Journal: Cells

Article Title: Cys340Ser Mutation Abolishing S-Nitrosylation Drives GRK2 Mitochondrial Localization and Dysfunction

doi: 10.3390/cells15050458

Figure Lengend Snippet: Group comparison of mitochondrial Drp1 ( A ) and Fis1 ( B ) expressions. Statistical significance was determined by ANOVA followed by Bonferroni’s post hoc test. Data are shown as mean ± SEM; ns, p = 0.2021 for Drp1; *, p < 0.05 ( p = 0.0041) for Fis1 ( n = 5–7 per group). Representative immunoblots demonstrating Drp1 and Fis1 protein levels in mitochondrial fractions and total cell lysates of AC16 cells infected with Ad.GFP or Ad.GRK2-C340S under normoxic or hypoxia/reoxygenation (H/R) conditions. VDAC served as a mitochondrial loading control and GAPDH served as a total protein marker. Densitometric analysis was performed using Li-Cor Image Studio software and normalized to the respective loading controls. All samples included in each comparative analysis were run on the same gel. Cropped blots are presented for clarity; uncropped blots with molecular weight markers are provided in .

Article Snippet: The primary antibodies used were as follows: GRK2 (13990-1-AP; ProteinTech, Rosemont, IL, USA), GRK2 (05-465; Sigma Aldrich, St. Louis, MO, USA), Drp1 (8570; Cell Signaling Technology, Danvers, MA, USA), Fis1 (32525; Cell Signaling Technology, Danvers, MA, USA), Mfn1 (14739; Cell Signaling Technology, Danvers, MA, USA), Mfn2 (11925; Cell Signaling Technology, Danvers, MA, USA), Opa1 (80471; Cell Signaling Technology), Pink (6946; Cell Signaling Technology, Danvers, MA, USA), Parkin (4211; Cell Signaling Technology, Danvers, MA, USA), LC3 (3868; Cell Signaling Technology, Danvers, MA, USA), Tom20 (42406; Cell Signaling Technology, Danvers, MA, USA), VDAC1 (sc-390996; Santa Cruz Biotechnology, Dallas, TX, USA), and GAPDH (sc-32233; Santa Cruz Santa Cruz Biotechnology, Dallas, TX, USA).

Techniques: Comparison, Western Blot, Infection, Control, Marker, Software, Molecular Weight