rabbit polyclonal anti fech  (Bioss)

Journal: Cell death & disease

Article Title: Oxidized-LDL inhibits testosterone biosynthesis by affecting mitochondrial function and the p38 MAPK/COX-2 signaling pathway in Leydig cells.

doi: 10.1038/s41419-020-02751-z

Figure Lengend Snippet: Fig. 4 oxLDL inhibits mitochondrial function in primary Leydig cells. Cells were treated with various concentrations of oxLDL or 200 μM H2O2 for 24 h, and then the mitochondrial membrane potential (MMP) (a, b) and intracellular ROS (c, d) were assayed by flow cytometry. H2O2 as a positive control. e Activities of mitochondrial complexes I, II, and III (CI, CII, and CIII) were assessed by spectrophotometry. f The expression of respiratory chain proteins, including Ndufs1 (a subunit of CI), SdhB (a subunit of CII), Uqcrfs1 (a subunit of CIII), Fech (a matrix enzyme ferrochelatase), and CytC (an intermembrane space protein cytochrome C), was detected by western blotting. Quantitation of mitochondrial proteins expression was determined by normalization to the internal control GAPDH. g The level of Intracellular ATP were detected. The vehicle with no oxLDL treatment was used as a control. The data are presented as the mean ± standard deviation of at least three independent experiments. *P < 0.05, **P < 0.01, compared with the control.

Article Snippet: Rabbit polyclonal anti-oxidized LDL (cat. no. ab14519), rabbit polyclonal anti-COX-2 (cat. no. ab15191), rabbit monoclonal anti-3β-HSD (cat. no. ab150384), mouse monoclonal anti-phospho-p38 (cat. no. ab45381), rabbit monoclonal anti-p38 (cat. no. ab170099), rabbit monoclonal anti-phospho-JNK (cat. no. ab124956), and rabbit monoclonal anti-JNK (cat. no. ab208035) were purchased from Abcam (Cambridge, MA, USA); rabbit monoclonal anti-phospho-ERK1/2 (cat. no. #4370) and rabbit monoclonal anti-ERK1/2 (cat. no. #4695) were purchased from Cell Signaling Technology (Beverly, MA, USA); rabbit polyclonal anti-STAR (cat. no. 12225-1-AP), rabbit polyclonal anti-p450scc (cat. no. 13363-1-AP), rabbit polyclonal anti-CD36 (cat. no. 18836-1-AP), rabbit polyclonal antiNdufs1 (cat. no. 12444-1-AP), rabbit polyclonal anti-SdhB (cat. no. 10620-1-AP), rabbit polyclonal anti-Uqcrfs1 (cat. no. 18443-1-AP), rabbit polyclonal anti- Fech (cat. no. 14466-1-AP), and rabbit polyclonal anti-CytC(cat. no. 10993-1-AP) were purchased from ProteinTech Group (Chicago, IL, USA); rabbit polyclonal anti-17β-HSD (cat. no. sc-32872) was purchased from Santa Cruz Biotechnology (Eugene, OR, USA); mouse monoclonal GAPDH (cat. no. KC-5G5) was purchased from KangChen Biotech (Shanghai, China); and horseradish peroxidase (HRP)-conjugated goat anti-rabbit IgG (H+ L) secondary antibody and HRP-conjugated goat anti-mouse IgG (H+ L) secondary antibody were purchased from Thermo Fisher Scientific, Inc. (Waltham, MA, USA).

Techniques: Membrane, Cytometry, Positive Control, Spectrophotometry, Expressing, Western Blot, Quantitation Assay, Control, Standard Deviation

Journal: Cell reports

Article Title: The mitochondrial carrier SFXN1 is critical for complex III integrity and cellular metabolism

doi: 10.1016/j.celrep.2021.108869

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: anti-FECH, rabbit polyclonal , Proteintech , Cat#14466-1-AP; RRID:AB_2231579.

Techniques: Bioprocessing, Recombinant, Labeling, Transfection, Activity Assay, Reverse Transcription, Bicinchoninic Acid Protein Assay, CyQUANT Assay, Proliferation Assay, Multiplex sample analysis, Control, Software

Journal: Cell metabolism

Article Title: A Single Adaptable Cochaperone-Scaffold Complex Delivers Nascent Iron-Sulfur Clusters to Mammalian Respiratory Chain Complexes I-III.

doi: 10.1016/j.cmet.2017.03.010

Figure Lengend Snippet: Figure 1. The HSC20-HSPA9-holo-ISCU Fe-S Transfer Complex Interacts with the Complex III Accessory Factor LYRM7 and with Fe-S Subunits of Respiratory Complexes I–III (A) Immunoprecipitation (IP) of endogenous LYRM7 from HEK293 mitochondrial matrix extracts (MMEs), followed by immunoblots (IBs) to HSC20, HSPA9, ISCU, and UQCRFS1. (B) IP of endogenous UQCRFS1 from HEK293 MMEs, followed by IBs to LYRM7, HSC20, HSPA9, and ISCU. (C) Proteins that interact with HSC20 were identified by mass spectrometry (MS) analysis after IP of endogenous HSC20. Some of the notable interacting proteins and relative numbers of peptides detected in the MS are shown (see also Table S1 for complete listing). (D) CRISPR/Cas9 HSC20-knockout (KO) HEK293 and HeLa cell mitochondrial lysates. Levels of HSPA9, LYRM7, UQCRFS1, mitochondrial transporters ABCB7 and mitoferrin 2 (MFRN2), and Fe-S scaffold protein NFU1 are also shown. TOM20 was used as loading control (crRNA NT CTRL is a scramble CRISPR RNA; crRNA HSC20 is the CRISPR RNA targeting exon 1 of HSC20). (E) Expression of UQCRFS1-FLAG in control and HSC20-KO cells, followed by anti-FLAG IP. (F) IPs of LYRM7-FLAG/MYC wild-type (WT) or the mutants, as indicated, expressed in HEK293 cells for 48 hr. The IP sample for the LYRM7LFK-AAA-F/M mutant was normalized as described in the STAR Methods to compensate for the reduced levels detected in the input. (G) In vitro pull-down assays of 35S-labeled HA-tagged LYRM7 wild-type or its mutants, as indicated, were performed in the presence of FLAG-tagged HSC20. (A, B, D, E, F, n = 5 biological replicates; G, n = 4 biological replicates). See also Figure S1 and Table S1.

Article Snippet: Rouault Epitope: YDLLEKNINIVRKRLNR See the STAR Methods Immunoblots Rabbit polyclonal anti-MFRN1 (Chen et al., 2009) N/A Rabbit polyclonal anti-FECH Proteintech 14466-1-AP, RRID: AB_2231579 Mouse monoclonal anti-TOM20 Santa Cruz Biotechnology SC-136211, RRID: AB_2207538 Chemicals, Peptides, and Recombinant Proteins Anti-FLAG M2 Affinity Gel Sigma-Aldrich Cat#A2220 3 X FLAG Peptide Sigma-Aldrich Cat#F4799 Sodium pyruvate solution Sigma-Aldrich Cat#S8636 Uridine Sigma-Aldrich Cat# U6381 Iron-55 Radionuclide, Ferric Chloride in 0.5M HCl Perkin-Elmer Cat#NEZ043002M (Continued on next page) e1 Cell Metabolism 25, 945–953.e1–e6, April 4, 2017

Techniques: Immunoprecipitation, Western Blot, Mass Spectrometry, CRISPR, Knock-Out, Control, Expressing, Mutagenesis, In Vitro, Labeling

Journal: Cell metabolism

Article Title: A Single Adaptable Cochaperone-Scaffold Complex Delivers Nascent Iron-Sulfur Clusters to Mammalian Respiratory Chain Complexes I-III.

doi: 10.1016/j.cmet.2017.03.010

Figure Lengend Snippet: Figure 4. Assembly of Respiratory Complexes I–III and Supercomplexes Was Abrogated by KO of HSC20, whereas KD of LYRM7 Solely Inhibited Complex III Formation (A) 55Fe radiolabeling of respiratory complexes was performed in HEK293 cells and in HSC20-KO HEK293 cells or in cells transfected with siRNAs to KD the expression of endogenous LYRM7 for 7 days. Control cells were crRNA NT CTRL for HSC20-KO cells (scramble non-targeting crRNA) and NT siRNAs for the LYRM7 KD cells. (Ft is ferritin). The lower panel shows IBs, which were performed to assess levels of HSC20, LYRM7, UQCRFS1, and SDHB in HSC20-KO cells and in LYRM7 KD cells (7 days KD). (B) Native IBs were performed to assess levels of respiratory complexes, including the Complex III subunits UQCRC2 and UQCRFS1, the CII subunit SDHA, the CIV subunit MTCO1, and the CI subunit NDUFS1. (C) Native IBs were performed after KD of LYRM7 for 3 or 6 days, as indicated. (D) SDS immunoblots were performed on samples loaded in (C). (E) CI and CIV in-gel activity assays were performed on samples equivalent to those loaded in (B) and (C); (A and B, n = 3 biological replicates; C–E, n = 5 biological replicates). See also Figure S3.

Article Snippet: Rouault Epitope: YDLLEKNINIVRKRLNR See the STAR Methods Immunoblots Rabbit polyclonal anti-MFRN1 (Chen et al., 2009) N/A Rabbit polyclonal anti-FECH Proteintech 14466-1-AP, RRID: AB_2231579 Mouse monoclonal anti-TOM20 Santa Cruz Biotechnology SC-136211, RRID: AB_2207538 Chemicals, Peptides, and Recombinant Proteins Anti-FLAG M2 Affinity Gel Sigma-Aldrich Cat#A2220 3 X FLAG Peptide Sigma-Aldrich Cat#F4799 Sodium pyruvate solution Sigma-Aldrich Cat#S8636 Uridine Sigma-Aldrich Cat# U6381 Iron-55 Radionuclide, Ferric Chloride in 0.5M HCl Perkin-Elmer Cat#NEZ043002M (Continued on next page) e1 Cell Metabolism 25, 945–953.e1–e6, April 4, 2017

Techniques: Radioactivity, Transfection, Expressing, Control, Western Blot, Activity Assay