459240
antibody from Invitrogen Antibodies
Targeting: ATP5F1A
ATP5A, ATP5A1, ATP5AL2, ATPM, hATP1, OMR, ORM
Antibody data
- Antibody Data
- Antigen structure
- References [37]
- Comments [0]
- Validations
- Western blot [2]
- Immunocytochemistry [1]
- Other assay [17]
Submit
Validation data
Reference
Comment
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- Product number
- 459240 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- ATP5A1 Monoclonal Antibody (7H10BD4F9)
- Antibody type
- Monoclonal
- Antigen
- Other
- Reactivity
- Human, Mouse, Rat, Bovine
- Host
- Mouse
- Isotype
- IgG
- Antibody clone number
- 7H10BD4F9
- Vial size
- 100 µL
- Concentration
- 1.0 mg/mL
- Storage
- 4° C
Submitted references Cardiac disruption of SDHAF4-mediated mitochondrial complex II assembly promotes dilated cardiomyopathy.
Enhancement of anaerobic glycolysis - a role of PGC-1α4 in resistance exercise.
Combination of Anoectochilus roxburghii Polysaccharide and Exercise Ameliorates Diet-Induced Metabolic Disorders in Obese Mice.
Tetracyclines promote survival and fitness in mitochondrial disease models.
Inner mitochondrial membrane protein MPV17 mutant mice display increased myocardial injury after ischemia/reperfusion.
PPARδ Attenuates Alcohol-Mediated Insulin Resistance by Enhancing Fatty Acid-Induced Mitochondrial Uncoupling and Antioxidant Defense in Skeletal Muscle.
Increased Skeletal Muscle Fiber Cross-Sectional Area, Muscle Phenotype Shift, and Altered Insulin Signaling in Rat Hindlimb Muscles in a Prenatally Androgenized Rat Model for Polycystic Ovary Syndrome.
The Alzheimer's disease-associated C99 fragment of APP regulates cellular cholesterol trafficking.
Regulation of vitamin D system in skeletal muscle and resident myogenic stem cell during development, maturation, and ageing.
Single amino acid mutations in the Saccharomyces cerevisiae rhomboid peptidase, Pcp1p, alter mitochondrial morphology.
Effects of prolonged type 2 diabetes on mitochondrial function in cerebral blood vessels.
CerS6-Derived Sphingolipids Interact with Mff and Promote Mitochondrial Fragmentation in Obesity.
Metabolic Reprogramming in Astrocytes Distinguishes Region-Specific Neuronal Susceptibility in Huntington Mice.
Loss of the mitochondrial i-AAA protease YME1L leads to ocular dysfunction and spinal axonopathy.
Microglia pre-activation and neurodegeneration precipitate neuroinflammation without exacerbating tissue injury in experimental autoimmune encephalomyelitis.
L-Arabinose Elicits Gut-Derived Hydrogen Production and Ameliorates Metabolic Syndrome in C57BL/6J Mice on High-Fat-Diet.
Absence of TXNIP in Humans Leads to Lactic Acidosis and Low Serum Methionine Linked to Deficient Respiration on Pyruvate.
Dynamic association of human mRNP proteins with mitochondrial tRNAs in the cytosol.
Cardiac CaMKII activation promotes rapid translocation to its extra-dyadic targets.
MTSS1/Src family kinase dysregulation underlies multiple inherited ataxias.
Impaired Mitochondrial Respiration in Large Cerebral Arteries of Rats with Type 2 Diabetes.
Mutant desmin substantially perturbs mitochondrial morphology, function and maintenance in skeletal muscle tissue.
Control of mitochondrial function and cell growth by the atypical cadherin Fat1.
Deregulation of mitochondrial F1FO-ATP synthase via OSCP in Alzheimer's disease.
Two different pathogenic mechanisms, dying-back axonal neuropathy and pancreatic senescence, are present in the YG8R mouse model of Friedreich's ataxia.
CCN6 regulates mitochondrial function.
Loss of OMA1 delays neurodegeneration by preventing stress-induced OPA1 processing in mitochondria.
The mitochondrial function of the cerebral vasculature in insulin-resistant Zucker obese rats.
Differentiation-Associated Downregulation of Poly(ADP-Ribose) Polymerase-1 Expression in Myoblasts Serves to Increase Their Resistance to Oxidative Stress.
Dynamics of enhanced mitochondrial respiration in female compared with male rat cerebral arteries.
Sustained mitochondrial functioning in cerebral arteries after transient ischemic stress in the rat: a potential target for therapies.
α-Synuclein is localized to mitochondria-associated ER membranes.
Mitochondrial dynamics associated with oxygen-glucose deprivation in rat primary neuronal cultures.
Cytosolic p53 inhibits Parkin-mediated mitophagy and promotes mitochondrial dysfunction in the mouse heart.
Effect of moderate dietary restriction on visceral organ weight, hepatic oxygen consumption, and metabolic proteins associated with energy balance in mature pregnant beef cows.
Effects of SRC and STAT3 upon gap junctional, intercellular communication in lung cancer lines.
Influence of pregnancy in mid-to-late gestation on circulating metabolites, visceral organ mass, and abundance of proteins relating to energy metabolism in mature beef cows.
Wang X, Zhang X, Cao K, Zeng M, Fu X, Zheng A, Zhang F, Gao F, Zou X, Li H, Li M, Lv W, Xu J, Long J, Zang W, Chen J, Gao F, Ding J, Liu J, Feng Z
Nature communications 2022 Jul 8;13(1):3947
Nature communications 2022 Jul 8;13(1):3947
Enhancement of anaerobic glycolysis - a role of PGC-1α4 in resistance exercise.
Koh JH, Pataky MW, Dasari S, Klaus KA, Vuckovic I, Ruegsegger GN, Kumar AP, Robinson MM, Nair KS
Nature communications 2022 Apr 28;13(1):2324
Nature communications 2022 Apr 28;13(1):2324
Combination of Anoectochilus roxburghii Polysaccharide and Exercise Ameliorates Diet-Induced Metabolic Disorders in Obese Mice.
Chen C, Kang M, Wang Q, Liu W, Yang M, Liang S, Xiang Q, Han X, Tao J
Frontiers in nutrition 2021;8:735501
Frontiers in nutrition 2021;8:735501
Tetracyclines promote survival and fitness in mitochondrial disease models.
Perry EA, Bennett CF, Luo C, Balsa E, Jedrychowski M, O'Malley KE, Latorre-Muro P, Ladley RP, Reda K, Wright PM, Gygi SP, Myers AG, Puigserver P
Nature metabolism 2021 Jan;3(1):33-42
Nature metabolism 2021 Jan;3(1):33-42
Inner mitochondrial membrane protein MPV17 mutant mice display increased myocardial injury after ischemia/reperfusion.
Madungwe NB, Feng Y, Imam Aliagan A, Tombo N, Kaya F, Bopassa JC
American journal of translational research 2020;12(7):3412-3428
American journal of translational research 2020;12(7):3412-3428
PPARδ Attenuates Alcohol-Mediated Insulin Resistance by Enhancing Fatty Acid-Induced Mitochondrial Uncoupling and Antioxidant Defense in Skeletal Muscle.
Koh JH, Kim KH, Park SY, Kim YW, Kim JY
Frontiers in physiology 2020;11:749
Frontiers in physiology 2020;11:749
Increased Skeletal Muscle Fiber Cross-Sectional Area, Muscle Phenotype Shift, and Altered Insulin Signaling in Rat Hindlimb Muscles in a Prenatally Androgenized Rat Model for Polycystic Ovary Syndrome.
DeChick A, Hetz R, Lee J, Speelman DL
International journal of molecular sciences 2020 Oct 25;21(21)
International journal of molecular sciences 2020 Oct 25;21(21)
The Alzheimer's disease-associated C99 fragment of APP regulates cellular cholesterol trafficking.
Montesinos J, Pera M, Larrea D, Guardia-Laguarta C, Agrawal RR, Velasco KR, Yun TD, Stavrovskaya IG, Xu Y, Koo SY, Snead AM, Sproul AA, Area-Gomez E
The EMBO journal 2020 Oct 15;39(20):e103791
The EMBO journal 2020 Oct 15;39(20):e103791
Regulation of vitamin D system in skeletal muscle and resident myogenic stem cell during development, maturation, and ageing.
Srikuea R, Hirunsai M, Charoenphandhu N
Scientific reports 2020 May 19;10(1):8239
Scientific reports 2020 May 19;10(1):8239
Single amino acid mutations in the Saccharomyces cerevisiae rhomboid peptidase, Pcp1p, alter mitochondrial morphology.
Huddleston ME, Xiao N, Both AP, Gordon DM
Cell biology international 2020 Jan;44(1):200-215
Cell biology international 2020 Jan;44(1):200-215
Effects of prolonged type 2 diabetes on mitochondrial function in cerebral blood vessels.
Merdzo I, Rutkai I, Sure VNLR, Katakam PVG, Busija DW
American journal of physiology. Heart and circulatory physiology 2019 Nov 1;317(5):H1086-H1092
American journal of physiology. Heart and circulatory physiology 2019 Nov 1;317(5):H1086-H1092
CerS6-Derived Sphingolipids Interact with Mff and Promote Mitochondrial Fragmentation in Obesity.
Hammerschmidt P, Ostkotte D, Nolte H, Gerl MJ, Jais A, Brunner HL, Sprenger HG, Awazawa M, Nicholls HT, Turpin-Nolan SM, Langer T, Krüger M, Brügger B, Brüning JC
Cell 2019 May 30;177(6):1536-1552.e23
Cell 2019 May 30;177(6):1536-1552.e23
Metabolic Reprogramming in Astrocytes Distinguishes Region-Specific Neuronal Susceptibility in Huntington Mice.
Polyzos AA, Lee DY, Datta R, Hauser M, Budworth H, Holt A, Mihalik S, Goldschmidt P, Frankel K, Trego K, Bennett MJ, Vockley J, Xu K, Gratton E, McMurray CT
Cell metabolism 2019 Jun 4;29(6):1258-1273.e11
Cell metabolism 2019 Jun 4;29(6):1258-1273.e11
Loss of the mitochondrial i-AAA protease YME1L leads to ocular dysfunction and spinal axonopathy.
Sprenger HG, Wani G, Hesseling A, König T, Patron M, MacVicar T, Ahola S, Wai T, Barth E, Rugarli EI, Bergami M, Langer T
EMBO molecular medicine 2019 Jan;11(1)
EMBO molecular medicine 2019 Jan;11(1)
Microglia pre-activation and neurodegeneration precipitate neuroinflammation without exacerbating tissue injury in experimental autoimmune encephalomyelitis.
Wimmer I, Scharler C, Zrzavy T, Kadowaki T, Mödlagl V, Rojc K, Tröscher AR, Kitic M, Ueda S, Bradl M, Lassmann H
Acta neuropathologica communications 2019 Jan 31;7(1):14
Acta neuropathologica communications 2019 Jan 31;7(1):14
L-Arabinose Elicits Gut-Derived Hydrogen Production and Ameliorates Metabolic Syndrome in C57BL/6J Mice on High-Fat-Diet.
Zhao L, Wang Y, Zhang G, Zhang T, Lou J, Liu J
Nutrients 2019 Dec 13;11(12)
Nutrients 2019 Dec 13;11(12)
Absence of TXNIP in Humans Leads to Lactic Acidosis and Low Serum Methionine Linked to Deficient Respiration on Pyruvate.
Katsu-Jiménez Y, Vázquez-Calvo C, Maffezzini C, Halldin M, Peng X, Freyer C, Wredenberg A, Giménez-Cassina A, Wedell A, Arnér ESJ
Diabetes 2019 Apr;68(4):709-723
Diabetes 2019 Apr;68(4):709-723
Dynamic association of human mRNP proteins with mitochondrial tRNAs in the cytosol.
Jády BE, Ketele A, Kiss T
RNA (New York, N.Y.) 2018 Dec;24(12):1706-1720
RNA (New York, N.Y.) 2018 Dec;24(12):1706-1720
Cardiac CaMKII activation promotes rapid translocation to its extra-dyadic targets.
Wood BM, Simon M, Galice S, Alim CC, Ferrero M, Pinna NN, Bers DM, Bossuyt J
Journal of molecular and cellular cardiology 2018 Dec;125:18-28
Journal of molecular and cellular cardiology 2018 Dec;125:18-28
MTSS1/Src family kinase dysregulation underlies multiple inherited ataxias.
Brown AS, Meera P, Altindag B, Chopra R, Perkins EM, Paul S, Scoles DR, Tarapore E, Magri J, Huang H, Jackson M, Shakkottai VG, Otis TS, Pulst SM, Atwood SX, Oro AE
Proceedings of the National Academy of Sciences of the United States of America 2018 Dec 26;115(52):E12407-E12416
Proceedings of the National Academy of Sciences of the United States of America 2018 Dec 26;115(52):E12407-E12416
Impaired Mitochondrial Respiration in Large Cerebral Arteries of Rats with Type 2 Diabetes.
Merdzo I, Rutkai I, Sure VN, McNulty CA, Katakam PV, Busija DW
Journal of vascular research 2017;54(1):1-12
Journal of vascular research 2017;54(1):1-12
Mutant desmin substantially perturbs mitochondrial morphology, function and maintenance in skeletal muscle tissue.
Winter L, Wittig I, Peeva V, Eggers B, Heidler J, Chevessier F, Kley RA, Barkovits K, Strecker V, Berwanger C, Herrmann H, Marcus K, Kornblum C, Kunz WS, Schröder R, Clemen CS
Acta neuropathologica 2016 Sep;132(3):453-73
Acta neuropathologica 2016 Sep;132(3):453-73
Control of mitochondrial function and cell growth by the atypical cadherin Fat1.
Cao LL, Riascos-Bernal DF, Chinnasamy P, Dunaway CM, Hou R, Pujato MA, O'Rourke BP, Miskolci V, Guo L, Hodgson L, Fiser A, Sibinga NE
Nature 2016 Nov 24;539(7630):575-578
Nature 2016 Nov 24;539(7630):575-578
Deregulation of mitochondrial F1FO-ATP synthase via OSCP in Alzheimer's disease.
Beck SJ, Guo L, Phensy A, Tian J, Wang L, Tandon N, Gauba E, Lu L, Pascual JM, Kroener S, Du H
Nature communications 2016 May 6;7:11483
Nature communications 2016 May 6;7:11483
Two different pathogenic mechanisms, dying-back axonal neuropathy and pancreatic senescence, are present in the YG8R mouse model of Friedreich's ataxia.
Mollá B, Riveiro F, Bolinches-Amorós A, Muñoz-Lasso DC, Palau F, González-Cabo P
Disease models & mechanisms 2016 Jun 1;9(6):647-57
Disease models & mechanisms 2016 Jun 1;9(6):647-57
CCN6 regulates mitochondrial function.
Patra M, Mahata SK, Padhan DK, Sen M
Journal of cell science 2016 Jul 15;129(14):2841-51
Journal of cell science 2016 Jul 15;129(14):2841-51
Loss of OMA1 delays neurodegeneration by preventing stress-induced OPA1 processing in mitochondria.
Korwitz A, Merkwirth C, Richter-Dennerlein R, Tröder SE, Sprenger HG, Quirós PM, López-Otín C, Rugarli EI, Langer T
The Journal of cell biology 2016 Jan 18;212(2):157-66
The Journal of cell biology 2016 Jan 18;212(2):157-66
The mitochondrial function of the cerebral vasculature in insulin-resistant Zucker obese rats.
Merdzo I, Rutkai I, Tokes T, Sure VN, Katakam PV, Busija DW
American journal of physiology. Heart and circulatory physiology 2016 Apr 1;310(7):H830-8
American journal of physiology. Heart and circulatory physiology 2016 Apr 1;310(7):H830-8
Differentiation-Associated Downregulation of Poly(ADP-Ribose) Polymerase-1 Expression in Myoblasts Serves to Increase Their Resistance to Oxidative Stress.
Oláh G, Szczesny B, Brunyánszki A, López-García IA, Gerö D, Radák Z, Szabo C
PloS one 2015;10(7):e0134227
PloS one 2015;10(7):e0134227
Dynamics of enhanced mitochondrial respiration in female compared with male rat cerebral arteries.
Rutkai I, Dutta S, Katakam PV, Busija DW
American journal of physiology. Heart and circulatory physiology 2015 Nov;309(9):H1490-500
American journal of physiology. Heart and circulatory physiology 2015 Nov;309(9):H1490-500
Sustained mitochondrial functioning in cerebral arteries after transient ischemic stress in the rat: a potential target for therapies.
Rutkai I, Katakam PV, Dutta S, Busija DW
American journal of physiology. Heart and circulatory physiology 2014 Oct 1;307(7):H958-66
American journal of physiology. Heart and circulatory physiology 2014 Oct 1;307(7):H958-66
α-Synuclein is localized to mitochondria-associated ER membranes.
Guardia-Laguarta C, Area-Gomez E, Rüb C, Liu Y, Magrané J, Becker D, Voos W, Schon EA, Przedborski S
The Journal of neuroscience : the official journal of the Society for Neuroscience 2014 Jan 1;34(1):249-59
The Journal of neuroscience : the official journal of the Society for Neuroscience 2014 Jan 1;34(1):249-59
Mitochondrial dynamics associated with oxygen-glucose deprivation in rat primary neuronal cultures.
Wappler EA, Institoris A, Dutta S, Katakam PV, Busija DW
PloS one 2013;8(5):e63206
PloS one 2013;8(5):e63206
Cytosolic p53 inhibits Parkin-mediated mitophagy and promotes mitochondrial dysfunction in the mouse heart.
Hoshino A, Mita Y, Okawa Y, Ariyoshi M, Iwai-Kanai E, Ueyama T, Ikeda K, Ogata T, Matoba S
Nature communications 2013;4:2308
Nature communications 2013;4:2308
Effect of moderate dietary restriction on visceral organ weight, hepatic oxygen consumption, and metabolic proteins associated with energy balance in mature pregnant beef cows.
Wood KM, Awda BJ, Fitzsimmons C, Miller SP, McBride BW, Swanson KC
Journal of animal science 2013 Sep;91(9):4245-55
Journal of animal science 2013 Sep;91(9):4245-55
Effects of SRC and STAT3 upon gap junctional, intercellular communication in lung cancer lines.
Geletu M, Guy S, Raptis L
Anticancer research 2013 Oct;33(10):4401-10
Anticancer research 2013 Oct;33(10):4401-10
Influence of pregnancy in mid-to-late gestation on circulating metabolites, visceral organ mass, and abundance of proteins relating to energy metabolism in mature beef cows.
Wood KM, Awda BJ, Fitzsimmons C, Miller SP, McBride BW, Swanson KC
Journal of animal science 2013 Dec;91(12):5775-84
Journal of animal science 2013 Dec;91(12):5775-84
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Supportive validation
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- Knockdown of ATP5A was achieved by transfecting Hep G2 with ATP5A specific siRNAs (Silencer® select Product # s1769). Western blot analysis (Fig. a) was performed using membrane enriched extracts from the ATP5A knockdown cells (lane 3), non-specific scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with ATP5A1 Monoclonal Antibody (7H10BD4F9) (Product # 459240, 1µg/mL and Goat anti-Mouse IgG (H+L), Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:4000 dilution). Densitometric analysis of this western blot is shown in histogram (Fig. b). Decrease in signal upon siRNA mediated knock down confirms that antibody is specific to ATP5A.
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- Western blot was performed using Anti-ATP5A Mouse Monoclonal Antibody (Product # 459240) and a 54 kDa band corresponding to an isoform of ATP5A was observed in cell lines and tissues tested. Membrane enriched extracts (30 µg lysate) of Hep G2 (Lane 1), HeLa (Lane 2), Caco-2 (Lane 3), HCT 116 (Lane 4), Mouse Heart (Lane 5), Rat Heart (Lane 6), Mouse Thymus (Lane 7) and Mouse Lung (Lane 8) were electrophoresed using Novex® NuPAGE® 4-12% Bis-Tris Protein Gel (Product # NP0322BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (1 µg/mL) and detected by chemiluminescence with Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005). A 25 kDa band (*) corresponding to circulting tissue IgG was observed in mouse tissues.
Supportive validation
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- Bovine pulmonary artery endothelial cells: Mitochondria were labeled using an anti-OxPhos Complex V subunit a, mouse IgG2b monoclonal antibody (Product # 459240) prelabeled with the Zenon® Alexa Fluor® 555 Mouse IgG2b Labeling Kit (Product # Z-25205). Tubulin was detected with an anti-a-tubulin mouse IgG2b monoclonal antibody prelabeled with the Zenon® Alexa Fluor® 488 Mouse IgG2b Labeling Kit (Product # Z-25202), and The nucleus was stained with DAPI (Product # D1306, D3571, D21490).
Supportive validation
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- Figure 5 F1FO-ATP synthase deregulation in 5xFAD mouse synaptic mitochondrial. ( a ) Synaptic mitochondria from 5xFAD mice demonstrated an age-dependent decrease in their respiratory control ratio (RCR). Six nonTg and 5 5xFAD mice at 4 months old and 6 nonTg and 5 5xFAD mice at 9 months old were used. ( b ) Synaptic mitochondria from 5xFAD mice had an age-dependent decrease in ATP synthesis. Six nonTg and 6 5xFAD mice at 4 months old and those from six nonTg and seven 5xFAD mice at 9 months old were used in the experiments. ( c ) Synaptic mitochondria from 5xFAD mice demonstrated an early decrease in the F1FO-ATP synthase catalytic activity at 4 months old which was exacerbated at 9 months old. Five mice of each group at 4 months old and seven nonTg and nine 5xFAD mice at 9 months old were used in the experiment. ( d ) Age-dependent increase in oligomycin-insensitive respiration of synaptic mitochondria from 5xFAD mice. Six nonTg and five 5xFAD mice at 4 months old, and six nonTg and five 5xFAD mice at 9 months old were used in the experiments. ( e , f ) Decreased oligomycin sensitivity of synaptic mitochondria from 5xFAD mice at 4 ( e ) and 9 months old ( f ). All data are presented as percentage of the activity of the corresponding vehicle-treated mitochondrial fractions. Six nonTg and five 5xFAD mice at 4 months old, and seven nonTg and seven 5xFAD mice at 9 months old were used in the experiments. ( g , h ) Increased F1 dissociation in synaptic mitochondria from 5xFAD mic
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- Figure 6 OSCP overexpression ameliorates Abeta-induced mitochondrial dysfunction in mouse neurons. Mouse cortical neurons were exposed to 500 nM oligomeric Abeta1-42 for 24 h. ( a ) Attenuated oligomeric Abeta1-42-induced OSCP reduction by OSCP overexpression. Western blot images show OSCP expression level in mouse primary neurons which is representative of seven samples in each group. ( b ) Shows immunofluorescent staining of OSCP (green) in primary cultured neurons. COXIV (red) was used to determine the localization of OSCP in mitochondria. * P
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- Figure 8 OSCP overexpression ameliorates Abeta-induced mitochondrial dysfunction in human neurons. ( a ) Human neurons were exposed to 500 nM oligomeric Abeta1-42 for 4 days, then subjected to immunoblotting detection of OSCP levels. Abeta induced a significant reduction in OSCP expression level. The lower panel shows representative immunoblotting images. Tom40 was used as the loading control. n =4-5 samples of each group. ( b ) Co-immunoprecipitation of OSCP and Abeta in Abeta-treated human neurons. ( c ) Significantly increased OSCP expression in the OSCP overexpressing neurons. The lower panel shows representative immunoblotting images. Tom40 was used as the loading control. n =4 samples of each group. ( d ) Preserved ATP production by OSCP overexpression. * P
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- Fig 9 PARP-1 protein levels in mitochondrial and nuclear fractions of myoblasts and myotubes. Western blot analysis detected elevated expression of PARP-1 in myoblasts than in myotubes for both nuclear (A) and mitochondrial (B) fractions. Densitometric analysis of PARP-1 protein level in myoblast was set as 100%. Statistical analyses of n = 3 independent experiments were assessed; where ** indicates p
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- Fig. 2. Assessment of mitochondrial bioenergetics in neuronal tissues (brainstem, posterior columns, nerve roots and dorsal root ganglia). A quantitative western blot assay was developed to measure FXN (A), COXI and COXII (B), cytochrome c (C), ATP synthase (D) and TOM22 (E) in neuronal tissues [brainstem, posterior columns (PC), nerve roots and dorsal root ganglia (DRG)]. (A) Representative western blot of human FXN expression in YG8R mice in the four neuronal tissues used in the study. Human FXN was only detected in YG8R mice, in which the transgene was present, and not in wild type (WT; data not shown). Western blot results were quantified for each lane using Fujifilm's Multi-Gauge software. To allow for loading variation, values were normalized to the actin control. Final values were expressed as a ratio to the value of FXN expression in nerve roots. DRG and nerve roots (distal tissues) expressed less FXN than the posterior columns and brainstem (proximal tissues). Values are expressed as mean+-s.e.m.; *** P
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- Figure 6 Expression of the mitochondrial marker ATP synthase is higher in gastrocnemius muscle in PNA rats. Gastrocnemius ( A - C ), tibialis anterior ( D - F ), and soleus ( G - I ) skeletal muscles from 16-week-old female rats exposed to sesame oil ( n = 4-5) or testosterone ( n = 4-5) were cryosectioned at 20 um thickness prior to indirect immunolabeling for ATP synthase. Scale bars, 200 um. For each rat, 3 distinct fields per muscle sample were imaged and the fluorescence intensity quantified ( C , F , I ). Fluorescence intensity data were analyzed using a Mann-Whitney U test. Fluorescence intensity trended higher in muscles from PNA rats for gastrocnemius (# p = 0.0556), trended slightly lower in TA (# p = 0.0979), but was not significantly different in soleus muscle. Quartiles with the median are shown. ( J - L ) Protein expression was examined by Western blot and quantified by densitometric analysis. Samples of ( J ) GN, ( K ) TA, and ( L ) SOL muscles from rats exposed to sesame oil ( n = 8) or testosterone ( n = 8) were separated by SDS-PAGE and transferred to nitrocellulose prior to detection with antibodies to the alpha subunit of ATP synthase or GAPDH (top panels). Values for ATP synthase were normalized to GAPDH and analyzed using an unpaired t test or Mann-Whitney U test, which indicated significantly greater expression of ATP synthase in GN, but not TA or SOL, from PNA rats (bottom panels). Values shown are means with SD. ** p < 0.005.
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- Fig. 4 Citrate synthase and complex I activities and expression levels of single subunit proteins of respiratory complexes I-V in R349P desmin knock-in mice. a Determination of the citrate synthase activity by a spectrophotometric assay using identical amounts of total protein extracts. Note the marked activity reduction in homozygous mice. For this approach, soleus muscles obtained from five mice of each genotype (WT, HET, HOM) were pooled and subjected to four-time repeated measurements. Column chart shows mean values of these technical replicates. b Determination of complex I activity by a colorimetric assay normalized to muscle weight. Though this analysis, performed in duplicate on non-pooled samples derived from three animals per genotype, showed a reduction of complex I activity in homozygous mice, the data failed to reach statistical significance. An additional normalization of the obtained complex I values to the citrate synthase values resulted in similar activity levels for the three genotypes (data not shown). Column chart shows mean values and standard errors. c Immunoblotting using antibodies directed against specific complex I-V proteins revealed increased, decreased, and missing signals of complex I subunit proteins in heterozygous and homozygous mice, whereas the signal intensities of complex II-V subunit proteins showed no obvious differences. Desmin immunoblotting confirmed a markedly decreased level of mutant desmin in homozygous mice. In heteroz
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- Figure 5 Pcp1p mutants have altered steady-state levels for select adenosine triphosphate (ATP) synthase subunits . (A) Nonfunctional pcp1 alleles have reduced levels of ATP synthase alpha subunit. Proteins from isolated mitochondria were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) followed by immunoblot analysis using anti-ATPV alpha subunit and anti-Tom40 antibodies. Tom40p was used as a control to confirm equal loading of samples. (B) Densitometric analysis of the levels of ATP synthase alpha subunit relative to Tom40p and normalized to the PCP1 wild-type control. Data presented as the average +- standard deviation from four experiments. * P = 0.03 for Delta pcp1 and 0.05 for G315D). (C) Tim11p levels vary across the pcp1 mutants. Proteins isolated as described above were analyzed by immunoblot for Tim11p and anti-Tom40 levels. (D) Densitometric analysis of the level of Tim11p relative to Tom40p and normalized to the PCP1 wild-type strain. Data represent the average +- standard deviation from six experiments. * P = 0.008.
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- Figure 6 Nonfunctional pcp1 alleles have reduced adenosine triphosphate (ATP) synthase super-complexes formed . ATP synthase complex oligomerization and ATPase activity were measured following blue native-polyacrylamide gel electrophoresis (PAGE). (A) Representative immunoblot showing ATP synthase oligomerization status using ATP synthase alpha subunit antibodies. Mitochondrial proteins from the strains indicated were solubilized in buffer at a digitonin to protein ratio of 1.4:1 (g/g) and 0.7:1 (g/g). (B) The corresponding sodium dodecyl sulfate PAGE (SDS-PAGE) of Tom40p levels by immunoblot analysis for samples presented in (A) confirm equal loading. (C) Representative in-gel ATP synthase activity for the same pcp1 strains under the two solubilization conditions described above ( n = 3).