Journal: International Journal of Molecular Sciences

Article Title: Reduced VDAC1, Maintained Mitochondrial Dynamics and Enhanced Mitochondrial Biogenesis in a Transgenic Tau Mouse Model of Alzheimer’s Disease

doi: 10.3390/ijms23158561

Figure Lengend Snippet: Western blot, immunofluorescence, and quantification analysis of proteins regulating mitochondrial dynamics in 6-month-old WT, VDAC1 +/− , TAU, and VDAC1 +/− /TAU mice. ( A ) Representative immunoblots. ( B ) Quantitative densitometry study of mitochondrial dynamics found that the fission of DRP1 (* p < 0.05) and FIS1 (** p < 0.01) was dramatically decreased, whereas the fusion of MFN1, MFN2, and OPA1 (* p < 0.05) was significantly enhanced in VDAC1 +/− /TAU animals compared to TAU mice. Forty micrograms (g) of total protein were put into each lane. The loading control was carried out with the help of the housekeeping protein beta-actin. Data are from three independent experiments showed similar results (N = 3). Three animals were randomly selected from each group/genotype for immunoblotting and immunofluorescence analyses from a total of ten animals studied for behavioral phenotype . ( C ) Representative immunofluorescence images of 10-micron coronal sections (10×). ( D ) Fluorescence intensity analysis of mitochondrial dynamics-DRP1 (**** p < 0.0001), FIS1 (**** p < 0.0001) (fission) was significantly decreased, MFN1 (*** p < 0.001), MFN2 (**** p < 0.0001) and OPA1 (**** p < 0.0001) (fusion) were significantly increased in VDAC1 +/− /TAU mice compared to TAU mice. The data are from three separate experiments, all of which yielded comparable findings (N = 3), and each mouse was exposed to 10–15 fields. Scale bar: 200 μm. The results were presented as the mean accompanied by the standard error of the mean; ns denotes that the difference did not reach statistical significance; * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001; one-way ANOVA followed by Turkey’s test for multiple comparisons.

Article Snippet: After that, the sections were incubated with the appropriate primary antibodies [DRP1 (NB110-55288, Rabbit Polyclonal 1:100; Novus Biologicals), FIS1 (NB100-56646, Rabbit Polyclonal 1:100; Novus Biologicals), fusion proteins: MFN1: (13798-1-AP, Rabbit Polyclonal 1:100; Protein Tech Group), MFN2 (9482, Rabbit Polyclonal 1:100; Cell Signaling Technology), OPA1 (NBP2-59770, Rabbit Polyclonal 1:100; Novus Biologicals), PGC1A (NBP1-04676, Rabbit Polyclonal 1:100; Novus Biologicals), NRF1 (ab34682, Rabbit Polyclonal 1:100; Abcam), NRF2 (NBP1-32822, Rabbit Polyclonal 1:100; Novus Biologicals), TFAM (NBP2-19437, Rabbit Polyclonal 1:100; Novus Biologicals)] for a whole night at 4 degrees Celsius.

Techniques: Western Blot, Immunofluorescence, Control, Fluorescence

Journal: Bone research

Article Title: FAR591 promotes the pathogenesis and progression of SONFH by regulating Fos expression to mediate the apoptosis of bone microvascular endothelial cells.

doi: 10.1038/s41413-023-00259-8

Figure Lengend Snippet: Fig. 8 FAR591 recruited TAF15 and Pol II in the promoter region of the Fos gene to promote the expression of Fos by transcriptional activation. ChIRP-Seq detection of cis-acting elements interacting with FAR591 (n = 3): a Motif analysis results. b GO enrichment analysis. c KEGG enrichment analysis. d Visualization of FAR591 enrichment on chromosome 6. e Triplet diagram of FAR591 binding to the Fos gene promoter. f ChIRP-PCR was used to verify the binding site of FAR591 on the Fos gene promoter (n = 3). ChIRP-MS detection of trans-acting factors interacting with FAR591 (n = 3): g MS identified four specific peptides in TAF15. h ChIRP-western blot analysis verified the interaction of FAR591 with TAF15 (n = 4). ChIP-PCR was used to detect the binding sites of TAF15 (i) and Rpb1 (j) on the Fos gene promoter (n = 3). After overexpression or knockout of FAR591 in BMECs: The enrichment levels of FAR591 (k), TAF15 (l), and Rpb1 (m) on the Fos gene promoter were detected by ChIRP/ChIP-PCR (n = 3). n, o The expression of Fos protein was detected by western blotting (n = 4). In (f, i–m, o), data are presented as the means ± SDs; statistical significance was calculated by one-way ANOVA with Tukey’s post hoc tests; *P < 0.05, **P < 0.01, ***P < 0.001. Abbreviations: chromatin isolation by RNA purification (ChIRP), high-throughput sequencing (Seq), triplex target site (TTS), triplex- forming oligonucleotide (TFO), mass spectrometry (MS), control (CON), FAR591 (FAR), immunoprecipitation (IP), supernatant (SN), RNA polymerase II subunit 1 (Rpb1)

Article Snippet: Then, TAF15 (D8V6Q) rabbit mAb (1:50; 28409; Cell Signaling Technology), Rpb1 NTD (D8L4Y) rabbit mAb (1:50; 14958; Cell Signaling Technology), and rabbit (DA1E) mAb IgG (1:50; 3900; Cell Signaling Technology) were added and incubated overnight at 4 °C.

Techniques: Expressing, Activation Assay, Binding Assay, Western Blot, Over Expression, Knock-Out, Isolation, Next-Generation Sequencing, Mass Spectrometry, Control, Immunoprecipitation

Journal: Bone research

Article Title: FAR591 promotes the pathogenesis and progression of SONFH by regulating Fos expression to mediate the apoptosis of bone microvascular endothelial cells.

doi: 10.1038/s41413-023-00259-8

Figure Lengend Snippet: Fig. 9 Schematic diagram of the mechanism of FAR591-mediated GC-induced femoral head necrosis. GCs interact with GR, and activated GR translocates into the nucleus to bind to the FAR591 gene promoter and activate FAR591 gene expression. FAR591 binds to the Fos gene promoter to form a stable RNA:DNA triplet structure. FAR591 recruits TAF15 and Pol II in the promoter region of the Fos gene, which promotes preinitiation complex formation and Fos gene expression by transcriptional activation. Fos activates the mitochondrial apoptosis pathway by regulating Bim and Puma and then mediates GC-induced BMEC apoptosis, which ultimately leads to bone microcirculation dysfunction and the pathogenesis and progression of SONFH. Abbreviations: expression (EXP), silent (SIL), promoter (P)

Article Snippet: Then, TAF15 (D8V6Q) rabbit mAb (1:50; 28409; Cell Signaling Technology), Rpb1 NTD (D8L4Y) rabbit mAb (1:50; 14958; Cell Signaling Technology), and rabbit (DA1E) mAb IgG (1:50; 3900; Cell Signaling Technology) were added and incubated overnight at 4 °C.

Techniques: Gene Expression, Activation Assay, Expressing

Journal: Stem Cell Research & Therapy

Article Title: The circ_0042103/TAF15/NER axis regulates inflammation and DNA damage in pulpitis

doi: 10.1186/s13287-025-04817-1

Figure Lengend Snippet: Identification and validation of TAF15 as a circ_0042103-interacting protein. A circ_0042103 downstream target proteins were predicted by CircAtlas, Starbase, and RBPmap databases. B , C Perform KEGG and GO analysis on the overlapping predicted target proteins. D-F Western blot and quantification of protein levels of candidate RBPs (DGCR8, FUS, TAF15, IGF2BP2) in DPSCs after circ_0042103 knockdown or overexpression. G-H Relative mRNA expression of DGCR8, FUS, TAF15, and IGF2BP2 in DPSCs after circ_0042103 knockdown or overexpression, determined by qRT-PCR. I The binding of circ_0042103 to TAF15 was detected by RNA pull down. J Immunofluorescence showing nuclear and cytoplasmic distribution of TAF15 (red) and nuclei (DAPI, blue) in NC, LPS, si-NC + LPS, and si-circ_0042103 + LPS groups. Scale bars = 20 μm. K The nuclear and cytoplasmic distribution of circ_0042103 in hDPSCs was determined by qRT-PCR. U6 and GAPDH were used as nuclear and cytoplasmic controls, respectively. Data are shown as the mean ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: hDPSCs were fixed in 4% paraformaldehyde for 15 min, washed with PBS, permeabilized with 0.5% Triton X-100 for 10 min at room temperature, and blocked with 5% BSA for 1 h. Cells were incubated overnight at 4 °C with primary antibody against TAF15 (1:100, Proteintech, China), washed, and then incubated for 1 h at room temperature with Coralite 555 conjugated goat anti-rabbit IgG (1:100, Proteintech, China).

Techniques: Biomarker Discovery, Western Blot, Knockdown, Over Expression, Expressing, Quantitative RT-PCR, Binding Assay, Immunofluorescence

Journal: Stem Cell Research & Therapy

Article Title: The circ_0042103/TAF15/NER axis regulates inflammation and DNA damage in pulpitis

doi: 10.1186/s13287-025-04817-1

Figure Lengend Snippet: TAF15 inhibits DNA damage and inflammatory expression. A , B Transfection efficiency of knockdown and overexpression of TAF15 were detected by qRT-PCR. C circ_0042103 expression after TAF15 knockdown or overexpression. D , E Western blot and quantification of protein levels of γ-H2AX in DPSCs after LPS treatment and TAF15 knockdown. F , G IL6 and IL8 mRNA expression were assessed by qRT-PCR following LPS stimulation and TAF15 silencing. H , I IL6 and IL8 protein secretion levels were assessed by ELISA following LPS stimulation and TAF15 silencing. J , K Western blot and quantification of protein levels of γ-H2AX in DPSCs after LPS treatment and TAF15 overexpression. L-O The expression of IL6 and IL8 was assessed by qRT-PCR and ELISA following LPS stimulation and TAF15 overexpression. Data are shown as the mean ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: hDPSCs were fixed in 4% paraformaldehyde for 15 min, washed with PBS, permeabilized with 0.5% Triton X-100 for 10 min at room temperature, and blocked with 5% BSA for 1 h. Cells were incubated overnight at 4 °C with primary antibody against TAF15 (1:100, Proteintech, China), washed, and then incubated for 1 h at room temperature with Coralite 555 conjugated goat anti-rabbit IgG (1:100, Proteintech, China).

Techniques: Expressing, Transfection, Knockdown, Over Expression, Quantitative RT-PCR, Western Blot, Enzyme-linked Immunosorbent Assay

Journal: Stem Cell Research & Therapy

Article Title: The circ_0042103/TAF15/NER axis regulates inflammation and DNA damage in pulpitis

doi: 10.1186/s13287-025-04817-1

Figure Lengend Snippet: NER suppression by circ_0042103/TAF15 axis. A GSEA of the NER pathway in pulpitis tissue. B Protein-protein interaction (PPI) network of potential TAF15-interacting proteins. C , D Western blot and quantification of NER proteins ERCC1 and PCNA in LPS-treated DPSCs after circ_0042103 knockdown or overexpression. E , F Western blot and quantification of NER proteins ERCC1 and PCNA in LPS-treated DPSCs after TAF15 knockdown or overexpression. G , H qRT-PCR analysis of ERCC1 and PCNA mRNA in LPS-stimulated hDPSCs following TAF15 knockdown (si-TAF15) or overexpression (OE-TAF15). I The ERCC1 and PCNA mRNA stability assay with ActD ( n = 3). Data are shown as the mean ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: hDPSCs were fixed in 4% paraformaldehyde for 15 min, washed with PBS, permeabilized with 0.5% Triton X-100 for 10 min at room temperature, and blocked with 5% BSA for 1 h. Cells were incubated overnight at 4 °C with primary antibody against TAF15 (1:100, Proteintech, China), washed, and then incubated for 1 h at room temperature with Coralite 555 conjugated goat anti-rabbit IgG (1:100, Proteintech, China).

Techniques: Western Blot, Knockdown, Over Expression, Quantitative RT-PCR, Stability Assay

Journal: Stem Cell Research & Therapy

Article Title: The circ_0042103/TAF15/NER axis regulates inflammation and DNA damage in pulpitis

doi: 10.1186/s13287-025-04817-1

Figure Lengend Snippet: Mechanistic diagram of circ_0042103/TAF15 regulation of inflammation and DNA damage in hDPSCs (Created in BioRender. https://BioRender.com/6ai51mm )

Article Snippet: hDPSCs were fixed in 4% paraformaldehyde for 15 min, washed with PBS, permeabilized with 0.5% Triton X-100 for 10 min at room temperature, and blocked with 5% BSA for 1 h. Cells were incubated overnight at 4 °C with primary antibody against TAF15 (1:100, Proteintech, China), washed, and then incubated for 1 h at room temperature with Coralite 555 conjugated goat anti-rabbit IgG (1:100, Proteintech, China).

Techniques:

Journal: Biomedicines

Article Title: Structural and Functional Alterations in Mitochondria-Associated Membranes (MAMs) and in Mitochondria Activate Stress Response Mechanisms in an In Vitro Model of Alzheimer’s Disease

doi: 10.3390/biomedicines9080881

Figure Lengend Snippet: Primary antibodies utilized for Western blot.

Article Snippet: FIS-1 , 1:500 , Rabbit , Novus Biologicals , NB100-56646 , Littleton, CO, USA.

Techniques: Western Blot, Transduction

Journal: Cancer Research

Article Title: Diagnostic and Therapeutic Potential of a Human Antibody Cloned from a Cancer Patient That Binds to a Tumor-Specific Variant of Transcription Factor TAF15

doi: 10.1158/0008-5472.can-09-2186

Figure Lengend Snippet: Figure 2. Purification and identification of PAT-BA4 antigen by affinity chromatography and MALDI mass spectroscopy. A, protein purification of PAT-BA4 antigen from pooled membrane extracts of pancreas carcinoma cell line BxPC-3. Purified protein eluate was gained by affinity chromatography with crude membrane extracts. Western blot analysis was done with only secondary antibody, control IgG, and PAT-BA4 antibody. A Coomassie gel of purified protein eluate was performed. The protein band marked with an arrow was excised from Coomassie-stained gel. B, identification of the excised 78-kDa protein band by MALDI peptide mass mapping. The peaks with asterisks matched the calculated masses of tryptic peptides of human TAF15 (NP_631961.1 isoform 1/NP_003478.1 isoform 2). Thereby, the peptide mass error was <30 ppm and the minimum sequence coverage of the corresponding amino acids equates 22%.

Article Snippet: On both blots, the 78-kDa band was detected with anti-TAF15 antibody (3 μg/ mL; AVIVA Systems Biology).

Techniques: Purification, Affinity Chromatography, Mass Spectrometry, Protein Purification, Membrane, Western Blot, Control, Staining, Sequencing

Journal: Cancer Research

Article Title: Diagnostic and Therapeutic Potential of a Human Antibody Cloned from a Cancer Patient That Binds to a Tumor-Specific Variant of Transcription Factor TAF15

doi: 10.1158/0008-5472.can-09-2186

Figure Lengend Snippet: Figure 3. Validation of TAF15 as target for PAT-BA4 antibody by siRNA technology. Pancreas carcinoma cell line BxPC-3 was transfected with siRNA against human TAF15. Forty-eight hours after transfection, protein level of TAF15 was monitored by FACS analysis. Nontransfected cells and cells transfected with scrambled siRNA showed a positive binding of (A) PAT-BA4 antibody and (B) anti-CD55 antibody. TAF15 siRNA-transfected cells showed a reduced PAT-BA4 antibody binding but an unchanged binding of anti-CD55 antibody. Percentages of cell surface antigens of (C) PAT-BA4 and (D) anti-CD55 antibody are depicted. Columns, mean; bars, SD. *, P < 0.05.

Article Snippet: On both blots, the 78-kDa band was detected with anti-TAF15 antibody (3 μg/ mL; AVIVA Systems Biology).

Techniques: Biomarker Discovery, Transfection, Binding Assay

Journal: Cancer Research

Article Title: Diagnostic and Therapeutic Potential of a Human Antibody Cloned from a Cancer Patient That Binds to a Tumor-Specific Variant of Transcription Factor TAF15

doi: 10.1158/0008-5472.can-09-2186

Figure Lengend Snippet: Figure 4. Comparison of TAF15PAT-BA4 and wild-type TAF15 by Western blotting, FACS analysis, and immunohistochemical staining. A, Western blot inhibition assay with PAT-BA4 (300 μg/mL) and anti-TAF15 (3 μg/mL) antibody. One assay of each antibody was preincubated with the opponent antibody to check blocking effects; the other one was performed without the opponent antibody. B, FACS analyses with either PAT-BA4 (300 μg/mL) or commercial anti-TAF15 antibody (30 μg/mL; Santa Cruz Biotechnology) were performed on BxPC-3 cells and lymphocytes. Both assays were done with the same cell amount (2 × 105) and the same antibody concentrations. C, immunohistochemical staining of pancreas normal tissue and pancreas adenocarcinoma tissue. The paraffin-embedded sections were incubated with either PAT-BA4 (100 μg/mL) or anti-TAF15 antibody (1:25).

Article Snippet: On both blots, the 78-kDa band was detected with anti-TAF15 antibody (3 μg/ mL; AVIVA Systems Biology).

Techniques: Comparison, Western Blot, Immunohistochemical staining, Staining, Inhibition, Blocking Assay, Incubation

Journal: Cancer Research

Article Title: Diagnostic and Therapeutic Potential of a Human Antibody Cloned from a Cancer Patient That Binds to a Tumor-Specific Variant of Transcription Factor TAF15

doi: 10.1158/0008-5472.can-09-2186

Figure Lengend Snippet: Figure 5. Important characteristics of TAF15 protein and PAT-BA4 antibody. Expression of TAF15 on (A) protein and (B) mRNA level. A, Western blot analysis with anti-TAF15 (3 μg/mL) antibody on cell lysates of BxPC-3 cells and lymphocytes made of the same cell amount (1.2 × 106). B, semiquantitative PCR with mRNA from malignant and nonmalignant lung tissue. GAPDH mRNA level was used as internal standard. Functional assays with PAT-BA4 on BxPC-3 cells. C, histogram of adhesion assay with BxPC-3 cells. For tumor cell engraftment, the BxPC-3 cells were incubated with PAT-BA4 antibody. Chrompure human IgG served as isotype control. The amount of adherent cells was determined after different periods of time. Untreated cells grown in complete RPMI 1640 were adjusted to 100%. D, motility assay with BxPC-3 cells. The tumor cells were grown in 24-well plates to a confluence of 80%. A cross was then scratched into the cell monolayer and PAT-BA4 antibody was added. Complete RPMI 1640 and unrelated human IgG served as negative controls. Columns, mean; bars, SD. *, P < 0.05.

Article Snippet: On both blots, the 78-kDa band was detected with anti-TAF15 antibody (3 μg/ mL; AVIVA Systems Biology).

Techniques: Expressing, Western Blot, Functional Assay, Cell Adhesion Assay, Incubation, Control, Motility Assay