pharmacological agents  (MedChemExpress)

Journal: Journal of Virology

Article Title: Galectin 3-binding protein suppresses PRRSV replication via Cullin3-mediated ubiquitination degradation of non-structural protein 12

doi: 10.1128/jvi.01083-25

Figure Lengend Snippet: LGALS3BP specifically degrades the PRRSV nsp12. ( A ) HEK-293T cells were transfected with PRRSV nsps and LGALS3BP for 24 h. The cell lysates were collected to analyze PRRSV nsps protein expression. ( B ) HEK-293T cells were transfected with different doses of LGALS3BP (0.1, 0.2, 0.4, 0.8, and 1.6 µg) and nsp12 for 24 h. The cell lysates were collected to analyze nsp12 protein expression. ( C ) HEK-293T cells were transfected with LGALS3BP and nsp12 for 12, 24, 36, and 48 h. The cell lysates were collected to analyze nsp12 protein expression. ( D ) HEK-293T cells were transfected with nsp12 and LGALS3BP for 12 h, and then treated with MG132 (0.1 µM), or 3-methyladenine (3-MA) (1 mM) for 24 h. ( E ) HEK-293T cells were transfected with nsp12 and LGALS3BP for 12 h and treated with or without MG132 (0.1 µM) for 24 h. Cells were collected to analyze by Co-IP assay. ( F ) HEK-293T cells were transfected with LGALS3BP and nsp12 for 24 h. Cells were collected to analyze by Co-IP assay. ( G ) HEK-293T cells were transfected with nsp12 and LGALS3BP for 24 h, and double stained with a rabbit anti-HA antibody and a mouse anti-Flag antibody, followed by fluorescein Alexa Fluor-conjugated anti-rabbit IgG (green) and Alexa Fluor-conjugated anti-mouse IgG (red). Cell nuclei were counterstained with 1 µg/mL of 4’, 6’-diamidino-2-phenylindole.

Article Snippet: Pharmacological agents MG132 (HY-13259) and 3-methyladenine (3-MA) (HY-19312) were purchased from MedChemExpress (MCE).

Techniques: Transfection, Expressing, Co-Immunoprecipitation Assay, Staining

Journal: Journal of Virology

Article Title: Galectin 3-binding protein suppresses PRRSV replication via Cullin3-mediated ubiquitination degradation of non-structural protein 12

doi: 10.1128/jvi.01083-25

Figure Lengend Snippet: LGALS3BP degrades nsp12 by recruiting the Cullin3 E3 ubiquitin ligase. ( A ) HEK-293T cells were transfected with Cullin3-specific siRNAs for 24 h, cells were collected to analyze the Cullin3 mRNA levels by RT-qPCR. ( B ) HEK-293T cells were transfected with Cullin3-specific siRNA for 24 hours, followed by co-transfection with LGALS3BP and nsp12 plasmids for another 24 hours. The cell lysates were collected to analyze nsp12 protein expression. ( C and D ) HEK-293T cells were transfected with Cullin3 and LGALS3BP ( C ) /nsp12 ( D ) plasmids for 36 h. Cells were collected to analyze by Co-IP assay. ( E ) HEK-293T cells were transfected for 12 h, then treated with MG132 (0.1 µM) or DMSO for another 24 h. Cells were collected to analyze by Co-IP assay. ( F and G ) HEK-293T cells were transfected and stained with mouse anti-HA ( F ) antibody or mouse anti-Flag ( G ) antibody, followed by Alexa Fluor-conjugated anti-mouse IgG (red). Cell nuclei were counterstained with 1 µg/mL of 4′,6′-diamidino-2-phenylindole. Statistical significance was determined by one-way ANOVA (****, P < 0.0001).

Article Snippet: Pharmacological agents MG132 (HY-13259) and 3-methyladenine (3-MA) (HY-19312) were purchased from MedChemExpress (MCE).

Techniques: Ubiquitin Proteomics, Transfection, Quantitative RT-PCR, Cotransfection, Expressing, Co-Immunoprecipitation Assay, Staining

Journal: bioRxiv

Article Title: Unbiased genetics identifies a glutamatergic signaling network as a mediator of daily sleep patterns

doi: 10.1101/2025.05.08.652901

Figure Lengend Snippet: A . Significant genome-wide QTLs, indicated by logarithm of odds (LOD) score, associated with sleep patterns in F2 mice were detected for sleep and circadian metrics, examples shown for %sleep light/dark ratio and cosinor amplitude. 95%, 90%, and 80% significance levels indicated: red, blue, green lines respectively. B . Spearman correlation between sleep and circadian metrics. The sizes of circles show the absolute value of corresponding correlation coefficients. Color indicates the value of correlation coefficients, positive in blue, negative in red. C . Map of chromosome 7 region containing candidate genes GRM5 and GCPII. D . qPCR measurement of GCPII, GRM5, and GRM3 mRNA isolated from cortex (CTX), hippocampus (Hip), and hypothalamus (Hyp) from CC036 and CC057 mice, values normalized to CC036. **P<0.01, ***P<0.001, ns: not significant, Unpaired Student’s t-test. N=3-6. E-F . Western blot of GRM5 and GRM3 from whole cortical lysate (WCL) or synapse enriched P2 fractions from CC036 and CC057 mice. Quantification of western blots shows trend in WCL and significantly increased expression in P2 of GRM5 and GRM3 in CC057 compared to CC036. *P<0.05, **P<0.01, ns: not significant, Unpaired Student’s t-test. N=3. G-I . Sleep recordings and pharmacology validation of QTL hits. CC036 and CC057 mice were injected with vehicle followed 72hrs later by drug, total sleep amount (%) was compared between vehicle and drug treatment days. Time of injection indicated by arrow. Mice injected at light onset with GRM5 activator LSN2814617 (LSN) ( G ), or GRM2/3 inhibitor LY341495 (LY) ( H ), or dark onset with GCPII inhibitor 2-PMPA (PMPA) ( I ). *P<0.05, **P<0.01, ***P<0.001 Paired Student’s t-test. N=5-6 per condition. J . Model depicting coordinated signaling of GRM5, GCPII and GRM3 across the tripartite synapse composed of pre- and post-synaptic compartments, and astrocyte peripheral processes.

Article Snippet: Pharmacology agents were purchased from Cayman Chemical (Anne Arbor MI, USA).

Techniques: Genome Wide, Isolation, Western Blot, Expressing, Biomarker Discovery, Injection