Fig 2: BRAF/MAPK/AP-1 axis promotes TBX3 transcription in PTC.a, b Western blot of TBX3 in PTC or normal thyroid cells (Nthy-ori-3-1) with BRAF or BRAFV600E over-expression (a) or K1 cells with BRAF knock-down (b). c K1 cells were treated with BRAF inhibitor PLX4032 or ERK1/2 inhibitor SCH772984 for 24 h, the level of TBX3 was analysis by western blot. d TBX3 and AP-1 factors levels in K1 cells over-expressing c-Jun, JunB, or c-Fos, and treated with PLX4032 or SCH772984. e TBX3 and AP-1 factors levels in K1 cells with BRAFV600E over-expression and AP-1 knock-down. f AP-1 binding motifs were analyzed through TBX3 promoter region using Jasper database. g Construction of TBX3 promoter GLuc with potential AP-1-binding sites. The predicted -358bp and -362bp site were overlapped as -360bp site. Arrows represented primers were used for qPCR. h, i Promoter GLuc activities were analyzed in HEK293T cells co-transfected with TBX3 truncated promoter (h) or TBX3-149-MUT promoter (i) and AP-1 expression constructs. j Quantitative ChIP (qChIP) analysis with anti-IgG, c-Jun, JunB and c-Fos for -149 site in K1 cells, primers targeting GAPDH gene were used as control. k Luciferase activities of GLuc-149 were analyzed in K1 cells treated with PLX4032 or SCH772984 for 12 h, 24 h or 36 h. l THCA datasets were downloaded from TCGA database to analyze the correlation between TBX3 and BRAF, n = 501. Densitometric analyses of western blot were shown (a–e). Two independent experiments were carried out with similar results for each kind of cells (a–c), and n = 3 biological independent samples (d, e, h–k). Data are shown as the mean ± s.d. (h–k). P values were calculated by unpaired two-tailed Student’s t test (h–k). Uncropped immunoblots and statistical source data are provided in Source Data.

Fig 3: JunB expression during skin maturation and upon stress. a Representative microphotographs with double immunostaining for JunB (green) and FABP5 (red), indicative of sebaceous glands, in skin derived from 1, 3, and 5 days old mice. Inset showing magnified view of a sebaceous gland. Scale bars, 50 µm. b Immunostaining of JunB (green) and FABP5 (red) in 60 days old mice. Inset showing magnified view. c Representative microphotographs with double immunostaining of JunB (green) and FABP5 (red) in dorsal skin of 60 days old mice following hair plucking or d after topical TPA application, a potent inducer of proliferation. e Immunostaining of JunB (green) and LRIG1 or CD34 (red) in dorsal skin of 60 days old mice following hair plucking. f Immunostaining of JunB (green) and FABP5 (red) in adult human skin. E, epidermis; D, dermis; HF, hair follicle; SG, sebaceous gland; SD, sebaceous duct; HS, hair shaft. Asterisk indicates hair shaft autofluorescence. Dashed line indicates the epidermal-dermal junction. Scale bars, 50 µm

Fig 4: Differential expression analysis of the dSPN proteome following CNO-induced activation of hM3Dq.a Experimental paradigm. Targeted co-expression of APEX and hM3Dq or fluorophore control in Drd1Cre+ dSPNs. Clozapine N-oxide (CNO) was administered (3 mg/kg i.p.) to selectively activate dSPNs via Gaq-coupled signaling cascades. Following CNO treatment, acute slice preparation and ex vivo biotinylation were performed to measure proteomic changes at 2.5–3 h post CNO administration. Some schematics were created with BioRender.com. b Western blot analysis of biotinylated striatal lysates for samples used in this dataset. Western blot analysis of these samples was performed once prior to orthogonal confirmation by proteomics. c Statistical analysis workflow for comparing the effect of CNO-induced changes in the dSPN proteome between hM3Dq-expressing and fluorophore control samples. d Hierarchical clustering heatmap of proteins and biological replicates. e Gene ontology analysis of H2B-enriched and NES-enriched protein lists. f Scatter plot of log2FC (mCherry-NES) vs log2FC (hM3Dq-NES) for the H2B-enriched nuclear proteome. Blue and red dots are downregulated and upregulated proteins, respectively (q value < 0.05). g Differential expression analysis for the nuclear proteome (hM3Dq – mCherry) (dotted line: q value = 0.05). h Differential expression analysis for the cytosolic proteome (hM3Dq – mCherry) (dotted line: q value = 0.05). i Immunostaining of JunB in the striatum 2.5 h post CNO administration (20 µm). n = 3 animals from two independent experiments. j Reporter ion quantification of JunB interacting proteins that were identified only by one unique peptide. For each protein, n reflects the number of samples where this protein was quantified, for mCherry and hM3Dq conditions, respectively: Fos n = 1, 4; Fosb n = 7, 7; Jund n = 4, 4; Fosl2 n = 2, 4; Pkia n = 3, 3; Smad3 n = 4, 4; and Mapk8 n = 4, 4. The upper and lower bounds of the box represent 75th and 25th percentiles, whiskers extend from minimum to maximum. The horizontal center line is the median. k Cytoscape network of JunB protein interactions detected in the dataset (color: log2FC (hM3Dq – mCherry)). Source data are provided as a Source Data file.

Fig 5: H2S regulates TNFSF14 transcription through JNK/JunB signal.A Schematic overview of the predicted transcription factors of TNFSF14 gene by PROMO dataset (upper) and JunB binding sites on TNFSF14 promoter predicted with JASPAR website (lower). B ChIP assays for JunB binding onto TNFSF14 promoter were carried out in HepG2 (upper) and PLC/PRF/5 (lower) cells. The right panel showed the statistical analysis for the ChIP assay. C NaHS was used to treat HepG2 (left) and PLC/PRF/5 (right) cells for 0, 1, 2, 8, and 24 h. Expression of p-JunB in nuclear of HCC cells was detecting using immunofluorescence staining, and Nuclei and p-JunB were stained with DAPI (blue) and FITC (green), respectively. The image is representative of three independent experiments. D The effects of different time of NaHS treatment on the expression of p-JunB in the nuclear and cytoplasm were analyzed with Western blotting in the HepG2 cells and PLC/PRF/5 cells. E The expression levels of p-JunB protein were detected by Western blotting in the HCC cells under exogenous and endogenous H2S treatment. F Western blotting for the effects of PPG (CSE inhibitor) and SP600125 (p-JNK inhibitor) in the expressions of JNK/p-JNK, JunB/p-JunB, and TNFSF14 proteins in HepG2 and PLC/PRF/5 cells. All Bars, ± SD; n = 3; *P < 0.05 compared with control group in HepG2 cells, #P < 0.05 compared with control group in PLC/PRF/5 cells.