Journal: Cancer biology & therapy

Article Title: CircATP5C1 promotes triple-negative breast cancer progression by binding IGF2BP2 to modulate CSF-1 secretion.

doi: 10.1080/15384047.2025.2479926

Figure Lengend Snippet: Figure 6. circATP5C1 increases the stability of CSF-1 mRNA through interaction with insulin-like growth factor 2 mRNA binding protein 2 (IGF2BP2). (a) RNA pull-down assay and silver staining were conducted to visualize the possible binding proteins in circATP5C1 probe. (b) RNA pull-down-western blot showed IGF2BP2 could bind with circATP5C1. (c) The specific peptide map for IGF2BP2 identified by mass spectrometry. (d) RNA fish-immunofluorescence in MDA-MB-231 and MDA-MB-468 cells exhibited the colocalization of circATP5C1 with IGF2BP2. (e) RNA immunoprecipitation (RIP) assays validate binding of circATP5C1 to IGF2BP2 protein. (f) RIP assays identified the interaction of IGF2BP2 protein with CSF-1 mRNA. (g) Knockdown of IGF2BP2 rescued the mRNA and protein levels of CSF-1 caused by circATP5C1 overexpression in MDA-MB-468 and HS-578T cells. (h) MeRIP assays validated CSF-1 mRNA contains m6A methylation. (i) Stability of CSF-1 mRNA was detected by rt- qPCR after actinomycin D treatment in transfected MDA-MB-231 and MDA-MB-468 cells. *P < 0.05, **P < 0.01, ***P < 0.001.

Article Snippet: The magnetic beads were incubated with 5 μg antibody against IGF2BP2 (proteintech, China) or anti-m6A antibody (proteintech, China), or control IgG (proteintech, China).

Techniques: Binding Assay, Pull Down Assay, Silver Staining, Western Blot, Mass Spectrometry, Immunofluorescence, RNA Immunoprecipitation, Knockdown, Over Expression, Methylation, Quantitative RT-PCR, Transfection

Journal: Frontiers in Pharmacology

Article Title: DNA methylation and immune regulation in osteoporosis: emerging epigenetic targets for drug discovery

doi: 10.3389/fphar.2025.1688305

Figure Lengend Snippet: The Triad of Osteoporosis: Pathophysiology, Molecular Pathways, and Treatment Approaches. From healthy bones to the development of osteoporosis. It integrates various factors: aging, hormonal abnormalities, inflammation, etc. It involves multi-layered mechanisms: from immune-bone interaction regulation to epigenetic regulation mediated by DNA methylation, analyzing the molecular mechanisms of bone metabolism disorders. Specifically, DNA methylation mediated by DNMT1/3A/3B exerts dual regulatory effects: on one hand, it targets and modulates key bone metabolism-related genes such as Runx2, SOST, and NFATc1, as well as signaling pathways including RANKL/RANK and Wnt/β-Catenin; on the other hand, it influences immune-osseous signaling pathways involving M1/M2 macrophages, T cells, IL-1RN, and NF-κB. Both regulatory effects ultimately drive the imbalance between bone resorption and bone formation. It covers multiple therapeutic strategies: various existing treatments, the potential use of DNMTi. And correlates bone turnover markers (BTM) with the diagnostic value of DNA methylation biomarkers.

Article Snippet: (Cell Death Dis) , Mouse and human cells , NFATc1 (RNA m6A; METTL14/YTHDF2) , Exosome-delivered METTL14 ↑ m6A at NFATc1 (4249A) → mRNA decay , Inhibits osteoclast resorption, preserves bone , .

Techniques: DNA Methylation Assay, Protein-Protein interactions, Diagnostic Assay

Journal: Theranostics

Article Title: SUMO1 modification of methyltransferase-like 3 promotes tumor progression via regulating Snail mRNA homeostasis in hepatocellular carcinoma.

doi: 10.7150/thno.42539

Figure Lengend Snippet: Figure 5. SUMOylation of Mettl3 regulates Snail mRNA homeostasis via m6A methyltransferase. (A) Mettl3-WT or -KR was transiently transfected into MHCC97H cells and detected by the dot-blot assay with the anti-m6A antibody. Equal loading of mRNAs was confirmed by methylene blue staining. (B) Mettl3 with or without Ubc9 was transfected into MHCC97H cells and detected by the dot-blot assay with the anti-m6A antibody. Equal loading of mRNAs was confirmed by methylene blue staining. (C) Suppression or overexpression of Mettl3 in HCC was determined by RT-qPCR, and GAPDH was used as the normalized control. (D) mRNA of Snail in Mettl3 suppression or overexpression cells. (E) Mettl3-WT or Mettl3-KR-expressing cells were transfected with the pEZX-PL01-Snail promoter reporter plasmid and negative control plasmid for 36 h. Results were expressed as the ratios between F-luc and R-luc activities. (F) IP immunoblot analysis was performed in Mettl3-WT- and Mettl3-KR-expressing cells with the anti-Mettl3 antibody, followed by western blotting with Mettl3, anti-Eef2, anti-eIF4E, and anti-NCBP1 antibodies. One-tenth of lysates as the input was immunoblotted with indicated antibodies. (G) Mettl3-WT- and Mettl3-KR-expressing cells were pretreated with MG-132 for 6 h and stimulated with serum for indicated times. Subsequently, Mettl3 and Snail protein expression levels were analyzed by western blotting. (H) Scramble or siMettl3-expressing cells were fed with CHX for the indicated times, and protein expression of Mettl3 and Snail was analyzed by western blotting. (I) Mettl3-WT or Mettl3-KR-expressing cells transfected with different plasmids were treated with CHX for the indicated times, and protein expression of Mettl3 and Snail was detected by western blotting. (J and K) The decay rate of mRNA and qPCR analysis of Snail at the indicated times after exposure to the transcription inhibitor actinomycin D (5 μg/mL) in MHCC97H (J) and HepG2 (K) cells. The relative expression level was normalized to β-actin. Data are presented as mean ± s.d. * p < 0.05, ** p < 0.01; Student’s t-test.

Article Snippet: The primary antibodies, including anti-GAPDH, anti-LaminB1, anti-His, anti-HA, anti-Flag, anti-MMP2 (from proteintech), anti-Mettl3 (from abcam, Bethyl); anti-m6a (from Synaptic Systems); SUMO-1 and SUMO2/3 (from abcam); anti-MMP9, anti-E-cadherin and normal rabbit IgG (from CST); UBC9, Snail and normal mouse IgG (from santa cruz); anti-rabbit, Anti-mouse peroxidase-conjugated secondary antibodies (from proteintech). m6A dot blot assay mRNA was enriched using Dynabeads mRNA DIRECT Purification Kit (InvitrogenTM, 61012).

Techniques: Transfection, Dot Blot, Staining, Over Expression, Quantitative RT-PCR, Control, Expressing, Plasmid Preparation, Negative Control, Western Blot