Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: MSL1 Promotes Liver Regeneration by Driving Phase Separation of STAT3 and Histone H4 and Enhancing Their Acetylation.

doi: 10.1002/advs.202301094

Figure Lengend Snippet: Figure 2. MSL1 enhances STAT3 transcriptional activity by interacting with STAT3 during liver regeneration. a) Immunoblot analysis of liver tissue lysates from WT and LKO mice at 24–72 h after partial hepatectomy (PH) using the indicated antibodies. b) Immunoblot analysis of primary mouse hepatocytes from WT and LKO mice treated with 50 × 10−6 m MG149 for 24 h with or without 30-min 10 ng mL−1 IL-6 treatment using the indicated antibodies. c,d) qRT-PCR analysis of c-Myc (c) and SOCS3 (d) mRNA expression in the regenerating liver (n = 4–5 mice per group). e) ELISA analysis of serum IL-6 level in WT and LKO mice at 24–72 h after PH (n = 3–4 mice per group). f) Coimmunoprecipitation analysis of MSL1 and STAT3 in liver tissues from WT mice. g) HEK293T cells were transfected with Myc-MSL1-encoding plasmids and with or without Flag-STAT3 plasmids, followed by culturing for 24 h without or with 10 ng mL−1 IL-6 for 30 min. Coimmunoprecipitation analysis was performed. h) Hep1-6 cells were transfected with plasmids for MSL1 or control vector for 24 h and treated without or with 10 ng mL−1 IL-6 for 30 min. Immunoblot analysis using the indicated antibodies. The data were expressed as means ± SD. Significant difference was presented at the level of *p < 0.05 by two-tailed Student’s t-test.

Article Snippet: HEK 293T cells, Hep1-6 cells, and primary mouse hepatocytes respectively treated with human IL-6 (MCE, HY-P7044) for 30 min for immunoblot analysis.

Techniques: Activity Assay, Western Blot, Quantitative RT-PCR, Expressing, Enzyme-linked Immunosorbent Assay, Transfection, Control, Plasmid Preparation, Two Tailed Test

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: MSL1 Promotes Liver Regeneration by Driving Phase Separation of STAT3 and Histone H4 and Enhancing Their Acetylation.

doi: 10.1002/advs.202301094

Figure Lengend Snippet: Figure 4. MSL1 phase separation modulated by acetyl-CoA in cells and in vitro. a) Primary mouse hepatocytes immunostained with an MSL1 antibody in WT and LKO mice. Scale bar, 5 μm. b) Fluorescence recovery after photo bleach (FRAP) time-lapse images and quantification of GFP-MSL1 condensates in live transfected HEK293T cells. Data are representative of three independent FRAP events. Scale bar, 5 μm. c) Confocal images and quantification of GFP-MSL1 condensates at increasing protein concentrations (2 × 10−6–16 × 10−6 m). n = 3 fields (100 × 100 μm2). Scale bar, 5 μm. d) FRAP assays and quantification of GFP-MSL1 (8 × 10−6 m) condensates. Data are representative of three independent FRAP events. Scale bar, 1 μm. e) Immunofluorescence images of MSL1 in primary hepatocytes treated with CMS-121 (1 × 10−6 m) or SB-204990 (10 × 10−6 m) for 24 h. Scale bar, 5 μm. f) Quantification of MSL1 nuclear puncta in primary hepatocytes (n = 20 cells). g) Confocal images of GFP-MSL1 condensates (10 × 10−6 m) at increasing acetyl-coenzyme A (Ac-CoA) concentrations (0–500 × 10−6 m). n = 3 fields (100 × 100 μm2). Scale bar, 5 μm. h) SDS–PAGE assay of MSL1 recovered from the aqueous phase or supernatant (S) and the condensed phase or pellet (P). Ac-CoA, PEG-8000, or 1.6-hex were added at the indicated concentrations. i) Quantification of the protein fraction recovered from pellets in the sedimentation assays. The sedimentation experiments were conducted in triplicates. j) Quantification of the Ac-CoA recovered from pellets in the sedimentation assays, Ac-CoA, PEG-8000, and 1.6-hex were added at the indicated concentrations. The sedimentation experiments were conducted in triplicates. The data were expressed as means ± SD. Significant difference was presented at the level of *p < 0.05, **p <0.01, and ***p <0.001 by two-tailed Student’s t-test.

Article Snippet: HEK 293T cells, Hep1-6 cells, and primary mouse hepatocytes respectively treated with human IL-6 (MCE, HY-P7044) for 30 min for immunoblot analysis.

Techniques: In Vitro, Fluorescence, Transfection, SDS Page, Sedimentation, Two Tailed Test

Journal: Nature Microbiology

Article Title: Evolution of enhanced innate immune suppression by SARS-CoV-2 Omicron subvariants

doi: 10.1038/s41564-023-01588-4

Figure Lengend Snippet: a – g , Calu-3 infection with 2,000 E copies per cell of Delta (yellow, Ο), BA.1 (blue, Ο), BA.2 (blue, Δ), BA.4 (purple, O) and BA.5 (purple, Δ), n = 3: mean viral E copies at 2 h.p.i. across three independent experiments ( a ); viral replication over time measured by RT–qPCR for intracellular E copies per microgram RNA ( b ); infection levels measured by nucleocapsid expression (% N+ by flow cytometry) ( c ); expression of IFNB , CXCL10 , IFIT1 , IFIT2 , RSAD2 , MX1 , MX2 and DDX58 in infected cells over time ( d ); IFNβ ( e ) and CXCL10 ( f ) secretion from infected Calu-3 cells measured by ELISA at 48 h.p.i.; rescue of viral replication by JAK1-inhibitor ruxolitinib in Calu-3 cells at 48 h.p.i., where relative infection levels are shown across three independent experiments determined by E copies per microgram RNA normalized to the median infection level of the untreated control ( g ). h – k , Primary bronchial HAEs were infected with the indicated variants at 1,500 E copies per cell: viral replication measured by intracellular E copies at 72 h.p.i. ( h ) and viral release into apical washes over time ( i ), with three biological replicates shown; expression of IFNB , CXCL10 , IFIT1 , IFIT2 , DDX58 and RSAD2 in HAEs at 72 h.p.i., with six biological replicates shown ( j ); intracellular viral E copies in HAEs in the presence or absence of 5 μM ruxolitinib at 72 h.p.i., with three biological replicates shown ( k ). For a , one-way analysis of variance (ANOVA) with Bonferroni post-test was used. n.s., not significant at P > 0.05 for all comparisons. For b – h and j , one-way ANOVA and Dunnett’s post-test were used. For i , two-way ANOVA with a Bonferroni post-test was used. For k , one-tailed unpaired Student’s t -test was used. Replicate measurements from one of three independent experiments. Fold change over mock is shown. Mean ± s.e.m. or individual datapoints are shown. h.p.i., hours post infection.

Article Snippet: IFNβ, IFNλ1/IFNλ3 and CXCL10 were measured using Human IFN-β Quantikine ELISA Kit, Human IL-29/IL-28B (IFNλ1/IFNλ3) DuoSet ELISA or Human CXCL10/IP-10 DuoSet ELISA reagents (Bio-Techne R&D Systems) according to the manufacturer’s instructions.

Techniques: Infection, Quantitative RT-PCR, Expressing, Flow Cytometry, Enzyme-linked Immunosorbent Assay, One-tailed Test