Journal: Life Science Alliance

Article Title: MIC26 and MIC27 cooperate to regulate cardiolipin levels and the landscape of OXPHOS complexes

doi: 10.26508/lsa.202000711

Figure Lengend Snippet: (A) Western blot showing the expression level of MIC26 or/and MIC27 in the stable cell lines of HAP1 WT expressing empty vector (ev) and DKO that contain either of the following constructs, ev (pMSCVpuro), pMSCVpuro-MIC26, pMSCVpuro-MIC27, and both pMSCVpuro-MIC26 and pMSCVpuro-MIC27 together. (B) Oxygen consumption rates (pmol O 2 /s), including basal respiration (Basal), proton leak, maximal respiration (Maximum) after uncoupling by FCCP, spare respiratory capacity (Spare capacity), non-mitochondrial respiration (Non-mitochondrial), and ATP production are shown for HAP1 WT+EV (empty vector) and DKO+EV, DKO+MIC26, DKO+MIC27, DKO+MIC26+MC27, and DKO+CRLS1. Data are normalized to basal respiration from HAP1 WT+EV and the mean ± SEM from three independent experiments is shown. Overexpression of both MIC26 and MIC27 in DKO cells significantly rescued respiration of DKO cells. * P -value ≤ 0.05, ** P -value ≤ 0.01 ( t test).

Article Snippet: CRLS1 ORF was obtained from Sino Biological (HG20234-U).

Techniques: Western Blot, Expressing, Stable Transfection, Plasmid Preparation, Construct, Over Expression

Journal: Life Science Alliance

Article Title: MIC26 and MIC27 cooperate to regulate cardiolipin levels and the landscape of OXPHOS complexes

doi: 10.26508/lsa.202000711

Figure Lengend Snippet: (A) Graph representing the levels of cardiolipin shown as arbitrary units normalized to mg of protein in each cell types show significant reduction in MIC26 KO and double knockout (DKO) cells. * P -value ≤ 0.05, ** P -value ≤ 0.01. t test was used for statistical analysis. (B) Graph showing the distribution of various cardiolipin species (arbitrary units normalized to mg of protein) in HAP1 WT, MIC26 KO, MIC27 KO, and DKO cells. (C) Blue-native gel electrophoresis for isolated mitochondria from HAP1 WT expressing empty vector (ev) and DKO cell lines that are stably expressing ev or MIC26 or MIC27 or MIC26 and MIC27 together or CRLS1 (cardiolipin synthase) were solubilized and blotted for antibodies specific for complex I (NDUFB4), complex III (UQCRC2), or complex IV (COX1V). The restoration of staining of respiratory chain (super) complexes compared with DKO (with ev) was found upon expression of MIC26 and MIC27 as well as CRLS1 in DKO cell lines. The part of the BN–PAGE stained with Coomassie is shown to represent the loading among the cell lines. (D) The scheme summarizing the phenotype occurring due to loss of MIC26 and MIC27 that show MIC26 and MIC27 are cooperatively required for the formation of crista junctions, maintenance of cardiolipin levels, and stability of respiratory chain (super) complexes and F 1 F O –ATP synthase. In addition, MIC26 and MIC27 are required for the assembly of F 1 F o –ATP synthase by facilitating the association of F 1 and F o part. Loss of MIC26 and MIC27 leads to impaired respiration.

Article Snippet: CRLS1 ORF was obtained from Sino Biological (HG20234-U).

Techniques: Double Knockout, Nucleic Acid Electrophoresis, Isolation, Expressing, Plasmid Preparation, Stable Transfection, Staining

Journal: Biology

Article Title: Linoleic Acid-Enriched Diet Increases Mitochondrial Tetralinoleoyl Cardiolipin, OXPHOS Protein Levels, and Uncoupling in Interscapular Brown Adipose Tissue during Diet-Induced Weight Gain

doi: 10.3390/biology12010009

Figure Lengend Snippet: Effect of LD and SO diet consumption on total CL, monolyso-CL, 72-carbon CL species, and gene expression of CL synthesis and remodeling enzymes in BAT. CL was measured using normal phase liquid chromatography coupled with electrospray ionization mass spectrometry and reported relative to quantified protein levels in the sample. ( a ) Total CL. ( b ) Total monolyso-CL (MLCL). ( c ) MLCL as a percent of the total MLCL + CL in each tissue sample. ( d ) Total mass:charge ( m / z ) 1448 (predominant acyl species L 4 CL) and ( e ) Am/z of 1448 as a percent of the total CL in each tissue sample. ( f ) The 72-carbon species of CL in each diet group expressed as a percent of the total 72C species with the major species of CL for each m / z value written above each set of bars. ( g ) Ratio between m / z 1448 and m / z 1456 (predominant acyl species O 4 CL). ( h ) Gene expression of cardiolipin synthase ( Crls1) and tafazzin ( Taz) , expressed as the relative fold change (2 −ΔΔct ). For all data, data are expressed as the mean + SEM. A two-sample t -test was used to determine significant differences between diet groups. An asterisk (*) designates p < 0.05, with p -values for < 0.10 reported in the figure. N = 5/group.

Article Snippet: Target gene expression was normalized to an endogenous control cyclophilin A (mouse PPIA; Mm02342430_g1); peroxisome proliferator-activated receptor γ (PPAR γ; Mm00440940_m1) well and cell death-inducing DNA fragmentation factor, alpha subunit-like ( Cidea ; Mm00423554_m1), peroxisome proliferator-activated receptor coactivator- 1α ( PGC-1α ; Mm01208835_m1), lipoprotein lipase ( Lpl ; Mm00434764_m1), uncoupling protein 1 ( Ucp1 ; Mm01244861_m1), Cardiolipin Synthase 1 ( Crls1 ; Mm01278100_m1), and taffazin ( Taz ; Mm00504978).

Techniques: Gene Expression, Normal Phase Liquid Chromatography, Mass Spectrometry

Journal: Biology of Sex Differences

Article Title: Spatiotemporal AMPKα2 deletion in mice induces cardiac dysfunction, fibrosis and cardiolipin remodeling associated with mitochondrial dysfunction in males only

doi: 10.1186/s13293-021-00394-z

Figure Lengend Snippet: Primers

Article Snippet: Cardiolipin synthase 1 , Crls1 , Cardiolipins biosynthesis , Mm00503002_m1.

Techniques: Binding Assay, Activation Assay

Journal: Biology of Sex Differences

Article Title: Spatiotemporal AMPKα2 deletion in mice induces cardiac dysfunction, fibrosis and cardiolipin remodeling associated with mitochondrial dysfunction in males only

doi: 10.1186/s13293-021-00394-z

Figure Lengend Snippet: Left ventricular cardiolipin profile in male and female mice 16 weeks after cardiac-specific Ampkα2 deletion. A Total cardiolipin content in male mice. B Total cardiolipin content in female mice. C Relative content of each identified cardiolipin species in male mice. D Relative content of each identified cardiolipin species in female mice. E Expression of genes related to cardiolipin biosynthesis and maturation: acyl-CoA synthetase long-chain family member 1 (Acsl1), CDP-diacylglycerol synthase 1 (Cds1), cardiolipin synthase 1 (Crls1), tafazzin (Taz), elongation of very long chain fatty acids-like 2 (Elovl2), elongation of long chain fatty acids (Elovl5), fatty acid desaturase 1 (Fads1) and fatty acid desaturase 2 (Fads2). ANOVA: µ p < 0.05, µµ p ≤ 0.01 for the sex effect; $ p < 0.05, $$ p < 0.01 for interaction; Post hoc Tukey test: * p < 0.05; ** p < 0.01, *** p < 0.001 Ampkα2 f/f vs Ampkα2ciKO (same sex); ¤ p < 0.05, ¤¤ p < 0.01 males vs females (same genotype)

Article Snippet: Cardiolipin synthase 1 , Crls1 , Cardiolipins biosynthesis , Mm00503002_m1.

Techniques: Expressing

Journal: Biology of Sex Differences

Article Title: Spatiotemporal AMPKα2 deletion in mice induces cardiac dysfunction, fibrosis and cardiolipin remodeling associated with mitochondrial dysfunction in males only

doi: 10.1186/s13293-021-00394-z

Figure Lengend Snippet: Cardiolipin profile and oxidative capacities 16 weeks after cardiac-specific Ampkα2 deletion in control (Sham) and ovariectomized (OVX) female mice. A Total cardiolipin content. B Relative content of major cardiolipin species. C Relative content of minor cardiolipin species. D Expression of genes related to cardiolipin biosynthesis and maturation: acyl-CoA synthetase long-chain family member 1 (Acsl1), CDP-diacylglycerol synthase 1 (Cds1), cardiolipin synthase 1 (Crls1), tafazzin (Taz), elongation of very long chain fatty acids-like 2 (Elovl2), elongation of long chain fatty acids (Elovl5), fatty acid desaturase 1 (Fads1) and fatty acid desaturase 2 (Fads2). E Rate of respiration after successive addition of pyruvate (1 mM), glutamate (10 mM), succinate (15 mM), amytal (1 mM). ( n = 4 to 7 per experimental group). ANOVA: # p ≤ 0.05 for the OVX effect; § p < 0.05 for the genotype effect, $ p < 0.05 interaction. Post hoc Tukey test: * p < 0.05, ** p < 0.01 CT-OVX vs KO-OVX

Article Snippet: Cardiolipin synthase 1 , Crls1 , Cardiolipins biosynthesis , Mm00503002_m1.

Techniques: Control, Expressing