN1: mitofusin 1; MFN2: mitofusin 2; MG132: carbobenzoxy-Leu-Leu-leucinal; MIEF1/MiD51: mitochondrial elongation factor 1; MIEF2/MiD49: mitochondrial elongation factor 2; MOMP: mitochondrial outer membrane permeabilization; MTR: MitoTracker Red; OA: oligomycin plus antimycin A; OCR: oxygen consumption rate; OMM: outer mitochondrial membrane; PARP: poly(ADP-ribose) polymerase; PI: propidium iodide; PINK1: PTEN induced kinase 1; PRKN: parkin RBR E3 ubiquitin protein ligase; ROS: reactive oxygen species; SD: standard deviation; STS: staurosporine; TNF: tumor necrosis factor; UPS: ubiquitin-proteasome system; VDAC1: voltage dependent anion channel 1.

Genetic ablation of TAZ induces HepG2 liver cancer cell apoptosis through activating the CaMKII/MIEF1 signaling pathway.

BACKGROUND AND OBJECTIVE: Transcriptional coactivator with PDZ-binding motif (TAZ) has been found to be associated with tumor progression. Mitochondrial homeostasis regulates cancer cell viability and metastasis. However, the roles of TAZ and mitochondrial homeostasis in liver cancer viability have not been explored. The aim of our study was to investigate the influence of TAZ on HepG2 liver cancer cell apoptosis. MATERIALS AND METHODS: HepG2 liver cancer cell was used in the present study, and shRNA against TAZ was transfected into HepG2 cell to knockdown TAZ expression. Mitochondrial function was analyzed using Western blotting, immunofluorescence assay, and flow cytometry. Pathway blocker was used to confirm the role of CaMKII pathway in TAZ-mediated cancer cell death. RESULTS: Our results indicated that TAZ deletion induced death in HepG2 cell via apoptosis. Biological analysis demonstrated that mitochondrial stress, including mitochondrial bioenergetics disorder, mitochondrial oxidative stress, and mitochondrial apoptosis, were activated by TAZ deletion. Furthermore, we found that TAZ affected mitochondrial stress by triggering mitochondrial elongation factor 1 (MIEF1)-related mitochondrial dysfunction. The loss of MIEF1 sustained mitochondrial function and promoted cancer cell survival. Molecular investigation illustrated that TAZ regulated MIEF1 expression via the CaMKII signaling pathway. Blockade of the CaMKII pathway prevented TAZ-mediated MIEF1 upregulation and improved cancer cell survival. CONCLUSION: Taken together, our results highlight the key role of TAZ as a master regulator of HepG2 liver cancer cell viability via the modulation of MIEF1-related mitochondrial stress and the CaMKII signaling pathway. These findings define TAZ and MIEF1-related mitochondrial dysfunction as tumor suppressors that act by promoting cancer apoptosis via the CaMKII signaling pathway, with potential implications for new approaches to liver cancer therapy.

Yap-Hippo promotes A549 lung cancer cell death via modulating MIEF1-related mitochondrial stress and activating JNK pathway.

Although the role of Yes-associated protein (Yap) has been described in the progression of lung cancer, the downstream effector of the Yap-Hippo pathway has not been identified. Accordingly, the aim of our study is to explore whether Yap modulates the activity of lung cancer by controlling mitochondrial elongation factor 1 (MIEF1)-related mitochondrial stress in a manner dependent on the JNK pathway. Cell viability was determined via MTT, LDH release and immunofluorescence assays. ATP production, the mitochondrial membrane potential, and caspase-9 activity were investigated to assess mitochondrial function. siRNA transfection and pathway blockers were used to observe the roles of MIEF1 and JNK in Yap-modulated cell viability in lung cancer cells in vitro. Yap deletion reduced cell viability in A549 and H358 lung cancer cells. At the molecular level, Yap deletion promoted mitochondrial dysfunction, as evidenced by the decreased mitochondrial potential, increased mitochondrial oxidative stress, augmented mitochondrial pro-apoptotic factor leakage and elevated caspase-9 activity. In addition, we found that Yap modulated mitochondrial stress via MIEF1 and that loss of MIEF1 abolished the regulatory actions of Yap on mitochondrial stress and cell viability. Besides, we provided evidence to support the necessary role of JNK in Yap-mediated MIEF1 upregulation. Inhibition of JNK abolished the promotive effect of Yap deletion on MIEF1 activation. Taken together, our results identified the JNK-MIEF1 pathway and mitochondrial stress as downstream effectors of Yap in lung cancer. This finding suggests a novel approach for the treatment of lung cancer in clinical practice.