Journal: Nucleic Acids Research

Article Title: Transient SUMOylation inhibition in human pre-adipocytes stably imprints a transcriptional beiging fate

doi: 10.1093/nar/gkag232

Figure Lengend Snippet: Effect of transient SUMOylation inhibition in pre-adipocytes on cAMP-PKA-p38 signaling and adaptive thermogenesis in mature adipocytes. ( A ) Heatmap showing the stable upregulation of most genes involved in adaptive thermogenesis and cAMP-PKA-p38 signaling 22 days after adipogenic induction. Z -scores were calculated and plotted in GraphPad Prism. Treatments of pre-adipocytes were performed as shown in Fig. . ( B ) Western blot analysis of PKA substrates phosphorylation, assessed using a pan-PKA-target antibody, 22 days after adipogenic induction. TBP was used as a loading control. The asterisk (*) indicates substrates significantly affected by TAK-981 and/or rosiglitazone treatment. ( C ) Quantification of PKA substrate signals from panel (B). The signal for PKA substrates and TBP was quantified using Fiji software, with PKA substrate signals normalized to TBP. Error bars represent the standard deviation of four independent experiments. Student’s t -test was used to calculate P -values. ( D ) Western blot analysis of p-CREB, p-ATF1, p38, p-p38, and p-ATF2, 22 days after adipogenic induction. TBP was used as a loading control. ( E ) Quantification of p-CREB/ATF-1 signals from panel (D). Signals were normalized to TBP. Error bars represent the standard deviation of three independent experiments. Student’s t -test was used to calculate P -values. ( F ) Western blot analysis of p-CREB in the absence or presence of the PKA inhibitor RP-8-CPT-cAMPS. TBP was used as a loading control. Quantification of p-CREB, normalized to TBP, is showed in the lower panel. Error bars represent the standard deviation of two independent experiments. Quantification of p-p38 ( G ) and p-ATF2 ( H ) from experiment in panel (D). Signals were normalized to TBP for p-ATF2 and to p38 for p-p38. Error bars represent the standard deviation of three independent experiments. Student’s t -test was used to calculate P -values.

Article Snippet: The following antibodies were used: UCP1, abcam, ab209483; PKA phospho-substrates, Cell Signaling, 9624; p-CREB/p-ATF1, Cell Signaling, 9198; p38 MAPK, Cell Signaling, 9212; p-p38 MAPK, Cell Signaling, 9211; p-ATF2, Cell Signaling, 24329; SUMO2/3, abcam, ab81371; PPARG, Cell signaling, 2443; and TBP, Protein Tech, 22006-1-AP or abcam, 282715; γ-tubulin, Sigma, T5326: CEBPB, Santa Cruz, sc-7962 quantifications were performed using FiJi [ ].

Techniques: Inhibition, Western Blot, Phospho-proteomics, Control, Software, Standard Deviation

Journal: iScience

Article Title: THBS1 upregulation by KLF7 in hypopharyngeal squamous cell carcinoma contributes to lung metastasis through the p38 MAPK signaling

doi: 10.1016/j.isci.2026.114816

Figure Lengend Snippet: The p38 MAPK activator LX-3 rescues the malignant behavior of HPSCC cells repressed by the silencing of THBS1 (A) Western blot analysis of the extent of p38, ATF2, and MK2 phosphorylation, as well as total p38 expression in KD-THBS1-treated HPSCC cells. (B) Western blot analysis of the extent of p38, ATF2, and MK2 phosphorylation, as well as total p38 expression in HPSCC cells treated with 2 μM LX-3 after knockdown of THBS1. (C–E) HPSCC cell proliferative capacity was examined using EdU staining. Transwell assay detects changes in the migratory (D) and invasive (E) abilities of HPSCC cells. (F) Expression of E-cadherin and N-cadherin in HPSCC cells was examined using dual-labeling immunofluorescence. The results are expressed as the mean ± SD (A–F). These values are derived from three independent experiments. Statistical analysis was performed using an unpaired t test.

Article Snippet: The primary antibodies used were as follows: anti-THBS1 (1:250, A2125, BioAb), anti-E-cadherin (1:1000, 3195, Cell Signaling Technologies), anti-N-cadherin (1:100, PA5-19486m Invitrogen), anti-Phospho-p38 MAPK (Thr180/Tyr182) (1:1000, 4511, Cell Signaling Technologies), anti-total p38 (1:1000, 9212, Cell Signaling Technologies), anti- p -ATF2 (Thr71) (1:1000, 44-295G, Invitrogen), anti-p-MK2 (Thr334) (1:1000, 3007, Cell Signaling Technologies), anti-KLF7 (1:1000, CSB- PA126129 , Cusabio), anti-Slug (1:1000, 12129-1-AP, ProteinTech Group), anti-Vimentin (1:20000, 10366-1-AP, ProteinTech Group), anti-HDAC6 (1:1000, 12834-1-AP, ProteinTech Group), and anti-GAPDH (1:20000, AC001, BioAb).

Techniques: Western Blot, Phospho-proteomics, Expressing, Knockdown, Staining, Transwell Assay, Labeling, Immunofluorescence, Derivative Assay

Journal: iScience

Article Title: THBS1 upregulation by KLF7 in hypopharyngeal squamous cell carcinoma contributes to lung metastasis through the p38 MAPK signaling

doi: 10.1016/j.isci.2026.114816

Figure Lengend Snippet: Reactivation of THBS1 rescues the HPSCC cell malignant biological behavior curtailed by knockdown of KLF7 (A) Western blot analysis of protein expression of KLF7 and THBS1 in HPSCC cells treated with knockdown of KLF7 combined with the overexpression of THBS1. (B–D) HPSCC cell proliferative capacity was examined using EdU staining. Transwell assay detects changes in the migratory (C) and invasive (D) abilities of HPSCC cells. (E) Expression of E-cadherin and N-cadherin in HPSCC cells was examined using dual-labeling immunofluorescence. (F) Western blot analysis of the extent of p38, ATF2, and MK2 phosphorylation, as well as total p38 expression in HPSCC cells. The results are expressed as the mean ± SD (A–F). These values are derived from three independent experiments. Statistical analysis was performed using a one-way ANOVA followed by Tukey’s multiple comparison test.

Article Snippet: The primary antibodies used were as follows: anti-THBS1 (1:250, A2125, BioAb), anti-E-cadherin (1:1000, 3195, Cell Signaling Technologies), anti-N-cadherin (1:100, PA5-19486m Invitrogen), anti-Phospho-p38 MAPK (Thr180/Tyr182) (1:1000, 4511, Cell Signaling Technologies), anti-total p38 (1:1000, 9212, Cell Signaling Technologies), anti- p -ATF2 (Thr71) (1:1000, 44-295G, Invitrogen), anti-p-MK2 (Thr334) (1:1000, 3007, Cell Signaling Technologies), anti-KLF7 (1:1000, CSB- PA126129 , Cusabio), anti-Slug (1:1000, 12129-1-AP, ProteinTech Group), anti-Vimentin (1:20000, 10366-1-AP, ProteinTech Group), anti-HDAC6 (1:1000, 12834-1-AP, ProteinTech Group), and anti-GAPDH (1:20000, AC001, BioAb).

Techniques: Knockdown, Western Blot, Expressing, Over Expression, Staining, Transwell Assay, Labeling, Immunofluorescence, Phospho-proteomics, Derivative Assay, Comparison

Journal: iScience

Article Title: THBS1 upregulation by KLF7 in hypopharyngeal squamous cell carcinoma contributes to lung metastasis through the p38 MAPK signaling

doi: 10.1016/j.isci.2026.114816

Figure Lengend Snippet: Activation of the THBS1/p38 MAPK axis by KLF7 promotes the lung metastasis of HPSCC cells (A) Lung metastasis in nude mice injected with FaDu cells in the tail vein was assessed by HE staining. (B–D) Quantitative results of lung metastatic nodules. IHC scores (C) and protein expression (D) of Slug and Vimentin within lung metastases in nude mice. (E) RT-qPCR detection of mRNA expression levels of KLF7 and THBS1 in lung metastases. (F) IHC scores of KLF7 and THBS1 within lung metastases in nude mice. (G) Western blot analysis of the extent of p38, ATF2, and MK2 phosphorylation, as well as total p38 expression in lung metastases. The results are expressed as the mean ± SD (A–G). These values are derived from five mice. Statistical analysis was performed using a one-way ANOVA followed by Tukey’s multiple comparison test.

Article Snippet: The primary antibodies used were as follows: anti-THBS1 (1:250, A2125, BioAb), anti-E-cadherin (1:1000, 3195, Cell Signaling Technologies), anti-N-cadherin (1:100, PA5-19486m Invitrogen), anti-Phospho-p38 MAPK (Thr180/Tyr182) (1:1000, 4511, Cell Signaling Technologies), anti-total p38 (1:1000, 9212, Cell Signaling Technologies), anti- p -ATF2 (Thr71) (1:1000, 44-295G, Invitrogen), anti-p-MK2 (Thr334) (1:1000, 3007, Cell Signaling Technologies), anti-KLF7 (1:1000, CSB- PA126129 , Cusabio), anti-Slug (1:1000, 12129-1-AP, ProteinTech Group), anti-Vimentin (1:20000, 10366-1-AP, ProteinTech Group), anti-HDAC6 (1:1000, 12834-1-AP, ProteinTech Group), and anti-GAPDH (1:20000, AC001, BioAb).

Techniques: Activation Assay, Injection, Staining, Expressing, Quantitative RT-PCR, Western Blot, Phospho-proteomics, Derivative Assay, Comparison