Journal: bioRxiv

Article Title: EZH2 Serine 21 Phosphorylation Restrains Compact-State PRC2 Activation and H3K27me3 Propagation

doi: 10.64898/2026.06.02.729660

Figure Lengend Snippet: A. Domain schematic of the purified recombinant core PRC2 subunits EZH2, SUZ12, EED, and RBBP4, with annotated phosphorylation sites. Yellow circles denote phosphorylation sites reported in high-throughput proteomic studies, and red circles denote sites reported in low-throughput studies. B. In vitro histone methyltransferase assays comparing untreated recombinant PRC2 and PRC2 dephosphorylated with lambda phosphatase. Sf9-purified PRC2 complexes were incubated with recombinant nucleosomes and a 2-fold serial dilution of stimulatory H3K27me3 peptide, beginning at 40 μM. Reaction products were analyzed by immunoblotting. C. Quantification of PRC2 catalytic activity in response to increasing concentrations of stimulatory H3K27me3 peptide. Untreated or lambda phosphatase-treated recombinant PRC2 was incubated with recombinant nucleosomes, and methyltransferase activity was measured by incorporation of 3 H-SAM. Data are shown as mean ± SEM. D. In vitro methyltransferase activity of recombinant PRC2 complexes containing wild-type EZH2, EZH2 S21D, or EZH2 S21A, assayed before or after lambda phosphatase treatment. Complexes were incubated with recombinant nucleosomes, and activity was quantified by incorporation of 3 H-SAM. Data are shown as mean ± SEM, with individual replicate values indicated. Statistical comparisons are indicated above the bars. E. EZH2 domain schematic and multiple sequence alignment of the conserved N-terminal region surrounding S21. The AKT1 consensus motif and S21 residue are indicated. F. Immunoblot analysis of Ezh2 knockout MEFs reconstituted with FLAG-tagged wild-type EZH2, S21D, S21A, or empty vector. GAPDH serves as a loading control. G. Heatmaps and aggregate profiles showing spike-in-normalized H3K27me3 and EZH2 ChIP-seq signal centered on H3K27me3 or EZH2 peaks in Ezh2 -knockout MEFs expressing the indicated EZH2 transgenes or empty vector. ChIP-seq experiments were normalized using exogenous spike-in chromatin. H. Representative genome browser tracks showing spike-in-normalized H3K27me3 and EZH2 ChIP-seq signal at Polycomb target loci in Ezh2 -knockout MEFs expressing the indicated EZH2 transgenes or empty vector. I. Immunoblot analysis of FLAG coimmunoprecipitation from nuclear extracts of Ezh2 knockout MEFs expressing the indicated FLAG-tagged EZH2 transgenes or empty vector control.

Article Snippet: Sf9 insect cells were obtained from ATCC (CRL-1711; RRID:CVCL 0549) and were used for baculovirus-mediated expression of recombinant PRC2 complexes.

Techniques: Purification, Recombinant, Phospho-proteomics, High Throughput Screening Assay, In Vitro, Incubation, Serial Dilution, Western Blot, Activity Assay, Sequencing, Residue, Knock-Out, Plasmid Preparation, Control, ChIP-sequencing, Expressing

Journal: Journal of Pesticide Science

Article Title: Design, synthesis, and insecticidal activity evaluation of piperine-phenylene diamine derivatives

doi: 10.1584/jpestics.D25-065

Figure Lengend Snippet: Fig. 4. Morphological observation of Sf9 cells. (a) untreated, (b) treated with D15 , and (c) treated with azadirachtin.

Article Snippet: Spodoptera exigua were purchased from Henan Jiyuan Baiyun Industry Co., Ltd. Sf9 cells ( S . frugiperda ovary cells), thiazole blue (MTT) and dimethyl sulfoxide (DMSO) were purchased from Wuhan Procell Life Technology Co., Ltd. One Step TUNEL Apoptosis Assay Kit and universal genomic DNA purification mini spin kit were purchased from Shanghai Beyotime Institute of Biotechnology.

Techniques:

Journal: Journal of Pesticide Science

Article Title: Design, synthesis, and insecticidal activity evaluation of piperine-phenylene diamine derivatives

doi: 10.1584/jpestics.D25-065

Figure Lengend Snippet: Fig. 5. Morphological observation of Sf9 cells. (a) untreated, (b) treated with J1 , and (c) treated with azadirachtin.

Article Snippet: Spodoptera exigua were purchased from Henan Jiyuan Baiyun Industry Co., Ltd. Sf9 cells ( S . frugiperda ovary cells), thiazole blue (MTT) and dimethyl sulfoxide (DMSO) were purchased from Wuhan Procell Life Technology Co., Ltd. One Step TUNEL Apoptosis Assay Kit and universal genomic DNA purification mini spin kit were purchased from Shanghai Beyotime Institute of Biotechnology.

Techniques:

Journal: Journal of Pesticide Science

Article Title: Design, synthesis, and insecticidal activity evaluation of piperine-phenylene diamine derivatives

doi: 10.1584/jpestics.D25-065

Figure Lengend Snippet: Fig. 7. Annexin V-FITC/PI staining of Sf9 cells. (a) untreated, (b) treated with J1 , and (c) treated with azadirachtin.

Article Snippet: Spodoptera exigua were purchased from Henan Jiyuan Baiyun Industry Co., Ltd. Sf9 cells ( S . frugiperda ovary cells), thiazole blue (MTT) and dimethyl sulfoxide (DMSO) were purchased from Wuhan Procell Life Technology Co., Ltd. One Step TUNEL Apoptosis Assay Kit and universal genomic DNA purification mini spin kit were purchased from Shanghai Beyotime Institute of Biotechnology.

Techniques: Staining

Journal: Journal of Pesticide Science

Article Title: Design, synthesis, and insecticidal activity evaluation of piperine-phenylene diamine derivatives

doi: 10.1584/jpestics.D25-065

Figure Lengend Snippet: Fig. 8. Flow cytometry analysis of Sf9 cells. (a) untreated, (b) treated with D15 , and (c) treated with J1 .

Article Snippet: Spodoptera exigua were purchased from Henan Jiyuan Baiyun Industry Co., Ltd. Sf9 cells ( S . frugiperda ovary cells), thiazole blue (MTT) and dimethyl sulfoxide (DMSO) were purchased from Wuhan Procell Life Technology Co., Ltd. One Step TUNEL Apoptosis Assay Kit and universal genomic DNA purification mini spin kit were purchased from Shanghai Beyotime Institute of Biotechnology.

Techniques: Flow Cytometry