Journal: Frontiers in pharmacology

Article Title: Cajaninstilbene Acid Ameliorates Acetaminophen-Induced Liver Injury Through Enhancing Sestrin2/AMPK-Mediated Mitochondrial Quality Control.

doi: 10.3389/fphar.2022.824138

Figure Lengend Snippet: FIGURE 7 | APAP-induced Sestrin2/AMPK activation is enhanced in the liver of mice treated with cajaninstilbene acid (CSA). (A–C) Animal experiment was identical to that in Figure 1, and liver tissues were collected at 6 h after APAP overdose. Immunoblot analysis of phosphorylated (p-) or total protein in lysates of total liver extracts. The graphs show results of densitometric analyses of p-AMPK relative to total AMPK, CAMKK2, Sestrin2, LKB1 relative to GAPDH respectively (n = 6/group). (D,E) Compound C (10 mg/kg) was injected to mice 0.5 h prior to APAP, followed by CSA (75 mg/kg) administration. Liver tissues were collected at 6 h after APAP overdose. (D) Immunoblot analysis of whole liver homogenate. The graphs show results of densitometric analyses of p-JNK relative to total JNK, PGC-1α, TOM20 relative to GAPDH respectively. (E) Immunoblot analysis of mitochondrial LC3 II and the corresponding densitometry normalized to VDAC1. *p < 0.05, **p < 0.001, ***p < 0.001 compared with the vehicle control; #p < 0.05, ##p < 0.01, ###p < 0.001 compared with the corresponding APAP-treated group. N.S, nonsignificant.

Article Snippet: The membranes were blocked with 5% nonfat milk in TBST buffer for 1 h at room temperature and incubated with primary antibodies specific for STING (ABclonal Cat# A3575, RRID:AB_2765161), p-IκBα (ABclonal Cat# AP0707, RRID:AB_2863811), IκBα (ABclonal Cat# A11397, RRID:AB_2861556), NLRP3 (Cell Signaling Technology Cat# 15101, RRID:AB_2722591), p-p65 (Cell Signaling Technology Cat# 3033, RRID:AB_331284), p65 (Proteintech Cat# 10745-1-AP, RRID:AB_2178878), CYP2E1 (ABclonal Cat# A2160, RRID:AB_2764178), p-JNK (Cell Signaling Technology Cat# 9251, RRID:AB_331659), JNK (ABclonal Cat# A4867, RRID:AB_2863367), 3-Nitrotyrosine (Abcam Cat# ab53232, RRID:AB_869927), PGC-1α (Abcam Cat# ab54481, RRID:AB_881987), TFAM (Abcam, Cat# ab252432), PINK1 (ABclonal Cat# A7131, RRID:AB_2767686), LC3 (ABclonal Cat# A19665, RRID:AB_2862723), p62 (Proteintech Cat# 18420-1-AP, RRID:AB_10694431), Parkin (ABclonal Cat# A0968, RRID:AB_2757487), p-AMPKα (Cell Signaling Technology Cat# 2535, RRID:AB_331250), AMPKα (Abcam, ab207442), CAMKK2 (Proteintech Cat# 11549-1-AP, RRID:AB_2259441), Sestrin2 (Proteintech Cat# 10795-1-AP, RRID:AB_2185480), LKB1 (ABclonal Cat# A2122, RRID: AB_2764141), LC3B (Cell Signaling Technology Cat# 3868, RRID:AB_2137707), VDAC1 (Proteintech Cat# 55259-1-AP, RRID:AB_10837225) and GAPDH (ABclonal Cat# AC001, RRID:AB_2619673) at 4°C shaking overnight.

Techniques: Activation Assay, Western Blot, Injection, Control

Journal: Frontiers in pharmacology

Article Title: Cajaninstilbene Acid Ameliorates Acetaminophen-Induced Liver Injury Through Enhancing Sestrin2/AMPK-Mediated Mitochondrial Quality Control.

doi: 10.3389/fphar.2022.824138

Figure Lengend Snippet: FIGURE 8 | Schematic illustration of the mechanisms underlying therapeutic effect of cajaninstilbene acid (CSA) against APAP-induced liver injury. APAP overdose causes accumulation of N-acetyl-p-benzoquinone imine (NAPQI) predominantly via CYP2E1. Excess NAPQI depletes GSH, leading to the sustained activation of JNK- Sab-ROS loop and mitochondrial dysfunction, characterized by a loss of efficiency in the electron transport chain (ETC) and reductions in the synthesis of ATP. This ultimately results into hepatocyte cell death, which triggers the innate immune system. To maintain intracellular homeostasis, a highly conserved stress-inducible metabolic protein Sestrin2 is activated to stimulate AMPK-mediated mitochondrial quality control and repress ROS. CSA upregulated Sestrin2/AMPK signaling pathway, leading to enhanced mitochondrial quality control, thereby protects against APAP-induced liver injury.

Article Snippet: The membranes were blocked with 5% nonfat milk in TBST buffer for 1 h at room temperature and incubated with primary antibodies specific for STING (ABclonal Cat# A3575, RRID:AB_2765161), p-IκBα (ABclonal Cat# AP0707, RRID:AB_2863811), IκBα (ABclonal Cat# A11397, RRID:AB_2861556), NLRP3 (Cell Signaling Technology Cat# 15101, RRID:AB_2722591), p-p65 (Cell Signaling Technology Cat# 3033, RRID:AB_331284), p65 (Proteintech Cat# 10745-1-AP, RRID:AB_2178878), CYP2E1 (ABclonal Cat# A2160, RRID:AB_2764178), p-JNK (Cell Signaling Technology Cat# 9251, RRID:AB_331659), JNK (ABclonal Cat# A4867, RRID:AB_2863367), 3-Nitrotyrosine (Abcam Cat# ab53232, RRID:AB_869927), PGC-1α (Abcam Cat# ab54481, RRID:AB_881987), TFAM (Abcam, Cat# ab252432), PINK1 (ABclonal Cat# A7131, RRID:AB_2767686), LC3 (ABclonal Cat# A19665, RRID:AB_2862723), p62 (Proteintech Cat# 18420-1-AP, RRID:AB_10694431), Parkin (ABclonal Cat# A0968, RRID:AB_2757487), p-AMPKα (Cell Signaling Technology Cat# 2535, RRID:AB_331250), AMPKα (Abcam, ab207442), CAMKK2 (Proteintech Cat# 11549-1-AP, RRID:AB_2259441), Sestrin2 (Proteintech Cat# 10795-1-AP, RRID:AB_2185480), LKB1 (ABclonal Cat# A2122, RRID: AB_2764141), LC3B (Cell Signaling Technology Cat# 3868, RRID:AB_2137707), VDAC1 (Proteintech Cat# 55259-1-AP, RRID:AB_10837225) and GAPDH (ABclonal Cat# AC001, RRID:AB_2619673) at 4°C shaking overnight.

Techniques: Activation Assay, Control

Journal: iScience

Article Title: SESN2 maintains cartilage homeostasis by SREBP1-mediated lipid metabolism during osteoarthritis progression

doi: 10.1016/j.isci.2025.113097

Figure Lengend Snippet: SESN2 deficiency is correlated with the upregulation of lipogenic enzymes in OA cartilage (A) Safranin O-Fast Green (S.O.) and Alcian blue staining of intact (I) and damaged (D) articular cartilages from OA patients. (B) Immunohistochemical (IHC) staining of matrix metallopeptidase 13 (MMP13) and type II collagen gene α (COL2A1) in human OA cartilages. (C and D) IHC staining and quantitative analysis for SESN2 in intact (I) and damaged (D) human OA cartilages ( n = 6). (E) Immunofluorescence (IF) staining for SESN2 expression in the cartilage of sham or destabilization of medial meniscus (DMM)-induced mice for 8 weeks ( n = 6). (F) Indicated age mice S.O. staining and IF staining of knee joints from mice at indicated ages (8, 16, 28 weeks, 1 year) ( n = 3). (G) Quantitative analysis for SESN2 expression in the cartilage of indicated treatment mice ( n = 6) and indicated age mice ( n = 3). (H and I) IHC staining (H) and quantitative analysis (I) for SESN2 in intact (I) and damaged (D) human OA cartilages ( n = 6). (J and K) Relative mRNA expression (qPCR) (J) and correlation analysis quantification (K) of indicated proteins genes (SESN2 vs. MMP3, MMP13, FASN, SCD1) in paired intact (I) and damaged (D) ( n = 8) human OA cartilages. Scale bars, (B, C, H, and F) 25 and 100 μm, (E) 50 μm, (A) 200 μm. Data are shown as mean ± SD. ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001. Statistical analysis was performed by unpaired t test (D, G [left], I, and J), one-way ANOVA (G, right), and Pearson correlation analysis (K).

Article Snippet: Slides were incubated overnight at 4°C with primary antibodies: SESN2 (Proteintech, #66297-1-Ig, 1:200), FASN (Proteintech, #66591-1-Ig, 1:100), SCD1 (ABclonal, #A16429, 1:100), SREBP1 (Proteintech, #14088-1-AP, 1:200), PGC-1α (Abcam, #ab54481, 1:100).

Techniques: Staining, Immunohistochemical staining, Immunohistochemistry, Immunofluorescence, Expressing

Journal: iScience

Article Title: SESN2 maintains cartilage homeostasis by SREBP1-mediated lipid metabolism during osteoarthritis progression

doi: 10.1016/j.isci.2025.113097

Figure Lengend Snippet: SESN2 is involved in maintaining cartilage homeostasis (A and B) qPCR (A) and western blot (WB) (B) analysis of Sesn2 in mouse primary chondrocytes pretreated with si-nc or si- Sesn2 ( n = 3). (C) qPCR analysis of indicated genes in mouse primary chondrocytes induced by 10 ng/mL IL-1β and/or pretreated with si-nc or si- Sesn2 for 48 h ( n = 3). (D and E) WB analysis (D) and quantitation (E) of indicated proteins in mouse primary chondrocytes induced by 10 ng/mL IL-1β and/or pretreated with si-nc or si- Sesn2 for 48 h ( n = 3). (F and G) qPCR (F) and WB (G) analysis of Sesn2 in chondrocytes infected with lv-nc or lv- Sesn2 at the indicated multiplicity of infection (MOI) ( n = 3). (H) qPCR analysis of indicated genes in mouse primary chondrocytes induced by 10 ng/mL IL-1β and/or infected with lv-nc or lv- Sesn2 for 48 h ( n = 3). (I and J) WB analysis (I) and quantitation (J) of indicated proteins in mouse primary chondrocytes induced by 10 ng/mL IL-1β and/or infected with lv-nc or lv- Sesn2 for 48 h ( n = 3). Data are shown as mean ± SD. ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001. Statistical analysis was performed by unpaired t test (A and F) and one-way ANOVA (C, E, H, and J).

Article Snippet: Slides were incubated overnight at 4°C with primary antibodies: SESN2 (Proteintech, #66297-1-Ig, 1:200), FASN (Proteintech, #66591-1-Ig, 1:100), SCD1 (ABclonal, #A16429, 1:100), SREBP1 (Proteintech, #14088-1-AP, 1:200), PGC-1α (Abcam, #ab54481, 1:100).

Techniques: Western Blot, Quantitation Assay, Infection

Journal: iScience

Article Title: SESN2 maintains cartilage homeostasis by SREBP1-mediated lipid metabolism during osteoarthritis progression

doi: 10.1016/j.isci.2025.113097

Figure Lengend Snippet: SESN2 improves fatty acid metabolism disorder via the downregulation of lipogenic enzymes (A) qPCR analysis of indicated genes in mouse primary chondrocytes pretreated with si-nc or si- Sesn2 ( n = 3). (B and C) Western blot analysis (B) and quantitation (C) of mouse primary chondrocytes pretreated with si-nc or si- Sesn2 ( n = 3). (D) BODIPY493/503 staining (left) and quantitative analysis (right) of mouse primary chondrocytes pretreated with si-nc or si- Sesn2 ( n = 6). (E) Cellular level of FFA of mouse primary chondrocytes pretreated with si-nc or si- Sesn2 ( n = 3). (F) qPCR analysis of indicated genes in mouse primary chondrocytes infected with lv-nc or lv- Sesn2 for 48 h ( n = 3). (G and H) Western blot analysis (G) and quantitation (H) of indicated proteins in mouse primary chondrocytes infected with lv-nc or lv- Sesn2 for 48 h ( n = 3). (I) BODIPY493/503 staining (left) and quantitative analysis (right) of mouse primary chondrocytes induced by 10 ng/mL IL-1β and infected with lv-nc or lv- Sesn2 for 48 h ( n = 6). (J) Cellular level of FFA of mouse primary chondrocytes induced by 10 ng/mL IL-1β and infected with lv-nc or lv- Sesn2 for 48 h ( n = 3). Data are shown as mean ± SD. ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001. Statistical analysis was performed by unpaired t test (A, C, D, E, F, H, I, and J).

Article Snippet: Slides were incubated overnight at 4°C with primary antibodies: SESN2 (Proteintech, #66297-1-Ig, 1:200), FASN (Proteintech, #66591-1-Ig, 1:100), SCD1 (ABclonal, #A16429, 1:100), SREBP1 (Proteintech, #14088-1-AP, 1:200), PGC-1α (Abcam, #ab54481, 1:100).

Techniques: Western Blot, Quantitation Assay, Staining, Infection

Journal: iScience

Article Title: SESN2 maintains cartilage homeostasis by SREBP1-mediated lipid metabolism during osteoarthritis progression

doi: 10.1016/j.isci.2025.113097

Figure Lengend Snippet: SESN2 improves fatty acid metabolism disorders by inhibiting SREBP1 (A) qPCR analysis of Srebp1 in mouse primary chondrocytes pretreated with si-nc or si- Sesn2 ( n = 3). (B) Western blot analysis (left) and quantitation (right) of SREBP1 in mouse primary chondrocytes pretreated with si-nc or si- Sesn2 ( n = 3). (C) IF analysis (left) and quantitation (right) of SREBP1 in mouse primary chondrocytes pretreated with si-nc or si- Sesn2 ( n = 6). (D) SESN2 and SCAP Molecular Docking Diagrams (HDOCK Server). (E) Immunoblot analysis of mouse primary chondrocytes, followed by immunoprecipitation with anti-SESN2 and anti-SCAP beads, probed with indicated antibodies. (F) qPCR analysis of indicated genes in mouse primary chondrocytes treated as indicated for 48 h. (G and H) Western blot analysis (G) and quantitation (H) of indicated proteins in mouse primary chondrocytes treated as indicated for 48 h ( n = 3). Scale bars, 25 μm. (I) qPCR analysis of indicated genes in mouse primary chondrocytes treated as indicated for 48 h ( n = 3). (J and K) Western blot analysis (J) and quantitation (K) of mouse primary chondrocytes treated as indicated for 48 h ( n = 3). Scale bars, 50 μm. Data are expressed as mean ± SD. ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001; NS, nonsignificant. Statistical analysis was performed by unpaired t test (A and B) and one-way ANOVA (F, H, I, and K).

Article Snippet: Slides were incubated overnight at 4°C with primary antibodies: SESN2 (Proteintech, #66297-1-Ig, 1:200), FASN (Proteintech, #66591-1-Ig, 1:100), SCD1 (ABclonal, #A16429, 1:100), SREBP1 (Proteintech, #14088-1-AP, 1:200), PGC-1α (Abcam, #ab54481, 1:100).

Techniques: Western Blot, Quantitation Assay, Immunoprecipitation

Journal: iScience

Article Title: SESN2 maintains cartilage homeostasis by SREBP1-mediated lipid metabolism during osteoarthritis progression

doi: 10.1016/j.isci.2025.113097

Figure Lengend Snippet: SESN2 overexpression improves fatty acid metabolism disorders and ameliorates cartilage degeneration (A and B) IF (A) and corresponding quantitative analysis (B) of SESN2 in the articular cartilage of mice induced by sham or destabilization of medial meniscus (DMM) surgery with intraarticular injection of lv-nc and lv- Sesn2 ( n = 6). (C–E) S.O. staining (C) of mice treated as in (A) ( n = 6). Quantitation of Osteoarthritis Research Society International (OARSI) scores (D), chondrocyte numbers, and cartilage thickness (E) ( n = 6). (F and G) IHC (F) and corresponding quantitative analysis (G) of COL2A1 and MMP13 in the articular cartilage of mice treated as in (A) ( n = 6). (H and I) BODIPY493/503 staining, IF (H) of SREBP1, FASN, and SCD1, and corresponding quantitative analysis (I) in the articular cartilage of mice treated as in (A) ( n = 6). (J and K) IF of PGC-1α and TUNEL staining (J) and corresponding quantitative analysis (K) in the articular cartilage of mice treated as in (A) ( n = 6). Scale bars: (F, H, and J) 50 μm, (C) 50 and 100 μm. Data ( n = 6) are shown as mean ± SD. ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001. Statistical analysis was performed by one-way ANOVA (B, D, E, G, I, and K).

Article Snippet: Slides were incubated overnight at 4°C with primary antibodies: SESN2 (Proteintech, #66297-1-Ig, 1:200), FASN (Proteintech, #66591-1-Ig, 1:100), SCD1 (ABclonal, #A16429, 1:100), SREBP1 (Proteintech, #14088-1-AP, 1:200), PGC-1α (Abcam, #ab54481, 1:100).

Techniques: Over Expression, Injection, Staining, Quantitation Assay, TUNEL Assay

Journal: iScience

Article Title: SESN2 maintains cartilage homeostasis by SREBP1-mediated lipid metabolism during osteoarthritis progression

doi: 10.1016/j.isci.2025.113097

Figure Lengend Snippet: SESN2 overexpression alleviates OA pathogenesis and joint dysfunction (A) Micro-CT analysis and quantitation of knee joints from the different groups. The sagittal image of the medial articular cavity depicts alterations in subchondral bone plate (SBP) thickness. (B and D) H&E staining of the synovium (B) and synovitis grade (D) of mice treated as indicated ( n = 6). (C) Osteophyte count of knee joints from mice treated as indicated ( n = 6). (E) The weight (left) and the knee diameter (right) of mice treated as indicated. (F) The outcomes of gait analysis for various cohorts. Blue print: forepaw; red print: hind paw. (G) Quantification of step length, stride length, and the length of the front/rear paw prints is shown as the dotted lines ( n = 6). Scale bars, 1 mm. (H) The spontaneous activity of mice after DMM surgery decreases in the open field test. (I) Quantification of activity, distance, active time, and mean speed ( n = 6). (J) Pain sensitivity was assessed by quantifying the paw withdrawal mechanical threshold (PWMT) ( n = 6). All data represent mean ± S.D. ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001; NS, nonsignificant. Statistical analysis was performed by one-way ANOVA (C, D, E, G, and I).

Article Snippet: Slides were incubated overnight at 4°C with primary antibodies: SESN2 (Proteintech, #66297-1-Ig, 1:200), FASN (Proteintech, #66591-1-Ig, 1:100), SCD1 (ABclonal, #A16429, 1:100), SREBP1 (Proteintech, #14088-1-AP, 1:200), PGC-1α (Abcam, #ab54481, 1:100).

Techniques: Over Expression, Micro-CT, Quantitation Assay, Staining, Activity Assay

Journal: iScience

Article Title: SESN2 maintains cartilage homeostasis by SREBP1-mediated lipid metabolism during osteoarthritis progression

doi: 10.1016/j.isci.2025.113097

Figure Lengend Snippet: Scheme illustrating the suppression of SESN2 on SREBP1 activation, which inhibits lipogenic enzyme expression to protect against cartilage degeneration in osteoarthritis

Article Snippet: Slides were incubated overnight at 4°C with primary antibodies: SESN2 (Proteintech, #66297-1-Ig, 1:200), FASN (Proteintech, #66591-1-Ig, 1:100), SCD1 (ABclonal, #A16429, 1:100), SREBP1 (Proteintech, #14088-1-AP, 1:200), PGC-1α (Abcam, #ab54481, 1:100).

Techniques: Activation Assay, Expressing

Journal: Archives of biochemistry and biophysics

Article Title: Activation of Sestrin2 accelerates deep second-degree burn wound healing through PI3K/AKT pathway.

doi: 10.1016/j.abb.2023.109645

Figure Lengend Snippet: Fig. 1. SESN2 is upregulated during the deep second-degree burn wound healing process. (A) The healing process of deep second-degree burns in C57BL/6 mice. Relative (B) mRNA and (C) protein level of SESN2 in wound tissues collected at indicated healing time points. (D) Immunohistochemical analysis of SESN2 expression at murine wound edge tissues, scale bar: 100 μm. (E) SESN2 expression levels in uninjured and injured keratinocytes from GEO dataset (GSE30355). The data are presented as the mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001, n = 5.

Article Snippet: After incubation with SESN2 antibody (1:500) overnight at 4 ◦C, the sections were incubated with secondary antibody horseradish peroxidase (HRP)-labeled goat anti-rabbit IgG (1: 1000, Wuhan Boster Biological Technology, Wuhan, China) at room temperature for 30 min.

Techniques: Immunohistochemical staining, Expressing

Journal: Archives of biochemistry and biophysics

Article Title: Activation of Sestrin2 accelerates deep second-degree burn wound healing through PI3K/AKT pathway.

doi: 10.1016/j.abb.2023.109645

Figure Lengend Snippet: Fig. 2. Up-regulation of SESN2 accelerates the burn wound healing process (A) Representative images and the (B) quantification of the burn wound area of mice in control group, eupatilin treated group and si-SESN2 group at indicated time (0 d, 7 d, 14 d, and 21 d). (C) HE and (D) Masson staining of tissue sections from different groups (control, eupatilin and si-SESN2), scale bar for HE: 1 mm and Masson: 400 μm. The data are presented as the mean ± SD. *P < 0.05, **P < 0.01, n = 5.

Article Snippet: After incubation with SESN2 antibody (1:500) overnight at 4 ◦C, the sections were incubated with secondary antibody horseradish peroxidase (HRP)-labeled goat anti-rabbit IgG (1: 1000, Wuhan Boster Biological Technology, Wuhan, China) at room temperature for 30 min.

Techniques: Control, Staining

Journal: Archives of biochemistry and biophysics

Article Title: Activation of Sestrin2 accelerates deep second-degree burn wound healing through PI3K/AKT pathway.

doi: 10.1016/j.abb.2023.109645

Figure Lengend Snippet: Fig. 3. SESN2 promotes the re-epithelialization during burn wound healing process. (A) The mRNA level of cytokines including TNF-α, IL-1β and IL-6 in murine wound edge tissues during the wound healing process. (B-C) The immunofluorescence staining of SESN2 (B), Ki67 (C) at murine wound edges of mice in control group, eupatilin treated group and si-SESN2 group, scale bar: 200 μm. The data are presented as the mean ± SD. **P < 0.01.

Article Snippet: After incubation with SESN2 antibody (1:500) overnight at 4 ◦C, the sections were incubated with secondary antibody horseradish peroxidase (HRP)-labeled goat anti-rabbit IgG (1: 1000, Wuhan Boster Biological Technology, Wuhan, China) at room temperature for 30 min.

Techniques: Immunofluorescence, Staining, Control

Journal: Archives of biochemistry and biophysics

Article Title: Activation of Sestrin2 accelerates deep second-degree burn wound healing through PI3K/AKT pathway.

doi: 10.1016/j.abb.2023.109645

Figure Lengend Snippet: Fig. 4. SESN2 promotes keratinocyte proliferation and migration. SESN2 expression levels in HaCaT cell treated with or without heat were measured by (A) qRT-PCR and (B) western blotting. The (C) mRNA and (D) protein levels of SESN2 in HaCaT cells treated with indicated concentrations of eupatilin. HaCaT cells pre-treated with heat were treated with or without eupatilin. (E) Cell viability was measured by CCK-8 assay, and (F) EDU assay was employed to assess the cell proliferation. (G–H) Wound healing assay (G) images and (H) quantitative analysis of HaCaT cells treated with or without 50 μM eupatilin at indicated time. (I) Transwell assay images and quantitative analysis of the migration of HaCaT cells treated with or without 50 μM eupatilin. The data are presented as the mean ± SD. *P < 0.05, **P < 0.01.

Article Snippet: After incubation with SESN2 antibody (1:500) overnight at 4 ◦C, the sections were incubated with secondary antibody horseradish peroxidase (HRP)-labeled goat anti-rabbit IgG (1: 1000, Wuhan Boster Biological Technology, Wuhan, China) at room temperature for 30 min.

Techniques: Migration, Expressing, Quantitative RT-PCR, Western Blot, CCK-8 Assay, EdU Assay, Wound Healing Assay, Transwell Assay

Journal: Archives of biochemistry and biophysics

Article Title: Activation of Sestrin2 accelerates deep second-degree burn wound healing through PI3K/AKT pathway.

doi: 10.1016/j.abb.2023.109645

Figure Lengend Snippet: Fig. 5. Deficiency of SESN2 impairs keratinocyte proliferation and migration. The knockdown efficiency of si-SESN2 in HaCaT cell pretreated with heat was confirmed by (A) qRT-PCR and (B) western blot assay. HaCaT cells pre-treated with heat were transfected with si-SESN2 or si-NC. (C) Cell viability was measured by CCK-8 assay, and (D) EDU assay was employed to evaluate the cell proliferation. (E–F) Wound healing assay (E) images and (F) quantitative analysis of HaCaT cells transfected with si-SESN2 or si-NC at indicated time. (G) Transwell assay images and quantitative analysis of the migration of HaCaT cells transfected with si-SESN2 or si-NC. The data are presented as the mean ± SD. *P < 0.05, **P < 0.01.

Article Snippet: After incubation with SESN2 antibody (1:500) overnight at 4 ◦C, the sections were incubated with secondary antibody horseradish peroxidase (HRP)-labeled goat anti-rabbit IgG (1: 1000, Wuhan Boster Biological Technology, Wuhan, China) at room temperature for 30 min.

Techniques: Migration, Knockdown, Quantitative RT-PCR, Western Blot, Transfection, CCK-8 Assay, EdU Assay, Wound Healing Assay, Transwell Assay

Journal: Archives of biochemistry and biophysics

Article Title: Activation of Sestrin2 accelerates deep second-degree burn wound healing through PI3K/AKT pathway.

doi: 10.1016/j.abb.2023.109645

Figure Lengend Snippet: Fig. 6. PI3K/AKT pathway mediates SESN2-deriven keratinocyte proliferation and migration. (A) Western blotting and quantitative analysis of SESN2, PI3K, p-PI3K, AKT, and p-AKT expression in HaCaT cells treated with eupatilin or LY294002. (B) Western blotting and quantitative analysis of SESN2, PI3K, p-PI3K, AKT, and p- AKT expression in HaCaT cells treated with 740Y-P with or without the knockdown of SESN2. (C) CCK-8 assay of HaCaT cells treated with eupatilin or LY294002 at indicated time. (D) Transwell assay images and quantitative analysis of HaCaT cells treated with eupatilin or LY294002. (E) CCK-8 assay of HaCaT cells treated with 740Y-P with or without the knockdown of SESN2 at indicated time. (F) Transwell assay images and quantitative analysis of HaCaT cells treated with 740Y-P with or without the knockdown of SESN2. The data are presented as the mean ± SD. *P < 0.05, **P < 0.01, NS, no significance.

Article Snippet: After incubation with SESN2 antibody (1:500) overnight at 4 ◦C, the sections were incubated with secondary antibody horseradish peroxidase (HRP)-labeled goat anti-rabbit IgG (1: 1000, Wuhan Boster Biological Technology, Wuhan, China) at room temperature for 30 min.

Techniques: Migration, Western Blot, Expressing, Knockdown, CCK-8 Assay, Transwell Assay

Journal: Journal of cell science

Article Title: Osh6 requires Ist2 for localization to ER-PM contacts and efficient phosphatidylserine transport in budding yeast.

doi: 10.1242/jcs.243733

Figure Lengend Snippet: Fig. 1. Ist2 is required for Osh6 localization to the cortical ER. (A) Localization of

Article Snippet: Maeda et al., 2013 pAC100 pADH1 OSH6-mCherry (U) ADH1prOSH6-mCherry, CEN, URA3 (pRS315-based); marker swap This study pVA111 pCYC1 Osh6-mCherry CYC1prOSH6-mCherry, CEN, URA3 (pRS316-based); OSH6 subcloned This study pJMD_22 pADH1prOsh6 L141A D142A Quikchange mutagenesis of pAC100 This study pJMD_23 pCYC1prOsh6 L141A D142A Quikchange mutagenesis of pVA111 This study pJMD_24 pADH1prOsh6 L141K D142A Quikchange mutagenesis of pAC100 This study pJMD_25 pCYC1 Osh6 L141K D142A Quikchange mutagenesis of pVA111 This study pC2Lact-GFP C2lact-GFP GPDprC2Lact-GFP, CEN, URA3 (pRS416-based) Yeung et al., 2008 pAC107 C2lact-GFP (LEU) GPDprC2Lact-GFP, CEN, LEU2 (pRS416-based) Lipp et al., 2019 pJMD_26 crRNA Ist2 736-743 crRNA targeting IST2 codons 736-743, URA, 2 μ ori, based on pUD628 (Addgene Plasmid #103018) This study pUDC175 fnCpf1 TEF1prFnCPF1, CEN, NatMX (p414TEF1 Backbone) Addgene Plasmid #103019 Swiat et al., 2017 pVA117 pRS415-ADHprOsh6D35-VN ADHprOsh6D35 (Lipp et al., 2019) subcloned into pRS415-VN using SacI/BamHI digestion This study pVA119 pRS415-ADHprOsh6D69-VN ADHprOsh6D69 (Lipp et al., 2019) subcloned into pRS415-VN using SacI/BamHI digestion This study J.

Techniques:

Journal: Journal of cell science

Article Title: Osh6 requires Ist2 for localization to ER-PM contacts and efficient phosphatidylserine transport in budding yeast.

doi: 10.1242/jcs.243733

Figure Lengend Snippet: Fig. 2. Osh6 interacts with the disordered cytosolic tail of Ist2. (A) Schematic representation

Article Snippet: Maeda et al., 2013 pAC100 pADH1 OSH6-mCherry (U) ADH1prOSH6-mCherry, CEN, URA3 (pRS315-based); marker swap This study pVA111 pCYC1 Osh6-mCherry CYC1prOSH6-mCherry, CEN, URA3 (pRS316-based); OSH6 subcloned This study pJMD_22 pADH1prOsh6 L141A D142A Quikchange mutagenesis of pAC100 This study pJMD_23 pCYC1prOsh6 L141A D142A Quikchange mutagenesis of pVA111 This study pJMD_24 pADH1prOsh6 L141K D142A Quikchange mutagenesis of pAC100 This study pJMD_25 pCYC1 Osh6 L141K D142A Quikchange mutagenesis of pVA111 This study pC2Lact-GFP C2lact-GFP GPDprC2Lact-GFP, CEN, URA3 (pRS416-based) Yeung et al., 2008 pAC107 C2lact-GFP (LEU) GPDprC2Lact-GFP, CEN, LEU2 (pRS416-based) Lipp et al., 2019 pJMD_26 crRNA Ist2 736-743 crRNA targeting IST2 codons 736-743, URA, 2 μ ori, based on pUD628 (Addgene Plasmid #103018) This study pUDC175 fnCpf1 TEF1prFnCPF1, CEN, NatMX (p414TEF1 Backbone) Addgene Plasmid #103019 Swiat et al., 2017 pVA117 pRS415-ADHprOsh6D35-VN ADHprOsh6D35 (Lipp et al., 2019) subcloned into pRS415-VN using SacI/BamHI digestion This study pVA119 pRS415-ADHprOsh6D69-VN ADHprOsh6D69 (Lipp et al., 2019) subcloned into pRS415-VN using SacI/BamHI digestion This study J.

Techniques:

Journal: Journal of cell science

Article Title: Osh6 requires Ist2 for localization to ER-PM contacts and efficient phosphatidylserine transport in budding yeast.

doi: 10.1242/jcs.243733

Figure Lengend Snippet: Fig. 4. Osh6 binding site in Ist2 tail is sufficient for localizing Osh6 to the ER-PM contact sites.

Article Snippet: Maeda et al., 2013 pAC100 pADH1 OSH6-mCherry (U) ADH1prOSH6-mCherry, CEN, URA3 (pRS315-based); marker swap This study pVA111 pCYC1 Osh6-mCherry CYC1prOSH6-mCherry, CEN, URA3 (pRS316-based); OSH6 subcloned This study pJMD_22 pADH1prOsh6 L141A D142A Quikchange mutagenesis of pAC100 This study pJMD_23 pCYC1prOsh6 L141A D142A Quikchange mutagenesis of pVA111 This study pJMD_24 pADH1prOsh6 L141K D142A Quikchange mutagenesis of pAC100 This study pJMD_25 pCYC1 Osh6 L141K D142A Quikchange mutagenesis of pVA111 This study pC2Lact-GFP C2lact-GFP GPDprC2Lact-GFP, CEN, URA3 (pRS416-based) Yeung et al., 2008 pAC107 C2lact-GFP (LEU) GPDprC2Lact-GFP, CEN, LEU2 (pRS416-based) Lipp et al., 2019 pJMD_26 crRNA Ist2 736-743 crRNA targeting IST2 codons 736-743, URA, 2 μ ori, based on pUD628 (Addgene Plasmid #103018) This study pUDC175 fnCpf1 TEF1prFnCPF1, CEN, NatMX (p414TEF1 Backbone) Addgene Plasmid #103019 Swiat et al., 2017 pVA117 pRS415-ADHprOsh6D35-VN ADHprOsh6D35 (Lipp et al., 2019) subcloned into pRS415-VN using SacI/BamHI digestion This study pVA119 pRS415-ADHprOsh6D69-VN ADHprOsh6D69 (Lipp et al., 2019) subcloned into pRS415-VN using SacI/BamHI digestion This study J.

Techniques: Binding Assay

Journal: Journal of cell science

Article Title: Osh6 requires Ist2 for localization to ER-PM contacts and efficient phosphatidylserine transport in budding yeast.

doi: 10.1242/jcs.243733

Figure Lengend Snippet: Fig. 5. Mutations in Ist2 tail decrease steady-state levels of PS. Lipidomic analysis of WT,

Article Snippet: Maeda et al., 2013 pAC100 pADH1 OSH6-mCherry (U) ADH1prOSH6-mCherry, CEN, URA3 (pRS315-based); marker swap This study pVA111 pCYC1 Osh6-mCherry CYC1prOSH6-mCherry, CEN, URA3 (pRS316-based); OSH6 subcloned This study pJMD_22 pADH1prOsh6 L141A D142A Quikchange mutagenesis of pAC100 This study pJMD_23 pCYC1prOsh6 L141A D142A Quikchange mutagenesis of pVA111 This study pJMD_24 pADH1prOsh6 L141K D142A Quikchange mutagenesis of pAC100 This study pJMD_25 pCYC1 Osh6 L141K D142A Quikchange mutagenesis of pVA111 This study pC2Lact-GFP C2lact-GFP GPDprC2Lact-GFP, CEN, URA3 (pRS416-based) Yeung et al., 2008 pAC107 C2lact-GFP (LEU) GPDprC2Lact-GFP, CEN, LEU2 (pRS416-based) Lipp et al., 2019 pJMD_26 crRNA Ist2 736-743 crRNA targeting IST2 codons 736-743, URA, 2 μ ori, based on pUD628 (Addgene Plasmid #103018) This study pUDC175 fnCpf1 TEF1prFnCPF1, CEN, NatMX (p414TEF1 Backbone) Addgene Plasmid #103019 Swiat et al., 2017 pVA117 pRS415-ADHprOsh6D35-VN ADHprOsh6D35 (Lipp et al., 2019) subcloned into pRS415-VN using SacI/BamHI digestion This study pVA119 pRS415-ADHprOsh6D69-VN ADHprOsh6D69 (Lipp et al., 2019) subcloned into pRS415-VN using SacI/BamHI digestion This study J.

Techniques:

Journal: Journal of cell science

Article Title: Osh6 requires Ist2 for localization to ER-PM contacts and efficient phosphatidylserine transport in budding yeast.

doi: 10.1242/jcs.243733

Figure Lengend Snippet: Fig. 6. Binding of Osh6 to the disordered tail of Ist2 is required for PS transport to the PM.

Article Snippet: Maeda et al., 2013 pAC100 pADH1 OSH6-mCherry (U) ADH1prOSH6-mCherry, CEN, URA3 (pRS315-based); marker swap This study pVA111 pCYC1 Osh6-mCherry CYC1prOSH6-mCherry, CEN, URA3 (pRS316-based); OSH6 subcloned This study pJMD_22 pADH1prOsh6 L141A D142A Quikchange mutagenesis of pAC100 This study pJMD_23 pCYC1prOsh6 L141A D142A Quikchange mutagenesis of pVA111 This study pJMD_24 pADH1prOsh6 L141K D142A Quikchange mutagenesis of pAC100 This study pJMD_25 pCYC1 Osh6 L141K D142A Quikchange mutagenesis of pVA111 This study pC2Lact-GFP C2lact-GFP GPDprC2Lact-GFP, CEN, URA3 (pRS416-based) Yeung et al., 2008 pAC107 C2lact-GFP (LEU) GPDprC2Lact-GFP, CEN, LEU2 (pRS416-based) Lipp et al., 2019 pJMD_26 crRNA Ist2 736-743 crRNA targeting IST2 codons 736-743, URA, 2 μ ori, based on pUD628 (Addgene Plasmid #103018) This study pUDC175 fnCpf1 TEF1prFnCPF1, CEN, NatMX (p414TEF1 Backbone) Addgene Plasmid #103019 Swiat et al., 2017 pVA117 pRS415-ADHprOsh6D35-VN ADHprOsh6D35 (Lipp et al., 2019) subcloned into pRS415-VN using SacI/BamHI digestion This study pVA119 pRS415-ADHprOsh6D69-VN ADHprOsh6D69 (Lipp et al., 2019) subcloned into pRS415-VN using SacI/BamHI digestion This study J.

Techniques: Binding Assay

Journal: Journal of cell science

Article Title: Osh6 requires Ist2 for localization to ER-PM contacts and efficient phosphatidylserine transport in budding yeast.

doi: 10.1242/jcs.243733

Figure Lengend Snippet: Fig. 7. Identification of a conserved patch on Osh6 that interact with Ist2-tail. (A) Structure

Article Snippet: Maeda et al., 2013 pAC100 pADH1 OSH6-mCherry (U) ADH1prOSH6-mCherry, CEN, URA3 (pRS315-based); marker swap This study pVA111 pCYC1 Osh6-mCherry CYC1prOSH6-mCherry, CEN, URA3 (pRS316-based); OSH6 subcloned This study pJMD_22 pADH1prOsh6 L141A D142A Quikchange mutagenesis of pAC100 This study pJMD_23 pCYC1prOsh6 L141A D142A Quikchange mutagenesis of pVA111 This study pJMD_24 pADH1prOsh6 L141K D142A Quikchange mutagenesis of pAC100 This study pJMD_25 pCYC1 Osh6 L141K D142A Quikchange mutagenesis of pVA111 This study pC2Lact-GFP C2lact-GFP GPDprC2Lact-GFP, CEN, URA3 (pRS416-based) Yeung et al., 2008 pAC107 C2lact-GFP (LEU) GPDprC2Lact-GFP, CEN, LEU2 (pRS416-based) Lipp et al., 2019 pJMD_26 crRNA Ist2 736-743 crRNA targeting IST2 codons 736-743, URA, 2 μ ori, based on pUD628 (Addgene Plasmid #103018) This study pUDC175 fnCpf1 TEF1prFnCPF1, CEN, NatMX (p414TEF1 Backbone) Addgene Plasmid #103019 Swiat et al., 2017 pVA117 pRS415-ADHprOsh6D35-VN ADHprOsh6D35 (Lipp et al., 2019) subcloned into pRS415-VN using SacI/BamHI digestion This study pVA119 pRS415-ADHprOsh6D69-VN ADHprOsh6D69 (Lipp et al., 2019) subcloned into pRS415-VN using SacI/BamHI digestion This study J.

Techniques:

Journal: Journal of cell science

Article Title: Osh6 requires Ist2 for localization to ER-PM contacts and efficient phosphatidylserine transport in budding yeast.

doi: 10.1242/jcs.243733

Figure Lengend Snippet: Fig. 8. Osh6 D141/L142 mutants decrease interaction with Ist2 and PS transport. (A)

Article Snippet: Maeda et al., 2013 pAC100 pADH1 OSH6-mCherry (U) ADH1prOSH6-mCherry, CEN, URA3 (pRS315-based); marker swap This study pVA111 pCYC1 Osh6-mCherry CYC1prOSH6-mCherry, CEN, URA3 (pRS316-based); OSH6 subcloned This study pJMD_22 pADH1prOsh6 L141A D142A Quikchange mutagenesis of pAC100 This study pJMD_23 pCYC1prOsh6 L141A D142A Quikchange mutagenesis of pVA111 This study pJMD_24 pADH1prOsh6 L141K D142A Quikchange mutagenesis of pAC100 This study pJMD_25 pCYC1 Osh6 L141K D142A Quikchange mutagenesis of pVA111 This study pC2Lact-GFP C2lact-GFP GPDprC2Lact-GFP, CEN, URA3 (pRS416-based) Yeung et al., 2008 pAC107 C2lact-GFP (LEU) GPDprC2Lact-GFP, CEN, LEU2 (pRS416-based) Lipp et al., 2019 pJMD_26 crRNA Ist2 736-743 crRNA targeting IST2 codons 736-743, URA, 2 μ ori, based on pUD628 (Addgene Plasmid #103018) This study pUDC175 fnCpf1 TEF1prFnCPF1, CEN, NatMX (p414TEF1 Backbone) Addgene Plasmid #103019 Swiat et al., 2017 pVA117 pRS415-ADHprOsh6D35-VN ADHprOsh6D35 (Lipp et al., 2019) subcloned into pRS415-VN using SacI/BamHI digestion This study pVA119 pRS415-ADHprOsh6D69-VN ADHprOsh6D69 (Lipp et al., 2019) subcloned into pRS415-VN using SacI/BamHI digestion This study J.

Techniques: