syntaxin 4 (sx4 Search Results


syntaxin 4 (sx4; #110 042, rrid:ab_887853)  (Synaptic Systems)
90
Synaptic Systems syntaxin 4 (sx4; #110 042, rrid:ab_887853)
EHD2 KD in 3T3-L1 adipocytes is associated with impaired insulin signaling. (A) Representative Western blots of total phospho-tyrosine in 3T3-L1 adipocyte lysates collected 96 h after gene silencing with control siRNA or EHD2 siRNA, either untreated or following 20-min stimulation with 100 nM insulin ( n = 2) and of protein levels of EHD2, SNARE proteins Syntaxin4 <t>(Sx4),</t> SNAP23, VAMP2, and Syntaxin16 (Sx16), as well as regulatory protein Munc18c, and GLUT4 ( n = 3). GAPDH was used as a loading control and the calculations displayed in C were performed exclusively on basal samples. (B) Glucose uptake (2-deoxy-D-glucose) of 3T3-L1 adipocytes, 96 h after gene silencing with control siRNA (Control) or EHD2 siRNA (EHD2 KD), with (INS) or without (Basal) 20-min stimulation with 100 nM insulin. Data were corrected for nonspecific cellular isotope uptake by performing parallel assays in the presence of 10 μM cytochalasin B and normalized to those obtained in the insulin-stimulated control adipocytes for each data set. Mean ± SD of n = 4 independent experiments are shown. Statistical analysis was done using two-way ANOVA Tukey’s Honest Significant Difference (TukeyHSD), * p < 0.05. (C) Corresponding quantification of protein expression in EHD2 siRNA KD adipocytes in the absence of an acute insulin challenge (lanes labeled “-” in A) is normalized to GAPDH and expressed as a percentage of protein expression in control siRNA adipocytes. Mean and SD of n = 3 independent experiments are shown. Statistical analysis was conducted using unpaired two-sample t test, * p < 0.05, ** p < 0.01.
Syntaxin 4 (Sx4; #110 042, Rrid:Ab 887853), supplied by Synaptic Systems, 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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EHD2 KD in 3T3-L1 adipocytes is associated with impaired insulin signaling. (A) Representative Western blots of total phospho-tyrosine in 3T3-L1 adipocyte lysates collected 96 h after gene silencing with control siRNA or EHD2 siRNA, either untreated or following 20-min stimulation with 100 nM insulin ( n = 2) and of protein levels of EHD2, SNARE proteins Syntaxin4 (Sx4), SNAP23, VAMP2, and Syntaxin16 (Sx16), as well as regulatory protein Munc18c, and GLUT4 ( n = 3). GAPDH was used as a loading control and the calculations displayed in C were performed exclusively on basal samples. (B) Glucose uptake (2-deoxy-D-glucose) of 3T3-L1 adipocytes, 96 h after gene silencing with control siRNA (Control) or EHD2 siRNA (EHD2 KD), with (INS) or without (Basal) 20-min stimulation with 100 nM insulin. Data were corrected for nonspecific cellular isotope uptake by performing parallel assays in the presence of 10 μM cytochalasin B and normalized to those obtained in the insulin-stimulated control adipocytes for each data set. Mean ± SD of n = 4 independent experiments are shown. Statistical analysis was done using two-way ANOVA Tukey’s Honest Significant Difference (TukeyHSD), * p < 0.05. (C) Corresponding quantification of protein expression in EHD2 siRNA KD adipocytes in the absence of an acute insulin challenge (lanes labeled “-” in A) is normalized to GAPDH and expressed as a percentage of protein expression in control siRNA adipocytes. Mean and SD of n = 3 independent experiments are shown. Statistical analysis was conducted using unpaired two-sample t test, * p < 0.05, ** p < 0.01.

Journal: Molecular Biology of the Cell

Article Title: EHD2 regulates plasma membrane integrity and downstream insulin receptor signaling events

doi: 10.1091/mbc.E23-03-0078

Figure Lengend Snippet: EHD2 KD in 3T3-L1 adipocytes is associated with impaired insulin signaling. (A) Representative Western blots of total phospho-tyrosine in 3T3-L1 adipocyte lysates collected 96 h after gene silencing with control siRNA or EHD2 siRNA, either untreated or following 20-min stimulation with 100 nM insulin ( n = 2) and of protein levels of EHD2, SNARE proteins Syntaxin4 (Sx4), SNAP23, VAMP2, and Syntaxin16 (Sx16), as well as regulatory protein Munc18c, and GLUT4 ( n = 3). GAPDH was used as a loading control and the calculations displayed in C were performed exclusively on basal samples. (B) Glucose uptake (2-deoxy-D-glucose) of 3T3-L1 adipocytes, 96 h after gene silencing with control siRNA (Control) or EHD2 siRNA (EHD2 KD), with (INS) or without (Basal) 20-min stimulation with 100 nM insulin. Data were corrected for nonspecific cellular isotope uptake by performing parallel assays in the presence of 10 μM cytochalasin B and normalized to those obtained in the insulin-stimulated control adipocytes for each data set. Mean ± SD of n = 4 independent experiments are shown. Statistical analysis was done using two-way ANOVA Tukey’s Honest Significant Difference (TukeyHSD), * p < 0.05. (C) Corresponding quantification of protein expression in EHD2 siRNA KD adipocytes in the absence of an acute insulin challenge (lanes labeled “-” in A) is normalized to GAPDH and expressed as a percentage of protein expression in control siRNA adipocytes. Mean and SD of n = 3 independent experiments are shown. Statistical analysis was conducted using unpaired two-sample t test, * p < 0.05, ** p < 0.01.

Article Snippet: Mammalian uncoordinated-18c (Munc18c; #116 202, RRID:AB_2619785), Syntaxin 4 (Sx4; #110 042, RRID:AB_887853), vesicle-associated membrane protein 2 (VAMP2; #104 403, RRID:AB_2864782), synaptosome associated protein 23 (SNAP23; #111 213, RRID:AB_10805651), and Syntaxin 16 (Sx16; #110 162, RRID:AB_887799) were all from Synaptic Systems (Coventry, UK).

Techniques: Western Blot, Expressing, Labeling

Reduced cholesterol and altered PM lipid content in EHD2 KO adipocytes. (A) Whole-cell and (B) PM samples were subjected to immunoblotting for Munc18c, Sx4, and SNAP23. All data are presented as percentage of WT levels (WT = 100%, dashed line), n = 3 biological replicates (A) and n = 3–6 (B). Data are displayed as mean ± SD, and unpaired two-sample t test was used for statistical analysis. Significance was determined according to ** p ≤ 0.01. (C) Cholesterol levels in FC, PM, and serum (S) from WT and EHD2 KO mice. Differences assessed by two-way ANOVA, involving an interaction (x) between genotype (g) and sample (t) with a posthoc Student’s t test. (D) Score plots from OPLS-DA calculated on membrane lipidomic data acquired in positive (top) and negative (below) electrospray ionization mode. t1, first predictive component; to[1], first orthogonal component. WT in white, EHD2 in gray. (E) Lipids showing significant differences between WT and EHD2 KO PMs (variable importance of projection (VIP >1). PE, PE_ep, PC, and SM. Enrichment was accessed using χ 2 statistics; PC_ep, q = 0.0013, PE, q = 0.011, PE_ep, q = 0.00081, SM, q = 0.00022. Significance was determined according to * p ≤ 0.05, ** p ≤ 0.01 and *** p ≤ 0.001, n = 3 biological replicates. NKA = Na + /K + -ATPase. All displayed results were obtained from inguinal adipocytes 2 wk of HFD.

Journal: Molecular Biology of the Cell

Article Title: EHD2 regulates plasma membrane integrity and downstream insulin receptor signaling events

doi: 10.1091/mbc.E23-03-0078

Figure Lengend Snippet: Reduced cholesterol and altered PM lipid content in EHD2 KO adipocytes. (A) Whole-cell and (B) PM samples were subjected to immunoblotting for Munc18c, Sx4, and SNAP23. All data are presented as percentage of WT levels (WT = 100%, dashed line), n = 3 biological replicates (A) and n = 3–6 (B). Data are displayed as mean ± SD, and unpaired two-sample t test was used for statistical analysis. Significance was determined according to ** p ≤ 0.01. (C) Cholesterol levels in FC, PM, and serum (S) from WT and EHD2 KO mice. Differences assessed by two-way ANOVA, involving an interaction (x) between genotype (g) and sample (t) with a posthoc Student’s t test. (D) Score plots from OPLS-DA calculated on membrane lipidomic data acquired in positive (top) and negative (below) electrospray ionization mode. t1, first predictive component; to[1], first orthogonal component. WT in white, EHD2 in gray. (E) Lipids showing significant differences between WT and EHD2 KO PMs (variable importance of projection (VIP >1). PE, PE_ep, PC, and SM. Enrichment was accessed using χ 2 statistics; PC_ep, q = 0.0013, PE, q = 0.011, PE_ep, q = 0.00081, SM, q = 0.00022. Significance was determined according to * p ≤ 0.05, ** p ≤ 0.01 and *** p ≤ 0.001, n = 3 biological replicates. NKA = Na + /K + -ATPase. All displayed results were obtained from inguinal adipocytes 2 wk of HFD.

Article Snippet: Mammalian uncoordinated-18c (Munc18c; #116 202, RRID:AB_2619785), Syntaxin 4 (Sx4; #110 042, RRID:AB_887853), vesicle-associated membrane protein 2 (VAMP2; #104 403, RRID:AB_2864782), synaptosome associated protein 23 (SNAP23; #111 213, RRID:AB_10805651), and Syntaxin 16 (Sx16; #110 162, RRID:AB_887799) were all from Synaptic Systems (Coventry, UK).

Techniques: Western Blot, Membrane