fk506 binding protein 5  (Novus Biologicals)

Journal: Journal of Stroke

Article Title: Neuroprotective Effects of GV1001 in Animal Stroke Model and Neural Cells Subject to Oxygen-Glucose Deprivation/Reperfusion Injury

doi: 10.5853/jos.2021.00626

Figure Lengend Snippet: The effect of GV1001 on intracellular signaling proteins and cells. (A) Decreased levels of phosphorylated Akt (pAkt; Ser473), phospho-glycogen synthase kinase (pGSK-3β; Ser9), phosphorylated-extracellular signal-regulated kinase (pERK), B-cell lymphoma 2 (Bcl-2), and increased levels of Bcl-2 associated X (Bax) were noted in the peri-infarct regions of GV1001-treated rats compared to those of control rats. Administration of GV1001 (30 and 60 μM/kg) alleviated the changes. (B) Immunohistochemistry (IHC) staining showing increased pAkt (Ser473), pGSK-3β (Ser9), pERK, Bcl-2 levels and decreased Bax levels along with increased numbers of neuronal nuclei (NeuN)- or SRY-box transcription factor 2 (SOX2)-positive cells in GV1001-treated groups, scale bar: 50 μm. (C) IHC also revealed increased levels of nestin (a neuroectodermal stem cell marker), NeuN (a neuronal nuclear antigen), doublecortin (DCX; a neuronal differentiation marker), and SOX2 (a multipotent neural stem cell marker) in the peri-infarct region of GV1001-treated rats. Glial fibrillary acidic protein (GFAP; an astrocyte marker) expression was decreased in GV1001-treated rats, scale bar: 50 μm. (D) Levels of neurotoxic phenotype reactive astrocytes (uridine diphosphate glucuronosyltransferase 1A1 [UGT1A1] and FK506 binding protein 5 [FKBP5]) were increased and those of neuroprotective phenotype reactive astrocytes (sphingosine kinase type 1 [SPHK1]) were decreased in the peri-infarct area, but GV1001 significantly restored the expression of these markers. pIRS-1, phospho-insulin receptor substrate-1; DAPI, 4',6-diamidino-2-phenylindole. * P <0.05 (vs. sham group); † P <0.01 (vs. sham group); ‡ P <0.05 (vs. saline group); § P <0.01 (vs. saline group).

Article Snippet: The peri-infarct region in the control (sham and saline) group and the corresponding area in the GV1001-treated group, which were confirmed based on MRI, were used in Western blot analysis for phospho-insulin receptor substrate-1 (pIRS-1) (Ser636/639; 1:1,000, 2388, Cell Signaling Technology), phospho-phosphoinositide 3-kinase (pPI3K) p85 (Tyr458)/p55(Tyr199) (p85α PI3K; 1:1,000, 4228, Cell Signaling Technology), pAkt (Ser473), Akt (1:2,000, 9272, Cell Signaling Technology), pGSK-3β (1:1,000, 9336, Cell Signaling Technology), GSK-3β (1:2,000, sc-9166, Santa Cruz Biotechnology), pERK1/2 (Thr202/Tyr204), Bcl-2, Bax (1:1,000, 2772, Cell Signaling Technology), uridine diphosphate glucuronosyltransferase 1A1 (UGT1A1, 1:500, ab237810, Abcam), FK506 binding protein 5 (FKBP5, 0.4 μg/mL, NBP1-84676, Novus Biologicals, Centennial, CO, USA), sphingosine kinase type 1 (SPHK1, 1:1,000, ab71700, Abcam), and beta-tubulin (β-tubulin, 1:2,000, 2146, Cell Signaling Technology).

Techniques: Control, Immunohistochemistry, Marker, Expressing, Binding Assay, Saline

Journal: Oncology Letters

Article Title: Targeting of FK506 binding protein 5 by miR-203 affects the progression of breast cancer via regulating the fatty acid degradation pathway and potential drug-repurposing

doi: 10.3892/ol.2021.12607

Figure Lengend Snippet: Prediction of miR-203 targets, PPI network, and core genes in the PPI network. (A) A total of 17 target genes were identified between Down-DEGs and Targetscan 7.2. (B) PPI network of target genes. (C) Kaplan-Meier plot was used to estimate the overall survival of patients with breast cancer based on the expression of FKBP5. (D) The expression levels of FKBP5 in SUM159 cells transfected with miR-203 mimics or miR-203 NC were determined by western blot analysis. Data are expressed as the mean ± SD (n=3). *P<0.05 compared with the Cntrl group (SUM159 cells transfected with miR-203 NC; unpaired t-test). PPI, protein-protein interaction; Down-DEGs, downregulated differentially expressed genes; FKBP5, FK506 binding protein 5; Cntrl, control; HR, hazard ratio; miR, microRNA.

Article Snippet: Following blocking with 51% (w/v) skimmed milk in wash buffer (TBS and 0.051% Tween-20) for 1 h at room temperature, the membranes were incubated 4°C with the following primary antibodies: anti-GAPDH (1:10,000; cat. no. G9545; Sigma-Aldrich; Merck KGaA) and anti-FK506 binding protein 5 (FKBP5; 1:1,000; cat. no. sc-271547; Santa Cruz Biotechnology, Inc.).

Techniques: Expressing, Transfection, Western Blot, Binding Assay

Journal: Oncology Letters

Article Title: Targeting of FK506 binding protein 5 by miR-203 affects the progression of breast cancer via regulating the fatty acid degradation pathway and potential drug-repurposing

doi: 10.3892/ol.2021.12607

Figure Lengend Snippet: Migration capability of SUM159 breast cancer cells. To evaluate the migration ability of Cntrl (SUM159 cells transfected with miR-203 NC and empty control vector pCMV6-XL5), miR-203 (SUM159 cells transfected with miR-203 mimic), and FKBP5 (SUM159 cells transfected with pCMV6-XL5-FKBP5 vector), wound healing assays were carried out under a light phase-contrast microscope (magnification, ×40) (A) Representative images showing the wound healing at 0 and 15 h. Images were captured using an inverted microscope and the wound area was calculated at the indicated time points using ImageJ v.1.8.0 (B) Graph represents the normalized (to t=0 h) percentage of wound healing at 0 and 15 h following scratching. Data are expressed as the mean ± SD from 6 wound-healing assays for each group. Statistical analysis was performed using Kruskal Wallis non-parametric ANOVA with Dunn's post hoc test comparing every treatment and with its corresponding control (*P<0.05). (C) The liner map of vector pCMV6-XL5. Cntrl, control; FKBP5, FK506 binding protein 5; miR, microRNA.

Article Snippet: Following blocking with 51% (w/v) skimmed milk in wash buffer (TBS and 0.051% Tween-20) for 1 h at room temperature, the membranes were incubated 4°C with the following primary antibodies: anti-GAPDH (1:10,000; cat. no. G9545; Sigma-Aldrich; Merck KGaA) and anti-FK506 binding protein 5 (FKBP5; 1:1,000; cat. no. sc-271547; Santa Cruz Biotechnology, Inc.).

Techniques: Migration, Transfection, Plasmid Preparation, Microscopy, Inverted Microscopy, Binding Assay

Journal: Oncology Letters

Article Title: Targeting of FK506 binding protein 5 by miR-203 affects the progression of breast cancer via regulating the fatty acid degradation pathway and potential drug-repurposing

doi: 10.3892/ol.2021.12607

Figure Lengend Snippet: Potential drug-repurposing for FKBP5 by virtual screening. The 3D structure of ZINC000003944422 binding in FKBP5 is shown. Yellow and green arrows indicate hydrogen bonds and Pi interactions, respectively. Salt bridges are indicated by purple lines. FKBP5, FK506 binding protein 5; TYR, tyrosine; LYS, lysine; Pi interactions, salt bridge.

Article Snippet: Following blocking with 51% (w/v) skimmed milk in wash buffer (TBS and 0.051% Tween-20) for 1 h at room temperature, the membranes were incubated 4°C with the following primary antibodies: anti-GAPDH (1:10,000; cat. no. G9545; Sigma-Aldrich; Merck KGaA) and anti-FK506 binding protein 5 (FKBP5; 1:1,000; cat. no. sc-271547; Santa Cruz Biotechnology, Inc.).

Techniques: Binding Assay

Journal: Steroids

Article Title: The Requirement for p42/p44 MAPK Activity in Progesterone Receptor-Mediated Gene Regulation is Target Gene-Specific

doi: 10.1016/j.steroids.2012.12.014

Figure Lengend Snippet: A) T47D cells were cultured in serum-free medium for 24 hours before pretreatment with DMSO or U0126 (10 μM) for 1 hour, followed by treatment with ethanol or R5020 (10 nM) for 6 hours. PR and tubulin protein expression levels were determined by Western blot analysis. A representative experiment is shown. U0126 does not affect PR expression levels in the absence or presence of R5020. B) In parallel to the experiments in panel A, relative SGK and CCND1 mRNA expression was determined using real-time qPCR and normalized to 18S rRNA expression. SGK gene induction was insensitive to U0126, while CCND1 basal and hormone-mediated gene expression was decreased with U0126. * p<0.05 compared to respective treatment in DMSO group. C) T47D cells were cultured and pretreated as in panel A, followed by treatment with ethanol or R5020 (10 nM) for 24 hours. Relative SGK, NDRG1 and FKBP5 mRNA expression was determined using real-time qPCR and normalized to 18S rRNA expression. SGK gene induction was abrogated by U0126, while NDRG1 gene induction was robustly decreased by U0126. In contrast, FKBP5 gene induction was relatively insensitive to U0126. * p<0.05 compared to DMSO R5020. ****p<0.0001 compared to DMSO R5020.

Article Snippet: Primers for 18S (previously described in [ 30 ]), SGK and cyclin D1 (CCND1) (previously described in [ 20 ]), FK506 binding protein 5 (FKBP5) (sense 5′-GGATATACGCCAACATGTTCAA-3′, antisense 5′-CCATTGCTTTATTGGCCTCT-3′), N-myc downstream regulated 1 (NDRG1) (sense 5′-CTACCATGACATCGGCATGAA-3′, antisense 5′-CGTGGCAGACGGCAAAGT-3′), and interleukin 1 receptor, type 1 (IL1R1) (sense 5′-AATTGATGAAGATGACCCAGTGC-3′, antisense 5′-GCAGGATTTTCCACACTGTAATAGTCT-3′) were purchased from Sigma-Genosys (The Woodlands, TX).

Techniques: Cell Culture, Expressing, Western Blot