Journal: eLife

Article Title: Identification of a weight loss-associated causal eQTL in MTIF3 and the effects of MTIF3 deficiency on human adipocyte function

doi: 10.7554/eLife.84168

Figure Lengend Snippet:

Article Snippet: Antibody , Anti-ATP8 (rabbit polyclonal antibody) , Proteintech , Cat: 26723-1-AP , WB (1:2000).

Techniques: Recombinant, Plasmid Preparation, Sequencing, TaqMan Assay, DNA Extraction, Isolation, Sample Prep, Picogreen Assay

Journal: bioRxiv

Article Title: MLQ is responsible for stabilisation of subunit a in the holoenzyme of mammalian ATP synthase

doi: 10.1101/2020.02.03.931709

Figure Lengend Snippet: Mitochondria isolated from cell lines were solubilised by digitonin 2 g/g protein, separated on clear-native PAGE (CNE) and either stained in gel for ATP hydrolytic activity or used for WB detection of F1-α, Fo-a, A6L, DAPIT and MLQ subunits. Individual forms of ATP synthase are indicated: cV M – monomer, cV D – dimer, cV o – oligomer, cV M * and cV D * denote smaller form of the respective complexes.

Article Snippet: Membranes were blocked in 5% non-fat milk or 3% BSA, dissolved in TBS (Tris-buffered saline containing 150 mM NaCl, 10 mM Tris-HCl, pH 7.5) for 1 h prior to incubations with primary (2 h at room temperature or overnight at 4 °C) antibodies to subunits of ATP synthase (F 1 -α ( ) lot 20D6, F 1 -β (Abcam ab14730), F 1 -γ (GTX 114275), F 1 - δ (GTX101503), F o -a , F o -b (Abcam 117991), F o -c (Abcam 181243), F o -d (GTX 87685), F o -g (GTX 111014), A6L (Biorbyt orb215488), OSCP (Santa Cruz SC-98707), DAPIT (Proteintech 17716-1-AP), MLQ (Proteintech 14704-1-AP), Complex II (SDHA, Abcam 14715), AFG3L2 (Proteintech 14631-1-AP), OPA1 (BD Bio 612606), and actin (Calbiochem CPO1-1EA).

Techniques: Isolation, Clear Native PAGE, Staining, Activity Assay

Journal: bioRxiv

Article Title: MLQ is responsible for stabilisation of subunit a in the holoenzyme of mammalian ATP synthase

doi: 10.1101/2020.02.03.931709

Figure Lengend Snippet: Mitochondria isolated from the respective cell lines were solubilised by digitonin 2 g/g and separated on blue-native PAGE (BNE). WB detection of F1-α, Fo-a, A6L, DAPIT and MLQ subunits. ATP synthase forms: cV M – monomer, cV D – dimer, F 1 +c − F 1 and c subunits and F 1 subcomplexes.

Article Snippet: Membranes were blocked in 5% non-fat milk or 3% BSA, dissolved in TBS (Tris-buffered saline containing 150 mM NaCl, 10 mM Tris-HCl, pH 7.5) for 1 h prior to incubations with primary (2 h at room temperature or overnight at 4 °C) antibodies to subunits of ATP synthase (F 1 -α ( ) lot 20D6, F 1 -β (Abcam ab14730), F 1 -γ (GTX 114275), F 1 - δ (GTX101503), F o -a , F o -b (Abcam 117991), F o -c (Abcam 181243), F o -d (GTX 87685), F o -g (GTX 111014), A6L (Biorbyt orb215488), OSCP (Santa Cruz SC-98707), DAPIT (Proteintech 17716-1-AP), MLQ (Proteintech 14704-1-AP), Complex II (SDHA, Abcam 14715), AFG3L2 (Proteintech 14631-1-AP), OPA1 (BD Bio 612606), and actin (Calbiochem CPO1-1EA).

Techniques: Isolation, Blue Native PAGE

Journal: bioRxiv

Article Title: MLQ is responsible for stabilisation of subunit a in the holoenzyme of mammalian ATP synthase

doi: 10.1101/2020.02.03.931709

Figure Lengend Snippet: (A) MLQ KO and control cells were radioactively labelled for 3h with 35 S-methionin/cysteine and subsequently chased with cold methionine for 1, 2, and 3 days. Cells were harvested, solubilised with triton X-100 and ATP synthase was isolated by immune-affinity purification using ATP synthase immunocapture kit. IP-ATP synthase was than analysed by SDS-PAGE. Quantification of the radioactive signal of all ATP synthase subunits shows uniform kinetics of degradation in control HEK cells (B) , while (C) MLQ KO cells displayed faster turnover of subunits F o -a and A6L (red lines) but stabilization of the F 1 subunits (α and β, green lines) compared to subunits b, d and OSCP of the external stalk (blue line). Similar stabilisation of F 1 subunits show ΔTA cells ( E ) compared to control cybrids ( D ) as well as rho 0 cells ( G ) compared to control rho + cells ( F ).

Article Snippet: Membranes were blocked in 5% non-fat milk or 3% BSA, dissolved in TBS (Tris-buffered saline containing 150 mM NaCl, 10 mM Tris-HCl, pH 7.5) for 1 h prior to incubations with primary (2 h at room temperature or overnight at 4 °C) antibodies to subunits of ATP synthase (F 1 -α ( ) lot 20D6, F 1 -β (Abcam ab14730), F 1 -γ (GTX 114275), F 1 - δ (GTX101503), F o -a , F o -b (Abcam 117991), F o -c (Abcam 181243), F o -d (GTX 87685), F o -g (GTX 111014), A6L (Biorbyt orb215488), OSCP (Santa Cruz SC-98707), DAPIT (Proteintech 17716-1-AP), MLQ (Proteintech 14704-1-AP), Complex II (SDHA, Abcam 14715), AFG3L2 (Proteintech 14631-1-AP), OPA1 (BD Bio 612606), and actin (Calbiochem CPO1-1EA).

Techniques: Control, Isolation, Affinity Purification, SDS Page

Journal: PLoS ONE

Article Title: Kidney-Specific Reduction of Oxidative Phosphorylation Genes Derived from Spontaneously Hypertensive Rat

doi: 10.1371/journal.pone.0136441

Figure Lengend Snippet: A: mt-ND1 was reduced ~3.7 fold in HT BN/SHR-mt SHR (*P<0.05). mt- ND3 was reduced ~2.6 fold in HT SHR/BN-mt SHR (**P<0.01). mt-ND4 was reduced ~10.8 fold in HT SHR/BN-mt SHR (*P<0.05). mt-ND4L was reduced ~7.7 fold in HT SHR/BN-mt SHR (P<0.05). mt-ND5 was reduced ~2.7 fold in HT SHR/BN-mt SHR (P<0.05). mt-ND6 was reduced ~1.7 fold in HT SHR/BN-mt SHR (P>0.05) . mt-ND2 was not different between the two phenotypes (P>0.05). B: mt-CYB was reduced ~5 fold in HT BN/SHR-mt SHR (*P<0.05). C: mt-CO1 was reduced ~3.2 fold in HT BN/SHR-mt SHR (*P<0.05), mt-CO2 was reduced ~3.6 fold in HT BN/SHR-mt SHR (P<0.05), and mt-CO3 was reduced 4.1 fold in HT BN/SHR-mt SHR (P<0.05). D: mt-ATP6 was reduced ~2.3 fold in HT BN/SHR—mt SHR (*P<0.05), while mt-ATP8 was reduced ~3.1 fold in HT BN/SHR-mt SHR (*P<0.05). (NT: open bars; HT: closed bars).

Article Snippet: Pre-designed TaqMan primers and hydrolysis probes for all genes of interest were purchased from Applied Biosystems ( mt-ND1 - Rn03296764_s1, mt-ND2 - Rn03296765_s1, mt-ND3 - Rn03296825_s1, mt-ND4 - Rn03296781_s1, mt-ND4L - Rn03296792_s1, mt-ND5 - Rn03296799_s1, mt-ND6 - Rn03296815_s1, mt-CO1 - Rn03296721_s1, mt-CO2 - Rn03296737_s1, mt-CO3 - Rn03296820_s1, mt-CYB - Rn03296746_s1, mt-ATP6 - Rn03296710_s1, mt-ATP8 - Rn03296716_s1, NRF1 - Rn01455958_m1, NRF2a - Rn01767215_m1, NRF2b -Rn01514289_g1, Pgc1α -Rn00598552_m1, Tfam -Rn00580051_m1, Cyc1 -Rn01504159_g1, Cox6c -Rn00820983_gH, GAPDH - Rn01775763_g1).

Techniques: