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paper n a higd2a bioid2 ha  (Addgene inc)
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Paper N A Higd2a Bioid2 Ha, supplied by Addgene inc, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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2007 n a oligonucleotides higd1a forward  (ATCC)
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2007 N A Oligonucleotides Higd1a Forward, supplied by ATCC, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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rc209997 higd2a myc ddk  (OriGene)
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Figure 1. The Absence of Each HIGD Protein Affects Mitochondrial Respiratory Chain Performance (A) Immunoblot analysis of the steady-state levels of HIGD proteins and MRC subunits in whole-cell lysates of HEK293T WT, HIGD1A-KO (clones KO1–3), and <t>HIGD2A-KO</t> (clones KO1–3) lines reconstituted with a FLAG-tagged recombinant WT version of the KO gene or the empty vector (EV) pCMV6-A-Hygro. (B–D) Mitochondrial respiratory chain function in WT, HIGD-KO, and reconstituted cell lines. (B) Polarographic assessment of KCN-sensitive intact cell respiration. (C) Polarographic measurement in digitonin-permeabilized cells of KCN-sensitive oxygen consumption driven by succinate+G3P in the presence or absence of ADP and the uncoupler CCCP. Respiratory activity was calculated as nmol O2/min/106 cells. (D) Spectrophotometric measurement of CIII and CIV activities in whole-cell extracts. Enzyme activities were calculated as nmol cytochrome c reduced (for CIII) or oxidized (for CIV)/min/mg cellular protein and expressed as a fraction of WT values. Results are mean ± SD of 4 repetitions. (E) Immunoblot analysis of the levels of HIGD proteins and MRC complex subunits in cell lysates of the indicated cell lines. The signals were quantified and normalized by TOM20 levels. The values were expressed as a percentage relative to WT and presented as the mean ± SD of 3 independent experiments.
Rc209997 Higd2a Myc Ddk, supplied by OriGene, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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rc201223 cox7a2l myc ddk  (OriGene)
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OriGene rc201223 cox7a2l myc ddk
Figure 1. The Absence of Each HIGD Protein Affects Mitochondrial Respiratory Chain Performance (A) Immunoblot analysis of the steady-state levels of HIGD proteins and MRC subunits in whole-cell lysates of HEK293T WT, HIGD1A-KO (clones KO1–3), and <t>HIGD2A-KO</t> (clones KO1–3) lines reconstituted with a FLAG-tagged recombinant WT version of the KO gene or the empty vector (EV) pCMV6-A-Hygro. (B–D) Mitochondrial respiratory chain function in WT, HIGD-KO, and reconstituted cell lines. (B) Polarographic assessment of KCN-sensitive intact cell respiration. (C) Polarographic measurement in digitonin-permeabilized cells of KCN-sensitive oxygen consumption driven by succinate+G3P in the presence or absence of ADP and the uncoupler CCCP. Respiratory activity was calculated as nmol O2/min/106 cells. (D) Spectrophotometric measurement of CIII and CIV activities in whole-cell extracts. Enzyme activities were calculated as nmol cytochrome c reduced (for CIII) or oxidized (for CIV)/min/mg cellular protein and expressed as a fraction of WT values. Results are mean ± SD of 4 repetitions. (E) Immunoblot analysis of the levels of HIGD proteins and MRC complex subunits in cell lysates of the indicated cell lines. The signals were quantified and normalized by TOM20 levels. The values were expressed as a percentage relative to WT and presented as the mean ± SD of 3 independent experiments.
Rc201223 Cox7a2l Myc Ddk, supplied by OriGene, used in various techniques. Bioz Stars score: 92/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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rc201195 pcmv6 a entry hygro origene  (OriGene)
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Figure 1. The Absence of Each HIGD Protein Affects Mitochondrial Respiratory Chain Performance (A) Immunoblot analysis of the steady-state levels of HIGD proteins and MRC subunits in whole-cell lysates of HEK293T WT, HIGD1A-KO (clones KO1–3), and <t>HIGD2A-KO</t> (clones KO1–3) lines reconstituted with a FLAG-tagged recombinant WT version of the KO gene or the empty vector (EV) pCMV6-A-Hygro. (B–D) Mitochondrial respiratory chain function in WT, HIGD-KO, and reconstituted cell lines. (B) Polarographic assessment of KCN-sensitive intact cell respiration. (C) Polarographic measurement in digitonin-permeabilized cells of KCN-sensitive oxygen consumption driven by succinate+G3P in the presence or absence of ADP and the uncoupler CCCP. Respiratory activity was calculated as nmol O2/min/106 cells. (D) Spectrophotometric measurement of CIII and CIV activities in whole-cell extracts. Enzyme activities were calculated as nmol cytochrome c reduced (for CIII) or oxidized (for CIV)/min/mg cellular protein and expressed as a fraction of WT values. Results are mean ± SD of 4 repetitions. (E) Immunoblot analysis of the levels of HIGD proteins and MRC complex subunits in cell lysates of the indicated cell lines. The signals were quantified and normalized by TOM20 levels. The values were expressed as a percentage relative to WT and presented as the mean ± SD of 3 independent experiments.
Rc201195 Pcmv6 A Entry Hygro Origene, supplied by OriGene, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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rc202697 bcs1l myc ddk  (OriGene)
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Figure 1. The Absence of Each HIGD Protein Affects Mitochondrial Respiratory Chain Performance (A) Immunoblot analysis of the steady-state levels of HIGD proteins and MRC subunits in whole-cell lysates of HEK293T WT, HIGD1A-KO (clones KO1–3), and <t>HIGD2A-KO</t> (clones KO1–3) lines reconstituted with a FLAG-tagged recombinant WT version of the KO gene or the empty vector (EV) pCMV6-A-Hygro. (B–D) Mitochondrial respiratory chain function in WT, HIGD-KO, and reconstituted cell lines. (B) Polarographic assessment of KCN-sensitive intact cell respiration. (C) Polarographic measurement in digitonin-permeabilized cells of KCN-sensitive oxygen consumption driven by succinate+G3P in the presence or absence of ADP and the uncoupler CCCP. Respiratory activity was calculated as nmol O2/min/106 cells. (D) Spectrophotometric measurement of CIII and CIV activities in whole-cell extracts. Enzyme activities were calculated as nmol cytochrome c reduced (for CIII) or oxidized (for CIV)/min/mg cellular protein and expressed as a fraction of WT values. Results are mean ± SD of 4 repetitions. (E) Immunoblot analysis of the levels of HIGD proteins and MRC complex subunits in cell lysates of the indicated cell lines. The signals were quantified and normalized by TOM20 levels. The values were expressed as a percentage relative to WT and presented as the mean ± SD of 3 independent experiments.
Rc202697 Bcs1l Myc Ddk, supplied by OriGene, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Image Search Results


Figure 1. The Absence of Each HIGD Protein Affects Mitochondrial Respiratory Chain Performance (A) Immunoblot analysis of the steady-state levels of HIGD proteins and MRC subunits in whole-cell lysates of HEK293T WT, HIGD1A-KO (clones KO1–3), and HIGD2A-KO (clones KO1–3) lines reconstituted with a FLAG-tagged recombinant WT version of the KO gene or the empty vector (EV) pCMV6-A-Hygro. (B–D) Mitochondrial respiratory chain function in WT, HIGD-KO, and reconstituted cell lines. (B) Polarographic assessment of KCN-sensitive intact cell respiration. (C) Polarographic measurement in digitonin-permeabilized cells of KCN-sensitive oxygen consumption driven by succinate+G3P in the presence or absence of ADP and the uncoupler CCCP. Respiratory activity was calculated as nmol O2/min/106 cells. (D) Spectrophotometric measurement of CIII and CIV activities in whole-cell extracts. Enzyme activities were calculated as nmol cytochrome c reduced (for CIII) or oxidized (for CIV)/min/mg cellular protein and expressed as a fraction of WT values. Results are mean ± SD of 4 repetitions. (E) Immunoblot analysis of the levels of HIGD proteins and MRC complex subunits in cell lysates of the indicated cell lines. The signals were quantified and normalized by TOM20 levels. The values were expressed as a percentage relative to WT and presented as the mean ± SD of 3 independent experiments.

Journal: Cell reports

Article Title: Distinct Roles of Mitochondrial HIGD1A and HIGD2A in Respiratory Complex and Supercomplex Biogenesis.

doi: 10.1016/j.celrep.2020.107607

Figure Lengend Snippet: Figure 1. The Absence of Each HIGD Protein Affects Mitochondrial Respiratory Chain Performance (A) Immunoblot analysis of the steady-state levels of HIGD proteins and MRC subunits in whole-cell lysates of HEK293T WT, HIGD1A-KO (clones KO1–3), and HIGD2A-KO (clones KO1–3) lines reconstituted with a FLAG-tagged recombinant WT version of the KO gene or the empty vector (EV) pCMV6-A-Hygro. (B–D) Mitochondrial respiratory chain function in WT, HIGD-KO, and reconstituted cell lines. (B) Polarographic assessment of KCN-sensitive intact cell respiration. (C) Polarographic measurement in digitonin-permeabilized cells of KCN-sensitive oxygen consumption driven by succinate+G3P in the presence or absence of ADP and the uncoupler CCCP. Respiratory activity was calculated as nmol O2/min/106 cells. (D) Spectrophotometric measurement of CIII and CIV activities in whole-cell extracts. Enzyme activities were calculated as nmol cytochrome c reduced (for CIII) or oxidized (for CIV)/min/mg cellular protein and expressed as a fraction of WT values. Results are mean ± SD of 4 repetitions. (E) Immunoblot analysis of the levels of HIGD proteins and MRC complex subunits in cell lysates of the indicated cell lines. The signals were quantified and normalized by TOM20 levels. The values were expressed as a percentage relative to WT and presented as the mean ± SD of 3 independent experiments.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER siCOX11 Ambion Cat# oligo s3411 siMITRAC7 Invitrogen Cat# oligo SMIM20HSS158163 siCOX5B Dharmacon Cat# oligo L-013632-01-0005 siCOX6A1 Dharmacon Cat# oligo L-011836-01-0005 Recombinant DNA TAL Effector (F)-HIGD1A: TAAGCATGACAAATGTCAA Thermo-Invitrogen N/A TAL Effector (R)- HIGD1A: TAGGATGATTTCTGAGCCA Thermo-Invitrogen N/A TAL Effector (F)- HIGD2A: TTCTCCTGCTGCTGTGGCC Thermo-Invitrogen N/A TAL Effector (R)- HIGD2A: CCGGCCCTGTGATTCCGGA Thermo-Invitrogen N/A UQCR11 in pReceiver-B31 GeneCopoeia Cat# EX-I0287-B31 UQCRB in pReceiver-B31 GeneCopoeia Cat# EX-F0223-B31 UQCRFS1 or RISP in pReceiver-B31 GeneCopoeia Cat# EX-A3744-B31 MCS-BioID2-HA Addgene Cat# 74224 MTS-BioID2-HA This paper N/A HIGD2A-BioID2-HA This paper N/A HIGD1A- Myc-DDK in pCMV6-Entry Origene Cat# RC209997 HIGD2A- Myc-DDK in pCMV6-Entry Origene Cat# RC201223 COX7A2L- Myc-DDK in pCMV6-Entry Origene Cat# RC202697 BCS1L-Myc-DDK in pCMV6-Entry Origene Cat# RC201195 pCMV6-A-Entry-Hygro Origene Cat# PS100024 COX7A2L-Myc-DDK in pCMV6-A-Entry-Hygro This paper N/A HIGD1A-Myc-DDK in pCMV6-A-Entry-Hygro This paper N/A HIGD2A-Myc-DDK in pCMV6-A-Entry-Hygro This paper N/A BCS1L-Myc-DDK in pCMV6-A-Entry-Hygro This paper N/A HIGD2AR82A-Myc-DDK in pCMV6-A-Entry-Hygro This paper N/A HIGD2AR82AQ86A-Myc-DDK in pCMV6-A-Entry-Hygro This paper N/A HIGD2AQ86A-Myc-DDK in pCMV6-A-Entry-Hygro This paper N/A pCDF1-MCS2-EF1-Puro System Biosciences Cat# CD110B-1 pFIV-34N System Biosciences Cat# LV100A-1 pVSV-G System Biosciences Cat# LV100A-1 HIGD1A in pCDF1-MCS2-EF1-Puro This paper N/A Software and Algorithms SPSS IBM, v 21.0 N/A ImageJ NIH https://imagej.nih.gov/ij/ GraphPad Prism GraphPad Software v.5.0a N/A Scaffold Proteome Software, Inc N/A Other Pre-cast NativePAGE 4%–16% Bis-Tris gels Invitrogen Cat# BN1004BOX Pre-cast NativePAGE 3%–12% Bis-Tris gels Invitrogen Cat# BN2011BX10

Techniques: Western Blot, Clone Assay, Recombinant, Plasmid Preparation, Activity Assay

Figure 5. HIGD2A Associates with SCs to Modulate Supercomplexed CIV Assembly (A) coIP analysis of HIGD1A-FLAG-interacting proteins with anti-FLAG agarose beads (FLAG) or plain beads used as control (Ct), using digitonin-extracted mitochondria. (B) BN-PAGE analysis of the incorporation during 10 to 60 min of the radiolabeled HIGD2A into MRC components in mitochondria freshly isolated from the indicated cell lines. Import assays were performed in duplicates with similar results. (C) HIGD2A-containing complexes were analyzed by BN-PAGE and immunoblot in purified HEK293T mitochondria solubilized with digitonin. (D) BN-PAGE analysis in purified mitochondria prepared from 143B parental cells and COX1 and COX2 mutant cybrids in the presence of digitonin. (E) Model proposed for the mechanism of action of HIGD2A in CIV biogenesis. HIGD2A regulatory module, containing COX4-1, COX5B, and COX6A1 subunits, may coordinate the binding of the COX3 module to CIV but also the incorporation of CIV modules into SCs.

Journal: Cell reports

Article Title: Distinct Roles of Mitochondrial HIGD1A and HIGD2A in Respiratory Complex and Supercomplex Biogenesis.

doi: 10.1016/j.celrep.2020.107607

Figure Lengend Snippet: Figure 5. HIGD2A Associates with SCs to Modulate Supercomplexed CIV Assembly (A) coIP analysis of HIGD1A-FLAG-interacting proteins with anti-FLAG agarose beads (FLAG) or plain beads used as control (Ct), using digitonin-extracted mitochondria. (B) BN-PAGE analysis of the incorporation during 10 to 60 min of the radiolabeled HIGD2A into MRC components in mitochondria freshly isolated from the indicated cell lines. Import assays were performed in duplicates with similar results. (C) HIGD2A-containing complexes were analyzed by BN-PAGE and immunoblot in purified HEK293T mitochondria solubilized with digitonin. (D) BN-PAGE analysis in purified mitochondria prepared from 143B parental cells and COX1 and COX2 mutant cybrids in the presence of digitonin. (E) Model proposed for the mechanism of action of HIGD2A in CIV biogenesis. HIGD2A regulatory module, containing COX4-1, COX5B, and COX6A1 subunits, may coordinate the binding of the COX3 module to CIV but also the incorporation of CIV modules into SCs.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER siCOX11 Ambion Cat# oligo s3411 siMITRAC7 Invitrogen Cat# oligo SMIM20HSS158163 siCOX5B Dharmacon Cat# oligo L-013632-01-0005 siCOX6A1 Dharmacon Cat# oligo L-011836-01-0005 Recombinant DNA TAL Effector (F)-HIGD1A: TAAGCATGACAAATGTCAA Thermo-Invitrogen N/A TAL Effector (R)- HIGD1A: TAGGATGATTTCTGAGCCA Thermo-Invitrogen N/A TAL Effector (F)- HIGD2A: TTCTCCTGCTGCTGTGGCC Thermo-Invitrogen N/A TAL Effector (R)- HIGD2A: CCGGCCCTGTGATTCCGGA Thermo-Invitrogen N/A UQCR11 in pReceiver-B31 GeneCopoeia Cat# EX-I0287-B31 UQCRB in pReceiver-B31 GeneCopoeia Cat# EX-F0223-B31 UQCRFS1 or RISP in pReceiver-B31 GeneCopoeia Cat# EX-A3744-B31 MCS-BioID2-HA Addgene Cat# 74224 MTS-BioID2-HA This paper N/A HIGD2A-BioID2-HA This paper N/A HIGD1A- Myc-DDK in pCMV6-Entry Origene Cat# RC209997 HIGD2A- Myc-DDK in pCMV6-Entry Origene Cat# RC201223 COX7A2L- Myc-DDK in pCMV6-Entry Origene Cat# RC202697 BCS1L-Myc-DDK in pCMV6-Entry Origene Cat# RC201195 pCMV6-A-Entry-Hygro Origene Cat# PS100024 COX7A2L-Myc-DDK in pCMV6-A-Entry-Hygro This paper N/A HIGD1A-Myc-DDK in pCMV6-A-Entry-Hygro This paper N/A HIGD2A-Myc-DDK in pCMV6-A-Entry-Hygro This paper N/A BCS1L-Myc-DDK in pCMV6-A-Entry-Hygro This paper N/A HIGD2AR82A-Myc-DDK in pCMV6-A-Entry-Hygro This paper N/A HIGD2AR82AQ86A-Myc-DDK in pCMV6-A-Entry-Hygro This paper N/A HIGD2AQ86A-Myc-DDK in pCMV6-A-Entry-Hygro This paper N/A pCDF1-MCS2-EF1-Puro System Biosciences Cat# CD110B-1 pFIV-34N System Biosciences Cat# LV100A-1 pVSV-G System Biosciences Cat# LV100A-1 HIGD1A in pCDF1-MCS2-EF1-Puro This paper N/A Software and Algorithms SPSS IBM, v 21.0 N/A ImageJ NIH https://imagej.nih.gov/ij/ GraphPad Prism GraphPad Software v.5.0a N/A Scaffold Proteome Software, Inc N/A Other Pre-cast NativePAGE 4%–16% Bis-Tris gels Invitrogen Cat# BN1004BOX Pre-cast NativePAGE 3%–12% Bis-Tris gels Invitrogen Cat# BN2011BX10

Techniques: Control, Isolation, Western Blot, Mutagenesis, Binding Assay