Journal: Scientific Reports

Article Title: Oligogenic effect is associated with the clinical heterogeneity of autosomal dominant deafness-15

doi: 10.1038/s41598-025-85881-8

Figure Lengend Snippet: The pedigree and segregation pattern of the ADNSHL family. P(+/−): The heterozygous mutation of POU4F3 . P(−/−): Wild type of POU4F3 . S(+/−): The heterozygous mutation of STRC . G(+/−): The heterozygous mutation of GJB2 . C(+/−):The heterozygous mutation of CDC14A . # Individuals (III-7, IV-1) with a faster progression of hearing loss and an earlier age of onset in this family. *The individuals who carried the mutation of POU4F3 but have not yet experienced hearing loss.

Article Snippet: Proteins were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis with 8% acrylamide, then the transferred and blocked protein was incubated with rabbit polyclonal antibody to STRC (Biorbyt Ltd., Cambridge, UK), mouse monoclonal antibody to GJB2 (Invitrogen, Carlsbad, CA, USA), rabbit polyclonal antibody to CDC14A (Invitrogen, Carlsbad, CA, USA) and rabbit polyclonal antibody to Vinculin(GeneTex, Irvine, CA, USA ) at 1:1000 dilution, followed by peroxidase-conjugated affinipure goat anti-mouse/rabbit IgG (Jackson ImmunoResearch, USA) at a 1:10000 dilution.

Techniques: Mutagenesis

Journal: Scientific Reports

Article Title: Oligogenic effect is associated with the clinical heterogeneity of autosomal dominant deafness-15

doi: 10.1038/s41598-025-85881-8

Figure Lengend Snippet: POU4F3 promoted the expression by activating the promotors of STRC , GJB2 and CDC14A. ( A ) mRNA expression analyses of STRC , GJB2 and CDC14A by RT-qPCR after POU4F3 overexpression. The results revealed that compared with the blank vector group, overexpression of wild type POU4F3 in 293T cell significantly increased the mRNA transcription of endogenous STRC (1.83 fold, P-values = 0.0007, F = 31.55, df = 8), GJB2 (1.41 fold, P-values < 0.0001, F = 64.47, df = 8) and CDC14A (1.31 fold, P-values = 0.018, F = 8.44, df = 8), while overexpression of the mutant POU4F3 did not change the mRNA levels of three genes. ( B ) mRNA expression analyses of STRC , GJB2 and CDC14A by RT-qPCR after POU4F3 knockdown by transfected with siRNA2. After knock-down of the POU4F3, the mRNA transcription of STRC, GJB2 and CDC14A was accordingly reduced to 63% (P-values = 0.0048, t = 7.560, df = 3), 79% (P-values = 0.025, t = 4.178, df = 3) and 80% (P-values = 0.0004, t = 18.37, df = 3) of untreated 293T cells respectively. ( C ) Protein expression analyses of CDC14A by western blot after POU4F3 overexpression. The results revealed that compared with the blank vector group, overexpression of wild type POU4F3 in 293T cell significantly increased the protein expression of endogenous CDC14A (2.02 fold, P-values = 0.0009, F = 28.63, df = 8). D. Protein expression analyses of CDC14A by western blot after POU4F3 knockdown by transfected with siRNA2. After knock-down of the POU4F3, the protein expression of CDC14A was accordingly reduced to 90% (P-values = 0.0061, t = 12.79, df = 2). NC negative control, transfection of empty vector, WT transfection of wild type POU4F3 vector, siRNA2 transfection of wild type siRNA2, L236F transfection of mutant POU4F3 vector. Values represent mean ± SD, significance was determined using 1way ANOVA Tukey’s Multiple Comparison Test and Paired t test. * P < 0.05, ** P < 0.01, *** P < 0.001. This experiment is repeated three times or more in biology.

Article Snippet: Proteins were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis with 8% acrylamide, then the transferred and blocked protein was incubated with rabbit polyclonal antibody to STRC (Biorbyt Ltd., Cambridge, UK), mouse monoclonal antibody to GJB2 (Invitrogen, Carlsbad, CA, USA), rabbit polyclonal antibody to CDC14A (Invitrogen, Carlsbad, CA, USA) and rabbit polyclonal antibody to Vinculin(GeneTex, Irvine, CA, USA ) at 1:1000 dilution, followed by peroxidase-conjugated affinipure goat anti-mouse/rabbit IgG (Jackson ImmunoResearch, USA) at a 1:10000 dilution.

Techniques: Expressing, Quantitative RT-PCR, Over Expression, Plasmid Preparation, Mutagenesis, Knockdown, Transfection, Western Blot, Negative Control, Comparison

Journal: Scientific Reports

Article Title: Oligogenic effect is associated with the clinical heterogeneity of autosomal dominant deafness-15

doi: 10.1038/s41598-025-85881-8

Figure Lengend Snippet: Dual luciferase reporter gene assay of STRC , GJB2 and CDC14A promotors. The results suggested that wild type POU4F3 significantly increased the promotor vectors’ luciferase activity of STRC (2.45 fold, P-values = 0.0029, F = 18.03, df = 8), GJB2 (2.40 fold, P-values < 0.0001, F = 66.55, df = 8) and CDC14A ( P-values = 0.0002, F = 46.37, df = 8) NC: negative control, cotransfection of empty vector and the promotor vector; WT: transfection of wild type POU4F3 vectorand the promotor vector; L236F: transfection of mutant POU4F3 vector and the promotor vector. This experiment is repeated three times or more in biology.

Article Snippet: Proteins were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis with 8% acrylamide, then the transferred and blocked protein was incubated with rabbit polyclonal antibody to STRC (Biorbyt Ltd., Cambridge, UK), mouse monoclonal antibody to GJB2 (Invitrogen, Carlsbad, CA, USA), rabbit polyclonal antibody to CDC14A (Invitrogen, Carlsbad, CA, USA) and rabbit polyclonal antibody to Vinculin(GeneTex, Irvine, CA, USA ) at 1:1000 dilution, followed by peroxidase-conjugated affinipure goat anti-mouse/rabbit IgG (Jackson ImmunoResearch, USA) at a 1:10000 dilution.

Techniques: Luciferase, Reporter Gene Assay, Activity Assay, Negative Control, Cotransfection, Plasmid Preparation, Transfection, Mutagenesis

Journal: Scientific Reports

Article Title: Oligogenic effect is associated with the clinical heterogeneity of autosomal dominant deafness-15

doi: 10.1038/s41598-025-85881-8

Figure Lengend Snippet: EMSA results of direct interaction between POU4F3 and the putative binding sites. STRC-P1, STRC-P2, GJB2-P1, GJB2-P2, CDC14A-P1 and CDC14A-P2: the detection probe which were labeled by biotin at the 5’ end. NC nuclear/cytoplasmic protein: nuclear/cytoplasmic protein of empty vector transfected cell. WT nuclear/cytoplasmic protein: nuclear/cytoplasmic protein of wild type POU4F3 vector transfected cell. L236F nuclear/cytoplasmic protein: nuclear/cytoplasmic protein of mutant POU4F3 vector transfected cell. This experiment is repeated three times or more in biology.

Article Snippet: Proteins were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis with 8% acrylamide, then the transferred and blocked protein was incubated with rabbit polyclonal antibody to STRC (Biorbyt Ltd., Cambridge, UK), mouse monoclonal antibody to GJB2 (Invitrogen, Carlsbad, CA, USA), rabbit polyclonal antibody to CDC14A (Invitrogen, Carlsbad, CA, USA) and rabbit polyclonal antibody to Vinculin(GeneTex, Irvine, CA, USA ) at 1:1000 dilution, followed by peroxidase-conjugated affinipure goat anti-mouse/rabbit IgG (Jackson ImmunoResearch, USA) at a 1:10000 dilution.

Techniques: Binding Assay, Labeling, Plasmid Preparation, Transfection, Mutagenesis

Journal: Biomedicines

Article Title: Recent Therapeutic Progress and Future Perspectives for the Treatment of Hearing Loss

doi: 10.3390/biomedicines11123347

Figure Lengend Snippet: Preclinical therapeutic pipeline for inner ear hearing disorders.

Article Snippet: Genetic ( STRC -mediated HL) , Gene therapy , AAV.104 , Undisclosed , Decibel Therapeutics (collaboration with Regeneron) , AAV STRC gene transfer to selectively target outer HCs.

Techniques: Formulation, Inhibition