Journal: bioRxiv

Article Title: High throughput screening of eukaryotic release factor 1 variants to enhance noncanonical amino acid incorporation

doi: 10.1101/2025.09.10.675417

Figure Lengend Snippet: A) Schematic of the Tet-off strain YTH41, which enables endogenous SUP45 expression to be shut off in the presence of doxycycline. This facilitates evaluation of the ncAA incorporation phenotypes of eRF1 variants expressed from plasmids. B) Investigation of ncAA incorporation in YTH41 transformed with WT, N58A, or BR3T.4 (isolated triple mutant with N64D, K105R, and P116L) and AltTAG reporter. Median fluorescence intensity (MFI) of full-length reporter protein in BFP-positive cells. Inductions of reporters were performed in the presence of 1 mM OmeY and either in the presence or absence of 10 μg/mL doxycycline (dox). One-way ANOVA statistical analysis shown with the following p -value assignments: **** p ≤ 0.0001, ** p ≤ 0.01. WT clones were analyzed in biological duplicate and mutant clones were analyzed in biological triplicate. Flow cytometry dot plots can be found in Fig. S11. C) Schematic of the knockout strains ΔPAQ1 and ΔYIL014C-A derived from BY4741. Relative readthrough efficiencies (RRE) of cells transformed with wild type (WT) eRF1, S30P (single mutant clone) and BR3T.4 in yeast strains BY4741, BY4741ΔPAQ1, and BY4741ΔYIL014C-A. D) Median Fluorescence Intensity (MFI) of C-terminal (GFP) detection in BFP-positive cells. One-way ANOVA statistical analysis shown with the following p -value assignments: **** p ≤ 0.0001, *** p ≤ 0.001, * p ≤ 0.05, ns p > 0.05. For panels (C) and (D), three biological replicates were used for each condition investigated.

Article Snippet: O -Methyl-L-tyrosine (OmeY) was purchased from Chem-Impex International.

Techniques: Expressing, Transformation Assay, Isolation, Mutagenesis, Fluorescence, Clone Assay, Flow Cytometry, Knock-Out, Derivative Assay

Journal: Regenerative Therapy

Article Title: Orexin-A increases the differentiation of human olfactory sensory neurons through orexin receptor type 1

doi: 10.1016/j.reth.2024.10.014

Figure Lengend Snippet: Immunofluorescence of orexin receptors and dose-response analyses of orexin-A. (a) Immunofluorescence images of OSNs stained with OX1R and OX2R (green). (b) Schematic showing the proposed mechanism of orexin-mediated differentiation of OSNs and dose-response analyses of suvorexant. Western blot densitometric analysis of (c) ASCL1, (d) βIII Tubulin, and (e) OMP, respectively. (p > 0.05, compared to controls, n = 5).

Article Snippet: The orexin receptor type 1 (OX1R) antagonist, SB-674042 (HY-10898, MedChem Express, NJ, USA), and orexin receptor type 2 (OX2R) antagonist, TCS-OX2-29 (HY-100452, MedChem Express) were diluted in the dimethyl sulfoxide, DMSO with 1 μg/mL and 10 μg/mL, respectively.

Techniques: Immunofluorescence, Staining, Western Blot

Journal: Regenerative Therapy

Article Title: Orexin-A increases the differentiation of human olfactory sensory neurons through orexin receptor type 1

doi: 10.1016/j.reth.2024.10.014

Figure Lengend Snippet: The effect of OX1R antagonists (SB674042) and OX2R antagonists (TCS-OX2-29) on the differentiation of OSN. The expression of various markers was assessed through western blots (a and g) and subsequently analyzed densitometrically for ADCY3, Golf, ASCL1, βIII tubulin, and OMP (b-f and h-l). (Asterisks indicate p < 0.05, compared to controls, n = 5–6).

Article Snippet: The orexin receptor type 1 (OX1R) antagonist, SB-674042 (HY-10898, MedChem Express, NJ, USA), and orexin receptor type 2 (OX2R) antagonist, TCS-OX2-29 (HY-100452, MedChem Express) were diluted in the dimethyl sulfoxide, DMSO with 1 μg/mL and 10 μg/mL, respectively.

Techniques: Expressing, Western Blot