Journal: Nucleic Acids Research

Article Title: Reduced FBXO22 skews human trophoblast fate equilibrium toward syncytialization via polyubiquitinating the CoREST complex

doi: 10.1093/nar/gkag557

Figure Lengend Snippet: FBXO22 interacts directly with the CoREST complex. ( A ) Schematic diagram showing the workflow for the identification of FBXO22 interactors. ( B ) GO enrichment analysis of FBXO22-interacting nuclear proteins in hTSCs. ( C ) Venn diagram showing the nuclear proteins among the FBXO22 interactors. ( D ) Co-IP analysis of endogenous interactions between FBXO22 and the CoREST complex in hTSCs. ( E ) Co-IP analysis of exogenous interactions between FBXO22 and the CoREST complex in HEK293T cells (293T). ( F ) Co-IP analysis of endogenous interactions between LSD1, HDAC1, RCOR1, and FBXO22 in hTSCs. ( G ) PLA showing the proximity between FBXO22 and CoREST in FBXO22 -KD cells and control hTSCs. ( H ) Western blot analysis of CoREST complex proteins in nuclear extracts from control and FBXO22 -KD hTSCs. H3 was used as the nuclear loading control; n = 3. ( I ) RT-PCR analysis of CoREST complex expression in FBXO22 -KD cells and control hTSCs. The values were normalized to GAPDH; n = 5. NS, no significance. Data are means ± SEM, and are representative of at least three independent experiments (D–G); ** P < 0.01, *** P < 0.001, **** P < 0.0001 by unpaired two-tailed t test. The Eppendorf tube images in Fig. 3A were adapted from SciDraw under the Creative Commons license CC BY 4.0.

Article Snippet: Briefly, reactions were assembled in 1.5-ml tubes in a final volume of 50 μl containing 14 μl of dH 2 O, 5 μl of 10× ubiquitinylation buffer, 2.5 μl of 20× ubiquitin, 2.5 μl of 20× E1 enzyme, 5 μl of 10× E2 conjugating enzyme, 5 μg of SCF FBXO22 E3 complex, and 5 μg of recombinant HDAC1, RCOR1, or LSD1 protein (custom-made, MedChemExpress).

Techniques: Co-Immunoprecipitation Assay, Control, Western Blot, Reverse Transcription Polymerase Chain Reaction, Expressing, Two Tailed Test

Journal: Nucleic Acids Research

Article Title: Reduced FBXO22 skews human trophoblast fate equilibrium toward syncytialization via polyubiquitinating the CoREST complex

doi: 10.1093/nar/gkag557

Figure Lengend Snippet: FBXO22-mediated ubiquitination of the CoREST complex orchestrates trophoblast syncytialization. ( A ) Western blot analysis of CoREST complex proteins and FBXO22 in FBXO22 -OE cells and control hTSCs treated with MG132 or 3-MA for 6 h. ( B ) Western blot analysis of FBXO22 and CoREST complex protein levels in hTSCs treated with cycloheximide (CHX) or CHX combined with MG132 for 0, 2, 4, 8, and 12 h. ( C ) Western blot analysis of FBXO22 and CoREST complex protein levels in FBXO22 -KD cells and control hTSCs following CHX treatment for 0, 2, 4, 8, and 12 h. ( D ) Western blot analysis of V5-LSD1, Myc-RCOR1, and Flag-HDAC1 protein levels with increasing doses of HA-FBXO22 expression plasmid in HEK293T cells. ( E ) Western blot analysis of CoREST complex ubiquitination in FBXO22 -OE cells and control hTSCs. ( F ) Western blot analysis of CoREST complex ubiquitination in HEK293T cells co-transfected with an FBXO22 expression plasmid and si- FBXO22 . ( G ) Immunofluorescence staining of CDH1 and CGB in hTSCs, hTSCs-derived STBs, and hTSCs-derived STBs treated with Romidepsin (2 nM) or GSK-LSD1 (1 μM) for 72 h. ( H ) Immunofluorescence staining of CDH1 and CGB in BeWo cells treated with DMSO, Forskolin (FSK), or FSK combined with Romidepsin (2 nM) or GSK-LSD1 (1 μM) for 48 h. Data are means ± SEM; GAPDH or β-actin was used as the loading control; the data presented are representative of at least three independent experiments; * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001 by unpaired two-tailed t test and one-way ANOVA.

Article Snippet: Briefly, reactions were assembled in 1.5-ml tubes in a final volume of 50 μl containing 14 μl of dH 2 O, 5 μl of 10× ubiquitinylation buffer, 2.5 μl of 20× ubiquitin, 2.5 μl of 20× E1 enzyme, 5 μl of 10× E2 conjugating enzyme, 5 μg of SCF FBXO22 E3 complex, and 5 μg of recombinant HDAC1, RCOR1, or LSD1 protein (custom-made, MedChemExpress).

Techniques: Ubiquitin Proteomics, Western Blot, Control, Expressing, Plasmid Preparation, Transfection, Immunofluorescence, Staining, Derivative Assay, Two Tailed Test

Journal: Nucleic Acids Research

Article Title: Reduced FBXO22 skews human trophoblast fate equilibrium toward syncytialization via polyubiquitinating the CoREST complex

doi: 10.1093/nar/gkag557

Figure Lengend Snippet: FBXO22 deficiency leads to CoREST complex accumulation and contributes to RPL. ( A ) Volcano plot of genes differentially expressed in first-trimester villi from normal pregnancies and in RPL patients ( P < 0.05, log 2 (fold change) > 0.8). ( B ) GO enrichment analysis of genes down-regulated in RPL patients. ( C ) Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis of genes downregulated in RPL patients. ( D ) Venn diagram showing the overlap between the LSD1-interactome, transcriptome downregulated genes, and human ubiquitin ligases. ( E ) PLA assay showing the proximity between FBXO22 and CoREST in the early villous tissues. CDH1 and CGB were used as markers for the CTBs and STBs, respectively. The data presented are representative of at least three independent experiments. ( F ) Western blot analysis of FBXO22 and CoREST complex protein levels in normal pregnancies and in RPL patients. Cytokeratin 7 (CK7) was used as the trophoblast marker control; n = 5. ( G ) Immunohistochemical analysis of FBXO22 in the early villous tissues from normal pregnancies and from RPL patients. TEAD4 and CGB were used as markers for CTBs and STBs, respectively. ( H ) Immunohistochemical analysis of the CoREST complex in the early villous tissues from normal pregnancies and from RPL patients. ( I ) The expression levels and correlation analysis of FBXO22 and the CoREST complex in early villous tissues from normal pregnancies and from RPL patients; n = 12 for Nor, and n = 10 for RPL. Data are means ± SEM; **** P < 0.0001 by unpaired two-tailed t test.

Article Snippet: Briefly, reactions were assembled in 1.5-ml tubes in a final volume of 50 μl containing 14 μl of dH 2 O, 5 μl of 10× ubiquitinylation buffer, 2.5 μl of 20× ubiquitin, 2.5 μl of 20× E1 enzyme, 5 μl of 10× E2 conjugating enzyme, 5 μg of SCF FBXO22 E3 complex, and 5 μg of recombinant HDAC1, RCOR1, or LSD1 protein (custom-made, MedChemExpress).

Techniques: Ubiquitin Proteomics, Western Blot, Marker, Control, Immunohistochemical staining, Expressing, Two Tailed Test

Journal: bioRxiv

Article Title: Workload-induced changes to cell state contribute to β-cell failure in diabetes

doi: 10.64898/2026.05.13.725004

Figure Lengend Snippet: (a) Schematic of alleles and treatments used to inactivate Lsd1 in db/db mice. TM, tamoxifen; wks, weeks. (b) Time course of ad libitum-fed blood glucose levels in TM-treated mice of the indicated genotypes. db/+ Lsd1 fl/+β : n = 7 mice, db/db Lsd1 fl/+β : n = 8 mice, db/db Lsd1 Δβ : n = 11 mice, db/+ Lsd1 Δβ : n = 14 mice. * p <0.05, ** p <0.01, *** p <0.001 between db/db Lsd1 fl/+β and db/db Lsd1 Δβ mice. (c) Glucose tolerance tests in TM-treated mice of the indicated genotypes. db/+ Lsd1 fl/+β : n = 6 mice, db/db Lsd1 Δβ : n = 11 mice, db/db Lsd1 fl/+β : n = 13 mice. * p <0.05, ** p <0.01 between db/db Lsd1 fl/+β and db/db Lsd1 Δβ mice. (d) Serum insulin and blood glucose (glc) levels in mice of the indicated genotypes following a 6-hour fast or 10 min following intraperitoneal injection of glucose. db/+ Lsd1 fl/+β fast and 10’ glucose: n = 4 mice, db/db Lsd1 Δβ 10’ glucose: n = 5 mice, all other groups: n = 6 mice. (e and f) Static insulin secretion assays for islets from the indicated genotypes of mice stimulated with the indicated glucose (glc) concentrations (in mM) with or without 10 nM of exendin-4 (Ex4) or GIP at 7 wks (e) and 9 wks (f) of age. db/+ Lsd1 fl/+β 16.8 mM glc + Ex4 or GIP wk 7: n = 4 pools of 10 islets each, db/+ Lsd1 Δβ 16.8 mM glc + Ex4 or GIP wk 7 and db/db Lsd1 fl/+β 16.8 mM glc + Ex4 wk 7: n = 5 islet pools, db/+ Lsd1 fl/+β 16.8 mM glc wk 7: n = 10 islet pools, db/+ Lsd1 fl/+β 2.8 mM glc wk 9 and db/db Lsd1 fl/+β 16.8 mM glc wk 9: n = 11 islet pools, db/db Lsd1 fl/+β 2.8 mM glc wk 9, db/db Lsd1 Δβ 2.8 mM glc or 16.8 mM glc wk 9, db/+ Lsd1 fl/+β 16.8 mM glc wk 9, and all genotypes 16.8 mM glc + Ex4 wk 9: n = 12 islet pools, all other groups: n = 6 islet pools. (g and h) Islet insulin content for islets from the indicated genotypes of mice. db/+ Lsd1 Δβ wk 7: n = 12 pools of 10 islets each, db/+ Lsd1 fl/+β wk 7: n = 17 islet pools, db/+ Lsd1 Δβ wk 9: n = 23 islet pools, all other groups: n = 24 islet pools. (i) Schematic of S961 administration via transplanted minipumps (20 nmol/week). Veh, vehicle. (j) Time course of ad libitum-fed blood glucose levels in TM-treated Lsd1 fl/+ ; Pdx1-CreER mice ( Lsd1 fl/+ β ) and TM-treated Lsd1 fl/fl ; Pdx1-CreER mice ( Lsd1 Δβ ) administered S961 or vehicle. Lsd1 fl/+ β veh: n = 3, Lsd1 Δβ veh: n = 5, Lsd1 fl/+ β S961: n = 7, Lsd1 Δβ S961, n = 9. NS, not significant between S961-treated Lsd1 fl/+ β and Lsd1 Δβ mice. * p <0.05 between Lsd1 fl/+ β and Lsd1 Δβ mice (k and l) Blood glucose levels (k) and serum insulin levels (l) after a 6-hour fast in Lsd1 fl/+ β and Lsd1 Δβ treated with S961 or vehicle for the indicated weeks. Lsd1 fl/+ β veh: n = 3, Lsd1 Δβ veh: n = 5, Lsd1 fl/+ β S961: n = 7, Lsd1 Δβ S961, n = 9. Significance was determined by one-way ANOVA followed by Student’s t-test with Welch’s correction for unequal variance as necessary followed by Dunnett’s multiple comparisons test (g and h) or by two-way ANOVA for treatment or genotype interaction with time or stimulation condition followed by Sidak’s (b, c, j) or Benjamini, Krieger and Yekutieli multiple comparisons test (d - f, k, l). * p <0.05, ** p <0.01, *** p <0.001; NS, not significant.

Article Snippet: The following strains were used in this study: Lsd1 flox , Pdx1-creER , and db/db (Jackson labs strain 000642).

Techniques: Injection

Journal: Journal of Advanced Research

Article Title: Epigenetically silenced KAT2B suppresses de novo lipogenesis through destroying HDAC5/LSD1 complex assembly in renal cell carcinoma

doi: 10.1016/j.jare.2025.08.007

Figure Lengend Snippet: KAT2B destroyed HDAC5/LSD1 complex assembly and suppressed FASN transcriptional activity Co-IP assays were performed to verify the interaction strength between wild-type HDAC5 or the K726R mutant and Exportin1 with KAT2B overexpression. (B) Representative immunofluorescence images of wild-type HDAC5, K726R mutant HDAC5, and NES-deleted HDAC5 with KAT2B overexpression in RCC cells. (C-D) Western blots were used to assess HDAC5 and LSD1 expression in RCC cells with KAT2B overexpression or knockdown. (E) The interaction between HDAC5 and LSD1 was determined by Co-IP assays in RCC cells. (F) The interactions between HDAC5 (wild, K726Q, and K726R) and LSD1 were determined by Co-IP assays in 293 T cells. (G) RCC cells were treated with Eltanexor (60 nM) to inhibit Exportin1 activity. The levels of nuclear HDAC5, total HDAC5, Exportin1, and LSD1 were detected using Western blot. (H) Protein stability experiment of LSD1 in RCC cells with KAT2B overexpression after treated with 100 μM cycloheximide (CHX) for 0 h, 1 h, 2 h, 3 h, and 4 h and statistical diagram. (I) Following the addition of chloroquine (10 μM) or MG132 (8 μM) to RCC, LSD1 protein expression was assessed. (J) RCC cells with KAT2B overexpression were immunoprecipitated with LSD1 antibody, and the level of ubiquitin was detected. (K) LSD1 and FASN expression were detected in RCC cells with KAT2B (wild or dead) and/or HDAC5 (wild, 726Q or 726R) overexpression. (L) Schematic diagram illustrating KAT2B-mediated acetylation of HDAC5, promoting its cytoplasmic mislocalization, which resulted in the disruption of the HDAC5-LSD1 complex in the nucleus and subsequent LSD1 degradation.

Article Snippet: KAT2B overexpression and HDAC5 overexpression lentivirus, KAT2B‐targeted shRNA lentivirus, and overexpression plasmids of LSD1 and KAT2B (wild and enzyme-inactivated) was provided by Genechem Co. Ltd (China).

Techniques: Activity Assay, Co-Immunoprecipitation Assay, Mutagenesis, Over Expression, Immunofluorescence, Western Blot, Expressing, Knockdown, Immunoprecipitation, Ubiquitin Proteomics, Disruption

Journal: Journal of Advanced Research

Article Title: Epigenetically silenced KAT2B suppresses de novo lipogenesis through destroying HDAC5/LSD1 complex assembly in renal cell carcinoma

doi: 10.1016/j.jare.2025.08.007

Figure Lengend Snippet: The KAT2B/HDAC5/LSD1/FASN axis repressed RCC lipogenesis and progression in vivo B) The picture of xenografts using Caki-1 cells with KAT2B and/or HDAC5 stable overexpressing. The tumor weight was used for statistical comparison (n = 5). (C) The tumor volume of each group was measured every six days (n = 5). (D) Representative of immunohistochemical (IHC) staining for KAT2B, HDAC5, LSD1, FASN and Ki67 in tumor xenografts. (E) Oil red O staining of the tumor xenografts with KAT2B and/or HDAC5 overexpression. (F) Living fluorescence images of mice in the metastasis model. (G-H) The liver photo and H&E staining of liver tissue in the metastatic model. Data were analyzed by one-way ANOVA (B,C).

Article Snippet: KAT2B overexpression and HDAC5 overexpression lentivirus, KAT2B‐targeted shRNA lentivirus, and overexpression plasmids of LSD1 and KAT2B (wild and enzyme-inactivated) was provided by Genechem Co. Ltd (China).

Techniques: In Vivo, Comparison, Immunohistochemical staining, Immunohistochemistry, Staining, Over Expression, Fluorescence

Journal: Journal of Advanced Research

Article Title: Epigenetically silenced KAT2B suppresses de novo lipogenesis through destroying HDAC5/LSD1 complex assembly in renal cell carcinoma

doi: 10.1016/j.jare.2025.08.007

Figure Lengend Snippet: Graphic abstract of this research TET1-mediated promoter hypermethylation in RCC leaded to decreased KAT2B expression. Mechanistically, KAT2B acetylated HDAC5 at the K726 site and promoted its nucleus export, thereby failing to form a complex with LSD1 in nucleus. This leaded to increased histone methylation levels and decreased FASN expression, ultimately inhibiting lipogenesis and RCC progression. FASN inhibition might be useful in treating KAT2B-low RCC progression by targeting de novo lipogenesis.

Article Snippet: KAT2B overexpression and HDAC5 overexpression lentivirus, KAT2B‐targeted shRNA lentivirus, and overexpression plasmids of LSD1 and KAT2B (wild and enzyme-inactivated) was provided by Genechem Co. Ltd (China).

Techniques: Expressing, Methylation, Inhibition