p38γ inhibitors increase olfactory transduction pathway activity. (A) WB analysis of Hut78 cells treated with 0.5 to 10 μM of CSH18. (B) WB analysis of Hut78 cells treated with 0.5 to 10 μM of CSH71. Both CSH18 and CSH71 increased OR4N5 protein levels. Other olfactory factors OR10K2, OR2H2, and OR4K17 were also assessed, with dimerization of OR4N5 observed based on molecular weight (blue arrows indicate 50 kDa). (C) WB assays for lentiviral expression of sh-p38γ vs sh-Ctrl. OR4N5 bands at ∼30 and 60 kDa correspond to the monomer and dimer, respectively (sh-Ctrl lane). (D) OR4N5 3D structure prediction by AlphaFold (AF-Q8IXE1-F1, left). Right: localization of OR4N5 in the plasma membrane (green, confidence score = 5). Slight presence in the cytoskeleton (confidence score = 1) according to Genecard.com. (E) In-depth analysis of the OR4N5 amino acid sequence identified 7 transmembrane domains (underlined) and 2 cysteines that may contribute to OR4N5 dimerization. CDR, Complementarity Determining Region; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; hr, hour; sh-Ctrl, shRNAs targeting control; sh-p38γ, shRNAs targeting p38γ; TM, Transmembrane.
Figure 2.

p38γ inhibitors increase olfactory transduction pathway activity. (A) WB analysis of Hut78 cells treated with 0.5 to 10 μM of CSH18. (B) WB analysis of Hut78 cells treated with 0.5 to 10 μM of CSH71. Both CSH18 and CSH71 increased OR4N5 protein levels. Other olfactory factors OR10K2, OR2H2, and OR4K17 were also assessed, with dimerization of OR4N5 observed based on molecular weight (blue arrows indicate 50 kDa). (C) WB assays for lentiviral expression of sh-p38γ vs sh-Ctrl. OR4N5 bands at ∼30 and 60 kDa correspond to the monomer and dimer, respectively (sh-Ctrl lane). (D) OR4N5 3D structure prediction by AlphaFold (AF-Q8IXE1-F1, left). Right: localization of OR4N5 in the plasma membrane (green, confidence score = 5). Slight presence in the cytoskeleton (confidence score = 1) according to Genecard.com. (E) In-depth analysis of the OR4N5 amino acid sequence identified 7 transmembrane domains (underlined) and 2 cysteines that may contribute to OR4N5 dimerization. CDR, Complementarity Determining Region; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; hr, hour; sh-Ctrl, shRNAs targeting control; sh-p38γ, shRNAs targeting p38γ; TM, Transmembrane.

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