Elongation Of Mitochondria Research Articles

Mitochondrial Homeostasis Regulating Mitochondrial Number and Morphology Is a Distinguishing Feature of Skeletal Muscle Fiber Types in Marine Teleosts.

Fishes' skeletal muscles are crucial for swimming and are differentiated into slow-twitch muscles (SM) and fast-twitch muscles (FM) based on physiological and metabolic properties. Consequently, mitochondrial characteristics (number and morphology) adapt to each fiber type's specific functional needs. However, the mechanisms governing mitochondrial adaptation to the specific bioenergetic requirements of each fiber type in teleosts remain unclear. To address this knowledge gap, we investigated the mitochondrial differences and mitochondrial homeostasis status (including biogenesis, autophagy, fission, and fusion) between SM and FM in teleosts using Takifugu rubripes as a representative model. Our findings reveal that SM mitochondria are more numerous and larger compared to FM. To adapt to the increased mitochondrial number and size, SM exhibit elevated mitochondrial biogenesis and dynamics (fission/fusion), yet show no differences in mitochondrial autophagy. Our study provides insights into the adaptive mechanisms shaping mitochondrial characteristics in teleost muscles. The abundance and elongation of mitochondria in SM are maintained through elevated mitochondrial biogenesis, fusion, and fission, suggesting an adaptive response to fulfill the bioenergetic demands of SM that rely extensively on OXPHOS in teleosts. Our findings enhance our understanding of mitochondrial adaptations in diverse muscle types among teleosts and shed light on the evolutionary strategies of bioenergetics in fishes.

Altered Mitochondrial Function and Accelerated Aging Phenotype in Neural Stem Cells Derived from Dnm1l Knockout Embryonic Stem Cells.

Mitochondria are crucial for cellular energy metabolism and are involved in signaling, aging, and cell death. They undergo dynamic changes through fusion and fission to adapt to different cellular states. In this study, we investigated the effect of knocking out the dynamin 1-like protein (Dnm1l) gene, a key regulator of mitochondrial fission, in neural stem cells (NSCs) differentiated from Dnm1l knockout embryonic stem cells (Dnm1l-/- ESCs). Dnm1l-/- ESC-derived NSCs (Dnm1l-/- NSCs) exhibited similar morphology and NSC marker expression (Sox2, Nestin, and Pax6) to brain-derived NSCs, but lower Nestin and Pax6 expression than both wild-type ESC-derived NSCs (WT-NSCs) and brain-derived NSCs. In addition, compared with WT-NSCs, Dnm1l-/- NSCs exhibited distinct mitochondrial morphology and function, contained more elongated mitochondria, showed reduced mitochondrial respiratory capacity, and showed a metabolic shift toward glycolysis for ATP production. Notably, Dnm1l-/- NSCs exhibited impaired self-renewal ability and accelerated cellular aging during prolonged culture, resulting in decreased proliferation and cell death. Furthermore, Dnm1l-/- NSCs showed elevated levels of inflammation and cell stress markers, suggesting a connection between Dnm1l deficiency and premature aging in NSCs. Therefore, the compromised self-renewal ability and accelerated cellular aging of Dnm1l-/- NSCs may be attributed to mitochondrial fission defects.

Morphological and mitochondrial changes in murine choroid plexus epithelial cells during healthy aging

BackgroundChoroid plexuses (ChPs) are intraventricular structures mainly composed by specialized epithelial cells interconnected by tight junctions that establish the blood-cerebrospinal fluid (CSF) barrier. ChPs are essential to produce CSF and transport solutes from and into the brain. Deterioration of ChP function and morphology has been correlated to worsening of neurodegenerative disorders. We here map morpho-functional changes in the ChP epithelial cells during healthy aging, starting from young adult to 2-years old mice.MethodsWe used a multi-tiered approach, including transmission electron microscopy (TEM), immunohistochemistry, RT-qPCR, Western Blot and 2-photon microscopy (2-PM) at multiple timepoints ranging from young adult to 2-years old mice.ResultsWe identified distinct morpho-functional modifications in epithelial cells of ChP starting from 8 to 12 months of age, which mostly remained stable up to 2 years. These changes include flattening of the epithelium, reduction of microvilli length and an augmentation of interrupted tight junctions. We also found a decrease in mitochondria density together with elongation of mitochondria in older mice. Morphological mitochondrial rearrangements were accompanied by increased superoxide levels, decreased membrane potential and decreased mitochondrial motility in aged mice. Interestingly, most of the age-related changes were not accompanied by modification of protein and/or gene expression levels and aged mitochondria effectively responded to acute pharmacological stressful stimuli.ConclusionsOur study suggests a long-term progression of multiple morpho-functional features of the mouse choroid plexus epithelium during adulthood followed by structural remodeling during the aging process. These findings can lead to a better understanding on how functional and morphological rearrangements of ChP are correlated during aging.

Inflammatory Milieu Induces Mitochondrial Alterations and Neuronal Activations in Hypothalamic POMC Neurons in a Time-Dependent Manner.

Inflammation has been associated with numerous neurological disorders. Inflammatory environments trigger a series of cellular and physiological alterations in the brain. However, how inflammatory milieu affects neuronal physiology and how neuronal alterations progress in the inflammatory environments are not fully understood. In this study, we examined the effects of pro-inflammatory milieu on mitochondrial functions and neuronal activities in the hypothalamic POMC neurons. Treating mHypoA-POMC/GFP1 with the conditioned medium collected from LPS activated macrophage were employed to mimic the inflammatory milieu during hypothalamic inflammation. After a 24-h treatment, intracellular ROS/RNS levels were elevated, and the antioxidant enzymes were reduced. Mitochondrial respiration and mitochondrial functions, including basal respiratory rate, spared respiration capacity, and maximal respiration, were all significantly compromised by inflammatory milieu. Moreover, pro-inflammatory cytokines altered mitochondrial dynamics in a time-dependent manner, resulting in the elongation of mitochondria in POMC neurons after a 24-h treatment. Additionally, the increase of C-Fos and Pomc genes expression indicated that the neurons were activated upon the stimulation of inflammatory environment. This neuronal activation of were confirmed on the LPS-challenged mice. Collectively, a short-term to midterm exposure to inflammatory milieu stimulated metabolic switch and neuronal activation, whereas chronic exposure triggered the elevation of oxidative stress, the decrease of the mitochondrial respiration, and the alterations of mitochondrial dynamics.

Caveolin-1 signaling-driven mitochondrial fission and cytoskeleton remodeling promotes breast cancer migration.

Mitochondria are highly dynamic organelles that constantly divide and fuse to maintain their proper structure and function. Cancer cells are often accompanied by an imbalance of mitochondrial fusion and fission, cancer progression is greatly affected by this imbalance. Here, we found that high-metastatic breast cancer MDA-MB-231 cells possess higher caveolin-1 (Cav-1) expression compared with low-metastatic breast cancer MCF-7 cells or normal breast epithelial MCF-10A cells. Downregulation of Cav-1 decreases the migratory and invasive abilities of MDA-MB-231 cells. Our results further demonstrated that downregulation of Cav-1 facilitated DRP1 and MFN2 to translocate to mitochondria, increasing the inhibitory phosphorylation level of DRP1 at Ser637 by protein kinase A (PKA), resulting in mitochondria elongation. We also showed that downregulation of Cav-1 significantly reduced the Rac1 activity by affecting intracellular reactive oxygen species (ROS) generation, which then inhibited F-actin formation. Based on these findings, we proposed that Cav-1 mediated mitochondrial fission-affected intracellular ROS generation and activated Rho GTPases, leading to F-actin-dependent formation of lamellipodia and promotion of breast cancer motility.

Abstract 3773: Mitochondrial fusion is required to support higher rates of respiration in proliferating cancer cells

Abstract Proliferating cancer cells exhibit an elevated rate of glucose consumption and lactate excretion, a phenomenon described as the “Warburg effect”. Although increased glycolytic flux in proliferating cells is well established, metabolic changes associated with mitochondria still remain elusive. Here, using various inducible quiescence models and mass spectrometry-based metabolomics, we directly compared mitochondrial respiration and TCA cycle activity from the same cells in the proliferative and quiescent states. We found that mitochondrial respiration and TCA cycle flux is significantly increased during proliferation in both transformed and non-transformed cell lines. This increase was supported by mitochondrial fusion rather than mitochondrial biogenesis. Notably, as cells transitioned from a quiescent state to a proliferative state, the elongation of mitochondria occurred as quickly as 3 hours. Our results reveal that the Warburg effect, which was originally associated with dysfunctional mitochondria, does not necessitate decreased mitochondrial respiration. Rather, in many proliferating cancer cells, mitochondrial respiration is actually elevated, underscoring the importance of mitochondrial respiration during proliferation. Citation Format: Yahui Wang, Conghui Yao, Ethan Stancliffe, Ronald Fowle-Grider, Rencheng Wang, Cheng Wang, Michaela Schwaiger-Haber, Leah P. Shriver, Jason Weber, Gary J. Patti. Mitochondrial fusion is required to support higher rates of respiration in proliferating cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3773.

Abstract 12199: Sirtuin1 Mediates Pressure Overload-Induced Cardiac Pathological Hypertrophy Through Regulating Mitochondrial Dynamics

Introduction: Sirtuin 1 (SIRT1) is a nicotinamide adenine dinucleotide-dependent histone deacetylase. We revealed that the pressure-overload-induced SIRT1 can exacerbate cardiac systolic dysfunction. But the mechanisms by which SIRT1 undermines cardiac function under pathological conditions remain unclear. Hypothesis: SIRT1 induced by pathological hypotrophy disturbs mitochondrial dynamics homeostasis resulting in an exacerbated cardiac dysfunction. Methods: Wild-type littermates SIRT1 f/f (3-5 months) and inducible cardiomyocyte-specific SIRT1 knockout (icSIRT1 -/- ) (3-5 months) C57BL/6J mice were subjected to transverse aortic constriction (TAC) surgery for six weeks. Echocardiography assessed cardiac functions and CODA tail-cuff system measured blood pressure. The mitochondrial dynamics were examined by immunoblotting and transmission electron microscopy (TEM). Results: The echocardiography showed that there were significant reductions in ejection fraction (EF) and fractional shortening (FS) of the SIRT1 f/f mice after 6-weeks of TAC-induced pressure overload. The diastolic and systolic blood pressure were increased in SIRT1 f/f mice during TAC surgery. Intriguingly, icSIRT1 -/- versus SIRT1 f/f demonstrated significant resistance to pathological hypertrophy caused by TAC-induced pressure overload as determined with echocardiography and blood pressure measurements. The immunoblotting revealed that the six weeks of TAC surgery caused significantly higher levels of the mitochondrial fusion protein, mitofusin 2 (MFN2), in SIRT1 f/f versus icSIRT1 -/- hearts. In addition, TEM data showed more elongated mitochondria in SIRT1 f/f versus icSIRT1 -/- hearts, indicating a critical role of SIRT1 in maintaining mitochondrial dynamic homeostasis under pathological stress conditions. Metabolomics and proteomics showed that SIRT1 regulates carnitine palmitoyltransferase-1 and long-chain acyl CoA dehydrogenase to modulate fatty acid metabolism in response to TAC-induced stress. Conclusions: Cardiomyocyte SIRT1 plays a key role in cardiac mitochondrial dynamics during pathological stress conditions. Inhibition of SIRT1 activity could be a good strategy to ameliorate pressure overload-induced hypertrophy.

Hypoxia compromises the mitochondrial metabolism of Alzheimer's disease microglia via HIF1.

Genetic Alzheimer's disease (AD) risk factors associate with reduced defensive amyloid β plaque-associated microglia (AβAM), but the contribution of modifiable AD risk factors to microglial dysfunction is unknown. In AD mouse models, we observe concomitant activation of the hypoxia-inducible factor 1 (HIF1) pathway and transcription of mitochondrial-related genes in AβAM, and elongation of mitochondria, a cellular response to maintain aerobic respiration under low nutrient and oxygen conditions. Overactivation of HIF1 induces microglial quiescence in cellulo, with lower mitochondrial respiration and proliferation. In vivo, overstabilization of HIF1, either genetically or by exposure to systemic hypoxia, reduces AβAM clustering and proliferation and increases Aβ neuropathology. In the human AD hippocampus, upregulation of HIF1α and HIF1 target genes correlates with reduced Aβ plaque microglial coverage and an increase of Aβ plaque-associated neuropathology. Thus, hypoxia (a modifiable AD risk factor) hijacks microglial mitochondrial metabolism and converges with genetic susceptibility to cause AD microglial dysfunction.

Changes in the Expression of Mitochondrial Morphology-Related Genes during the Differentiation of Murine Embryonic Stem Cells.

During embryonic development, cells undergo changes in gene expression, signaling pathway activation/inactivation, metabolism, and intracellular organelle structures, which are mediated by mitochondria. Mitochondria continuously switch their morphology between elongated tubular and fragmented globular via mitochondrial fusion and fission. Mitochondrial fusion is mediated by proteins encoded by Mfn1, Mfn2, and Opa1, whereas mitochondrial fission is mediated by proteins encoded by Fis1 and Dnm1L. Here, we investigated the expression patterns of mitochondria-related genes during the differentiation of mouse embryonic stem cells (ESCs). Pluripotent ESCs maintain stemness in the presence of leukemia inhibitory factor (LIF) via the JAK-STAT3 pathway but lose pluripotency and differentiate in response to the withdrawal of LIF. We analyzed the expression levels of mitochondrial fusion- and fission-related genes during the differentiation of ESCs. We hypothesized that mitochondrial fusion genes would be overexpressed while the fission genes would be downregulated during the differentiation of ESCs. Though the mitochondria exhibited an elongated morphology in ESCs differentiating in response to LIF withdrawal, only the expression of Mfn2 was increased and that of Dnm1L was decreased as expected, the other exceptions being Mfn1, Opa1, and Fis1. Next, by comparing gene expression and mitochondrial morphology, we proposed an index that could precisely represent mitochondrial changes during the differentiation of pluripotent stem cells by analyzing the expression ratios of three fusion- and two fission-related genes. Surprisingly, increased Mfn2/Dnm1L ratio was correlated with elongation of mitochondria during the differentiation of ESCs. Moreover, application of this index to other specialized cell types revealed that neural stems cells (NSCs) and mouse embryonic fibroblasts (MEFs) showed increased Mfn2/Dnm1L ratio compared to ESCs. Thus, we suggest that the Mfn2/Dnm1L ratio could reflect changes in mitochondrial morphology according to the extent of differentiation.

Ghrelin effects on mitochondrial fitness modulates macrophage function

Over the past years, systemic derived cues that regulate cellular metabolism have been implicated in the regulation of immune responses. Ghrelin is an orexigenic hormone produced by enteroendocrine cells in the gastric mucosa with known immunoregulatory roles. The mechanism behind the function of ghrelin in immune cells, such as macrophages, is still poorly understood. Here, we explored the hypothesis that ghrelin leads to alterations in macrophage metabolism thus modulating macrophage function. We demonstrated that ghrelin exerts an immunomodulatory effect over LPS-activated peritoneal macrophages, as evidenced by inhibition of TNF-α and IL-1β secretion and increased IL-12 production. Concomitantly, ghrelin increased mitochondrial membrane potential and increased respiratory rate. In agreement, ghrelin prevented LPS-induced ultrastructural damage in the mitochondria. Ghrelin also blunted LPS-induced glycolysis. In LPS-activated macrophages, glucose deprivation did not affect ghrelin-induced IL-12 secretion, whereas the inhibition of pyruvate transport and mitochondria-derived ATP abolished ghrelin-induced IL-12 secretion, indicating a dependence on mitochondrial function. Ghrelin pre-treatment of metabolic activated macrophages inhibited the secretion of TNF-α and enhanced IL-12 levels. Moreover, ghrelin effects on IL-12, and not on TNF-α, are dependent on mitochondria elongation, since ghrelin did not enhance IL-12 secretion in metabolic activated mitofusin-2 deficient macrophages. Thus, ghrelin affects macrophage mitochondrial metabolism and the subsequent macrophage function.

The Fine Tuning of Drp1-Dependent Mitochondrial Remodeling and Autophagy Controls Neuronal Differentiation.

Mitochondria play a critical role in neuronal function and neurodegenerative disorders, including Alzheimer’s, Parkinson’s and Huntington diseases and amyotrophic lateral sclerosis, that show mitochondrial dysfunctions associated with excessive fission and increased levels of the fission protein dynamin-related protein 1 (Drp1). Our data demonstrate that Drp1 regulates the transcriptional program induced by retinoic acid (RA), leading to neuronal differentiation. When Drp1 was overexpressed, mitochondria underwent remodeling but failed to elongate and this enhanced autophagy and apoptosis. When Drp1 was blocked during differentiation by overexpressing the dominant negative form or was silenced, mitochondria maintained the same elongated shape, without remodeling and this increased cell death. The enhanced apoptosis, observed with both fragmented or elongated mitochondria, was associated with increased induction of unfolded protein response (UPR) and ER-associated degradation (ERAD) processes that finally affect neuronal differentiation. These findings suggest that physiological fission and mitochondrial remodeling, associated with early autophagy induction are essential for neuronal differentiation. We thus reveal the importance of mitochondrial changes to generate viable neurons and highlight that, rather than multiple parallel events, mitochondrial changes, autophagy and apoptosis proceed in a stepwise fashion during neuronal differentiation affecting the nuclear transcriptional program.

Deficiency in augmenter of liver regeneration accelerates liver fibrosis by promoting migration of hepatic stellate cell

BackgroundAugmenter of liver regeneration (ALR) protects liver from various injuries, however, the association of ALR with liver fibrosis, particularly its effect on hepatic stellate cells (HSC), remains unclear. In this study, we investigated the impact of ALR on the activation of HSC, a pivotal event in occurrence of liver fibrosis. MethodsLiver fibrosis was induced in vivo in mice with heterozygous ALR knockdown (ALR-KD) by administration of CCl4 or bile duct ligation. The effect of ALR-KD and ALR-overexpression on liver fibrosis was studied in mice and in HSC cells as well. ResultsHepatic collagen deposition and expression of α-smooth muscle actin (α-SMA) were significantly increased in the ALR-KD mice compared to wild-type mice. In vitro, ALR-shRNA resulted in the activation of HSC cell line (LX-2). Furthermore, ALR-shRNA promoted LX-2 cell migration, accompanied by increased filamentous actin (F-actin) assembly. The ALR-KD-mediated increase in HSC migration was associated with mitochondrial fusion, resulting in mitochondria elongation and enhancing ATP production. In contrast, ALR transfection (ALR-Tx) decelerated HSC migration and inhibited F-actin assembly, concomitantly enhancing mitochondrial fission and reducing ATP synthesis. Mechanically, stimulation of HSC migration by ALR-shRNA was attributed to the increased mitochondrial Ca2+ influx in HSCs. Treatment of ALR-shRNA-cells with Ruthenium Red (RuR), a specific inhibitor of mitochondrial calcium uniporter (MCU), significantly suppressed mitochondrial Ca2+ influx, HSC migration, mitochondrial fusion and ATP production. ALR-KD-induced HSC migration was verified in vitro in primary mouse HSCs. ConclusionInhibition of ALR expression aggravates liver fibrosis, probably via promoting HSC migration and mitochondrial fusion.

Abstract 280: MCL-1 Matrix Promotes Mitochondrial Fusion and Protects Against TAC Induced Hypertrophy

The anti-apoptotic BCL-2 protein MCL-1 is highly expressed in the myocardium, and we have previously found that deletion in cardiomyocytes leads to mitochondrial dysfunction and rapid development of heart failure. MCL-1 exists both in the outer mitochondrial membrane (MCL-1 OM ) and in the mitochondrial matrix (MCL-1 Matrix ). Because Mcl-1 -/- mice lack both MCL-1 Matrix and MCL-1 OM , the functional role of each form remains unknown. While studies have implicated MCL-1 OM in regulating apoptosis, very little is known about the functional role of MCL-1 Matrix . Here, we show that mitochondria are responsive to alterations in cellular metabolism, and culturing mouse embryonic fibroblasts (MEFs) in “oxidative” media containing galactose or acetoacetate led to a significant increase in the MCL-1 Matrix /MCL-1 OM ratio and elongation of mitochondria. We confirmed that MCL-1 Matrix overexpression promoted mitochondrial fusion and reduced maximal mitochondrial respiration under baseline conditions. Interestingly, the elongated mitochondria were initially protected against FCCP-induced fission and mitophagy. However, prolonged exposure to FCCP resulted in abrogation of MCL-1 Matrix import. Accumulation of MCL-1 Matrix on the outer membrane led to its degradation and subsequent induction of mitophagy. Furthermore, to investigate its functional role in the heart, we generated cardiac specific MCL-1 Matrix (αMHC-MCL-1 Matrix ) transgenic mice. The mice show no differences in cardiac structure and function compared to wild-type litter mates at 3 months. However, ultrastructural analysis revealed the presence of enlarged mitochondria in myocytes from αMHC-MCL-1 Matrix mice. Finally, we subjected WT and αMHC-MCL-1 Matrix mice to transverse aortic constriction (TAC) and we found that αMHC-MCL-1 Matrix mice had reduced heart weight to body weight (HW/BW) ratio and decreased expression of hypertrophy markers ANF, BNP, and MHC-β at two weeks post-TAC compared to WT. Thus, our studies indicate that cardiac overexpression of MCL-1 Matrix promotes mitochondrial enlargement in myocytes and alleviates stress placed on the heart during pressure overload.

Mitochondria elongation is mediated through SIRT1-mediated MFN1 stabilization

Mitochondria are highly dynamic organelles that change size and morphology by fusing together or dividing through fission. In response to cellular cues, signaling cascades may post-translationally modify mitochondria-shaping proteins, which lead to a change in mitochondria morphology. Here we show that nicotinamide (NAM), an inhibitor of sirtuin deacetylases, promotes degradation of mitochondria fusion protein mitofusin 1 (MFN1), suggesting that acetylation status of MFN1 is important for its protein stability. TIP60 but not PCAF acetyltransferase caused a reduction of MFN1 level. Meanwhile, siRNA-mediated knockdown of SIRT1 deacetylase caused a significant reduction of MFN1 whereas over-expression of SIRT1 increased its level in 293T cells. In vitro acetylation experiments showed that TIP60 increased the acetylation of MFN1 that was abolished by co-existence of SIRT1. Notably, MFN1 and SIRT1 levels were accumulated, along with mitochondria elongation under hypoxic conditions. Thus, the data suggest that mitochondria elongation under hypoxic condition is regulated through SIRT1-mediated MFN1 deacetylation and accumulation. The data provide an insight in the maintenance of cellular homeostasis through mitochondria morphological change.

Dynamic Changes of Mitochondrial Fusion and Fission in Brain Injury after Cardiac Arrest in Rats.

Mitochondria change their morphology dynamically by continual fusion and fission processes to fulfill their function. However, little is known about the effect of cardiac arrest on mitochondrial dynamics. This study aimed to investigate time-dependent change of the mitochondrial dynamics after brain ischemic injury in rats of cardiac arrest. After resuscitation, obvious neuronal injury, reduced adenosine triphosphate (ATP) levels, excessive reactive oxygen species (ROS) generation, decreased mitochondrial membrane potential (MMP), and increased release of mitochondrial cytochrome c were observed at 12 h and 24 h after cardiac arrest. Moreover, we found that elongation of mitochondria was observed at 4 h after cardiac arrest, whereas fragmented mitochondria were significantly increased, along with concomitant increase in the fission proteins Drp1 and Fis1 and a reduction in the fusion proteins Mfn1 and Mfn2 at 12 h and 24 h after cardiac arrest. Taken together, these findings suggest that imbalance in mitochondrial dynamics probably contributes to brain injury after cardiac arrest.

Abstract 103: MCL-1 Matrix Promotes Mitochondrial Fusion and Protects Against Mitophagy

MCL-1 is a less well characterized anti-apoptotic BCL-2 protein which is highly expressed in the myocardium. We recently discovered that loss of MCL-1 in cardiac myocytes leads to rapid mitochondrial dysfunction and development of heart failure. MCL-1 exists both in the outer mitochondrial membrane (MCL-1 OM ) and in the mitochondrial matrix (MCL-1 Matrix ). Because the Mcl-1 -/- knockout mice are missing both MCL-1 Matrix and MCL-1 OM , the functional importance of each form is still unknown. While studies have implicated MCL-1 OM in regulating apoptosis, very little is known about the functional role of MCL-1 Matrix . Here, we examined whether cardiac MCL-1 levels were altered in response to fasting. Our data show that the MCL-1 Matrix /MCL-1 OM protein ratio was significantly increased in mouse hearts after 24 and 48 h of fasting. This was mainly due to a decrease in MCL-1 OM , while MCL-1 Matrix levels were unchanged. We also confirmed that the reduction in MCL-1 OM did not lead to a significant activation of apoptosis. Mitochondria are responsive to alterations in cellular metabolism and culturing cells in “oxidative” media containing galactose or acetoacetate led to a significant increase in the MCL-1 Matrix /MCL-1 OM ratio due to a reduction in MCL-1 OM protein and to elongation of mitochondria. We also confirmed that the reduced MCL-1 OM levels did not result in activation of apoptosis. This suggests that the MCL-1 Matrix /MCL-1 OM ratio is altered in response to changes in metabolism and energy demand without activating apoptosis. We have started dissecting how altering the MCL-1 Matrix /MCL-1 OM ratio affects mitochondrial morphology and turnover. We confirmed that MCL-1 Matrix overexpression led to more fused mitochondria under baseline conditions. Interestingly, we also found that the elongated mitochondria were also more protected against FCCP-induced fission as well as degradation by autophagosomes. Thus, we propose that MCL-1 Matrix protects mitochondria from unnecessary mitophagy to preserve the bioenergetic capacity when nutrients are limited. We are currently evaluating how cardiac restricted overexpression of MCL-1 Matrix (αMHC-MCL-1 Matrix ) affects mitochondrial morphology, mitophagy and bioenergetics during fasting.

Dynamin-Related Protein 1 Deficiency Improves Mitochondrial Fitness and Protects against Progression of Diabetic Nephropathy.

Mitochondrial fission has been linked to the pathogenesis of diabetic nephropathy (DN). However, how mitochondrial fission affects progression of DN in vivo is unknown. Here, we report the effect of conditional podocyte-specific deletion of dynamin-related protein 1 (Drp1), an essential component of mitochondrial fission, on the pathogenesis and progression of DN. Inducible podocyte-specific deletion of Drp1 in diabetic mice decreased albuminuria and improved mesangial matrix expansion and podocyte morphology. Ultrastructure analysis revealed a significant increase in fragmented mitochondria in the podocytes of wild-type diabetic mice but a marked improvement in mitochondrial structure in Drp1-null podocytes of diabetic mice. When isolated from diabetic mice and cultured in high glucose, Drp1-null podocytes had more elongated mitochondria and better mitochondrial fitness associated with enhanced oxygen consumption and ATP production than wild-type podocytes. Furthermore, administration of a pharmacologic inhibitor of Drp1, Mdivi1, significantly blunted mitochondrial fission and rescued key pathologic features of DN in mice. Taken together, these results provide novel correlations between mitochondrial morphology and the progression of DN and point to Drp1 as a potential therapeutic target in DN.

Mitochondrial Dynamics Is a Distinguishing Feature of Skeletal Muscle Fiber Types and Regulates Organellar Compartmentalization

Skeletal muscle fibers differentiate into specific fiber types with distinct metabolic properties determined by their reliance on oxidative phosphorylation (OXPHOS). Using in vivo approaches, we find that OXPHOS-dependent fibers, compared to glycolytic fibers, contain elongated mitochondrial networks with higher fusion rates that are dependent on the mitofusins Mfn1 and Mfn2. Switching of a glycolytic fiber to an oxidative IIA type is associated with elongation of mitochondria, suggesting that mitochondrial fusion is linked to metabolic state. Furthermore, we reveal that mitochondrial proteins are compartmentalized to discrete domains centered around their nuclei of origin. The domain dimensions are dependent on fiber type and are regulated by the mitochondrial dynamics proteins Mfn1, Mfn2, and Mff. Our results indicate that mitochondrial dynamics is tailored to fiber type physiology and provides a rationale for the segmental defects characteristic of aged and diseased muscle fibers.

Abstract 1250: Role of mitochondria elongation in p53-induced cellular senescence

Abstract Various stresses such as oncogene activation, oxidative stress, and activation of tumor suppressors induce cellular senescence in both normal and cancer cells. This phenomenon, termed premature senescence or accelerated senescence, acts as an important barrier against tumorigenesis and is also considered to be a potential mechanism for controlling human cancers. Although mitochondria has been implicated in cancer, but the exact role of mitochondria in premature senescence has not been fully determined. Here, we studied how morphological and functional change in mitochondria involves in the onset of p53-induced premature senescence. Upon p53 expression in p53-defective cancer cells, extensive elongation of mitochondria network and functional defects in mitochondria were detected. Antioxidant NAC treatment indicates that mitochondria elongation induces increase the level of ROS, which is an important mediator for cellular senescence. Our results suggest that morphology change and subsequent dysfunction of mitochondria play a critical role in p53-induced premature senescence. Citation Format: Young Yeon Kim, Hye Jin Jee, Na Young Jeong, Jeanho Yun. Role of mitochondria elongation in p53-induced cellular senescence. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1250. doi:10.1158/1538-7445.AM2015-1250

MiR-27 regulates mitochondrial networks by directly targeting the mitochondrial fission factor.

Mitochondrial morphology is dynamically regulated by forming small, fragmented units or interconnected networks, and this is a pivotal process that is used to maintain mitochondrial homeostasis. Although dysregulation of mitochondrial dynamics is related to the pathogenesis of several human diseases, its molecular mechanism is not fully elucidated. In this study, we demonstrate the potential role of miR-27 in the regulation of mitochondrial dynamics. Mitochondrial fission factor (MFF) mRNA is a direct target of miR-27, whose ectopic expression decreases MFF expression through binding to its 3′-untranslated region. Expression of miR-27 results in the elongation of mitochondria as well as an increased mitochondrial membrane potential and mitochondrial ATP level. Our results suggest that miR-27 is a novel regulator affecting morphological mitochondrial changes by targeting MFF.