Journal: Oncogene

Article Title: PMM2 interacts with TRIM28 to recruit E2F4 and promote KIFC3-mediated tumor glycolysis and colorectal cancer progression

doi: 10.1038/s41388-026-03707-x

Figure Lengend Snippet: Co-immunoprecipitation (Co-IP) assays in HCT-116 and RKO cells showing interaction between endogenous PMM2 and TRIM28 ( A : IP with anti-PMM2; B : IP with anti-TRIM28). C Co-IP of HA-tagged TRIM28 with FLAG-tagged full-length (FL) or domain-deleted PMM2 (Δ1, Δ2, Δ3) in HEK293T cells. Glucose uptake ( D ), Lactate production ( E ), and ECAR ( F ) in HCT-116 shCtrl or shPMM2 cells transfected with NC, PMM2-FL or PMM2-Δ1. G Western blot analysis of PKM2, LDHA, and PMM2 in HCT-116 cells grouped as in ( D ). CCK-8 ( H ) and wound healing ( I ) assays in HCT-116 cells grouped as in ( D ). Data are mean ± SD ( n = 3). ** p < 0.01 vs. corresponding control.

Article Snippet: Anti-PMM2 (10666-1-AP), anti-E2F4 (10923-1-AP), anti-GAPDH (60004-1-lg), anti-β-actin (66009-1-lg), anti-LDHA (19987-1-AP), anti-PKM2 (15822-1-AP), anti-TRIM28 (15202-1-AP), anti-Histone H3 (17168-1-AP), anti-Flag (66008-3-lg) and anti-HA (51064-2-AP) were purchased from Proteintech; Goat anti-mouse (A0216) and goat anti-rabbit (A0208) were obtained from Beyotime.

Techniques: Immunoprecipitation, Co-Immunoprecipitation Assay, Transfection, Western Blot, CCK-8 Assay, Control

Journal: Oncogene

Article Title: PMM2 interacts with TRIM28 to recruit E2F4 and promote KIFC3-mediated tumor glycolysis and colorectal cancer progression

doi: 10.1038/s41388-026-03707-x

Figure Lengend Snippet: A Immunoprecipitation combined with mass spectrometry (IP-MS) analysis of TRIM28-interacting proteins in HCT-116 cells. Co-immunoprecipitation (Co-IP) assays in HCT-116 cells validating the interaction between endogenous TRIM28 and E2F4 ( B : IP with anti-TRIM28; C : IP with anti-E2F4). RT-qPCR ( D ) and Western blot ( E ) analysis of KIFC3 mRNA and protein levels in HCT-116 cells with E2F4 overexpression. F Dual-luciferase reporter assay in 293 T cells transfected with WT or Mut1 KIFC3 promoter constructs, with or without E2F4 overexpression. G ChIP assay showing E2F4 enrichment at the KIFC3 promoter site 1 in HCT-116 cells. H Dual-luciferase reporter assay in 293 T cells co-transfected with PMM2, E2F4, and KIFC3 promoter constructs. I ChIP assay showing PMM2 overexpression enhances E2F4 binding to the KIFC3 promoter site 1 in HCT-116 cells. RT-qPCR ( J ) and Western blot ( K ) analysis of KIFC3 expression in HCT-116 cells with PMM2 overexpression and/or TRIM28 knockdown. L Dual-luciferase reporter assay in 293T cells co-transfected with PMM2, TRIM28 shRNA, E2F4 shRNA, and KIFC3 promoter constructs. M ChIP assay showing TRIM28 knockdown partially inhibits PMM2-induced E2F4 binding to the KIFC3 promoter site 1 in HCT-116 cells. Data are mean ± SD ( n = 3). * p < 0.05, ** p < 0.01 vs. corresponding control. ns, not significant.

Article Snippet: Anti-PMM2 (10666-1-AP), anti-E2F4 (10923-1-AP), anti-GAPDH (60004-1-lg), anti-β-actin (66009-1-lg), anti-LDHA (19987-1-AP), anti-PKM2 (15822-1-AP), anti-TRIM28 (15202-1-AP), anti-Histone H3 (17168-1-AP), anti-Flag (66008-3-lg) and anti-HA (51064-2-AP) were purchased from Proteintech; Goat anti-mouse (A0216) and goat anti-rabbit (A0208) were obtained from Beyotime.

Techniques: Immunoprecipitation, Mass Spectrometry, Protein-Protein interactions, Co-Immunoprecipitation Assay, Quantitative RT-PCR, Western Blot, Over Expression, Luciferase, Reporter Assay, Transfection, Construct, Binding Assay, Expressing, Knockdown, shRNA, Control

Journal: Oncology reports

Article Title: TRIM28 promotes cervical cancer growth through the mTOR signaling pathway.

doi: 10.3892/or.2018.6235

Figure Lengend Snippet: Figure 1. TRIM28 expression is elevated in cervical cancer tissues and cell lines. (A) Analysis from the Oncomine database showing that TRIM28 mRNA expression is significantly elevated in cervical cancer tissues (n=40) compared with that in normal tissues (n=5). The data were obtained from a previous study (35). (B) TRIM28 mRNA levels in 20 paired cervical cancer samples and adjacent normal tissues examined by qRT‑PCR. (C) Immunohistochemical staining of normal and cervical cancer tissues with anti-TRIM28 antibody. The expression levels of TRIM28 in human cervical cancer cells and normal human cervical tissues were measured by qRT‑PCR (D) and western blotting (E). Data are presented as means ± SD. *P<0.05.

Article Snippet: After blocking the endogenous peroxidase with 3% hydrogen peroxide, the sections were washed with Tris-buffered saline (TBS) and then incubated with a primary antibody targeting TRIM28 (Proteintech, Hubei, China) for 1 h at 37 ̊C.

Techniques: Expressing, Immunohistochemical staining, Staining, Western Blot

Journal: Oncology reports

Article Title: TRIM28 promotes cervical cancer growth through the mTOR signaling pathway.

doi: 10.3892/or.2018.6235

Figure Lengend Snippet: Figure 3. TRIM28 enhances cervical cancer cell tumorigenicity. (A) Tumor volumes were measured on the indicated days. (B) At the end of the experiment, the weights were measured. (C) The proliferation index (right) was determined based on the percentage of Ki67-positive cells (left). (D) Western blot analysis of TRIM28 and p-mTOR protein expression in indicated tumor xenografts. Data are presented as means ± SD. *P<0.05.

Article Snippet: After blocking the endogenous peroxidase with 3% hydrogen peroxide, the sections were washed with Tris-buffered saline (TBS) and then incubated with a primary antibody targeting TRIM28 (Proteintech, Hubei, China) for 1 h at 37 ̊C.

Techniques: Western Blot, Expressing

Journal: Oncology reports

Article Title: TRIM28 promotes cervical cancer growth through the mTOR signaling pathway.

doi: 10.3892/or.2018.6235

Figure Lengend Snippet: Figure 2. TRIM28 promotes cervical cancer cell proliferation by accelerating the cell cycle. (A) Western blot analysis of TRIM28 protein expression in TRIM28-infected cells and TRIM28 shRNA-infected cells. GAPDH was used as the loading control. (B) Effects of TRIM28 upregulation or downregulation on the proliferation of SiHa or CaSki cells analyzed by the MTT assay. (C) Representative micrographs (left) and quantification (right) of colonies formed by the indicated cervical cancer cell lines. (D) Cell cycle progression of the indicated cervical cancer cell lines analyzed by flow cytometry. (E) Representative images (left) and quantification (right) of BrdU-positive cells in the indicated cervical cancer cell lines. Data are presented as means ± SD. *P<0.05.

Article Snippet: After blocking the endogenous peroxidase with 3% hydrogen peroxide, the sections were washed with Tris-buffered saline (TBS) and then incubated with a primary antibody targeting TRIM28 (Proteintech, Hubei, China) for 1 h at 37 ̊C.

Techniques: Western Blot, Expressing, Infection, shRNA, Control, MTT Assay, Flow Cytometry

Journal: Oncology reports

Article Title: TRIM28 promotes cervical cancer growth through the mTOR signaling pathway.

doi: 10.3892/or.2018.6235

Figure Lengend Snippet: Figure 4. TRIM28 promotes cervical cancer cell proliferation via the mTOR signaling pathway. (A) Western blot analysis of TRIM28, p-mTOR(S2448), mTOR, p-S6K1(T389), and S6K1 in TRIM28 shRNA-infected CaSki cells and TRIM28-infected SiHa cells in the presence or absence of 20 nM evero- limus (EVE) for 48 h. GAPDH served as the loading control. (B-D) Cell proliferation, cell cycle, and BrdU incorporation were analyzed in TRIM28-infected SiHa cells in the presence or absence of EVE for 48 h. Data are presented as means ± SD. *P<0.05.

Article Snippet: After blocking the endogenous peroxidase with 3% hydrogen peroxide, the sections were washed with Tris-buffered saline (TBS) and then incubated with a primary antibody targeting TRIM28 (Proteintech, Hubei, China) for 1 h at 37 ̊C.

Techniques: Western Blot, shRNA, Infection, Control, BrdU Incorporation Assay

Journal: Scientific Reports

Article Title: Hypomethylation of ERVs in the sperm of mice haploinsufficient for the histone methyltransferase Setdb1 correlates with a paternal effect on phenotype

doi: 10.1038/srep25004

Figure Lengend Snippet: (a ) Pedigree produced from a Setdb1 MD13 /+ sire and pseudoagouti ( A vy ) dam. Wild type (WT) offspring from Setdb1 MD13 /+ sires showed a significant increase (Chi-square test: P < 0.001) in the proportion of animals with yellow coats when compared with WT offspring from WT sires. Offspring heterozygous for Setdb1 MD13 demonstrated a significant shift in penetrance toward yellow when compared to WT littermates (Chi-square test: P < 0.02). ( b ) Pedigree produced from a Trim28 MD9 /+ sire and pseudoagouti ( A vy ) dam. Trim28 MD9 heterozygotes exhibited a significant shift in penetrance toward yellow when compared with WT littermates (Chi-square test: P < 0.05). ( c ) Pedigree produced from a pseudoagouti ( A vy ) sire and Setdb1 MD13 /+ dam. Offspring showed no shift in penetrance at A vy . For all crosses data were produced from at least five different mating pairs. Offspring not carrying the A vy allele have been omitted.

Article Snippet: Overnight incubation in primary antibodies anti-Setdb1 antibody (11231-1-AP, Proteintech, Chicago, IL, USA) at 1:200, anti-Trim28 antibody (MP-2365522, Merck-Millipore) at 1:1500 and anti-histone H3 (trimethyl K9) (ab8898, Abcam) at 1:400 at 4 °C was followed by secondary biotinylated universal antibody (BA-1400, Vector Laboratories) incubation and ABC reagent incubation before development of DAB stain (SK-4100, Peroxidase substrate Kit, Vector Laboratories).

Techniques: Produced

Journal: Scientific Reports

Article Title: Hypomethylation of ERVs in the sperm of mice haploinsufficient for the histone methyltransferase Setdb1 correlates with a paternal effect on phenotype

doi: 10.1038/srep25004

Figure Lengend Snippet: Effect of parental haploinsufficiency of epigenetic modifiers on expression of A vy in offspring.

Article Snippet: Overnight incubation in primary antibodies anti-Setdb1 antibody (11231-1-AP, Proteintech, Chicago, IL, USA) at 1:200, anti-Trim28 antibody (MP-2365522, Merck-Millipore) at 1:1500 and anti-histone H3 (trimethyl K9) (ab8898, Abcam) at 1:400 at 4 °C was followed by secondary biotinylated universal antibody (BA-1400, Vector Laboratories) incubation and ABC reagent incubation before development of DAB stain (SK-4100, Peroxidase substrate Kit, Vector Laboratories).

Techniques: Expressing, Mutagenesis, Derivative Assay

Journal: Scientific Reports

Article Title: Hypomethylation of ERVs in the sperm of mice haploinsufficient for the histone methyltransferase Setdb1 correlates with a paternal effect on phenotype

doi: 10.1038/srep25004

Figure Lengend Snippet: ( a ) Detection of Setdb1, Trim28 and H3K9me3 by immunohistochemistry in testes from 3 months old males. For each, a secondary antibody control (2°Ab) is shown to indicate the specificity of the immunohistochemical staining (first panel of each row), followed by specific staining at 20x and 100x magnification. Setdb1 was found to be present in spermatogonia, Trim28 in spermatocytes and spermatids and H3K9me3 in spermatogonia and spermatids. ( b ) Box plot of the average DNA methylation in 10Kb windows across the genome (n = 246, 281), excluding outliers (1.5× interquartile range). ( c ) Plot showing the weighted average DNA methylation for sequence elements annotated as ERVK in sperm from Setdb1 MD13 /+ and Setdb1 +/+ mice (n = 16,331). Elements for which the DNA methylation difference was greater than 10 percentage points are colored blue (n = 607). ( d ) Counts of ERVK subfamilies that were either hypermethylated or hypomethylated in Setdb1 MD13 /+ relative to Setdb1 +/+ and for which DNA methylation differed by more than 10 percentage points. The 20 most frequently represented subfamilies are shown. ( e ) UCSC genome browser screen capture showing a reduction of mCG at an IAPLTR3 (top left panel) and a RNERVK23-int (top right panel) in Setdb1 MD13 heterozygotes. Sanger sequencing of bisulphite converted sperm genomic DNA from independent males confirmed reduced mCG levels of the IAPLTR3 (bottom left panel; T-test P = 0.05) and the RNERVK23-int (bottom right panel; T-test P < 0.05) in Setdb1 MD13 heterozygotes. Each boxed block of lines comprises clones derived from one bisulphite conversion and each column represents one individual with the percentage of methylated CpGs indicated above each individual. Open circles indicate an unmethylated CpG, and closed circles a methylated CpG.

Article Snippet: Overnight incubation in primary antibodies anti-Setdb1 antibody (11231-1-AP, Proteintech, Chicago, IL, USA) at 1:200, anti-Trim28 antibody (MP-2365522, Merck-Millipore) at 1:1500 and anti-histone H3 (trimethyl K9) (ab8898, Abcam) at 1:400 at 4 °C was followed by secondary biotinylated universal antibody (BA-1400, Vector Laboratories) incubation and ABC reagent incubation before development of DAB stain (SK-4100, Peroxidase substrate Kit, Vector Laboratories).

Techniques: Immunohistochemistry, Control, Immunohistochemical staining, Staining, DNA Methylation Assay, Sequencing, Blocking Assay, Clone Assay, Derivative Assay, Methylation