Journal: Cell reports

Article Title: Lysosome-related organelle integrity suppresses TIR-1 aggregation to restrain toxic propagation of p38 innate immunity

doi: 10.1016/j.celrep.2024.114674

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: For NativePAGE analysis, protein lysates were prepared in 1x NativePAGE buffer (Invitrogen), 1% digitonin, and HALT protease inhibitor (Thermo Fisher).

Techniques: Virus, Recombinant, Protease Inhibitor, cDNA Synthesis, DC Protein Assay, SYBR Green Assay, Software

Journal: PLoS Pathogens

Article Title: Epstein-Barr virus microRNAs regulate B cell receptor signal transduction and lytic reactivation

doi: 10.1371/journal.ppat.1007535

Figure Lengend Snippet: A. BJAB-NFkB-GL4.32, stably transfected with a NF-kappaB-responsive firefly luciferase reporter, were transduced with pLCE empty vector or individual EBV miRNA vectors. Cells were plated in 96-well black-well plates, treated with 10 ug/mL anti-IgM for 18 hrs, and analyzed for luciferase activity. Values are normalized to mock treated cells and reported relative to pLCE control. Shown are the averages of three independent experiments performed in quadruplicate. B. BJAB-NFkBLuc cells, expressing a NF-kappaB-responsive firefly luciferase reporter and a renilla luciferase reporter for internal control, were transduced with pLCE empty vector or individual EBV miRNA vectors. Cells were treated with 5 ug/mL anti-IgM for 18 hr, and then lysed in 1X passive lysis buffer. Luciferase activity was measured using the dual luciferase reporter kit. Values are reported relative to mock-treated control (pLCE) cells. Shown are the averages of four independent experiments performed in triplicate. C. BJAB-NFkBLuc cells transduced with pLCE or pLCE-BHRF1-2 were stimulated with 100 ng/mL LPS for 6 hrs, then analyzed for luciferase activity as in (A.). Shown are the averages of three independent experiments performed in triplicate. D. and E. BJAB-AP1-GL4.44 cells, expressing an AP1-responsive firefly luciferase reporter, were transduced with pLCE-based EBV miRNA expression vectors, treated with 10 ug/mL anti-IgM for 6 hrs (D.) or 18 hrs (E.), then analyzed for luciferase activity. Values are normalized to mock treated cells. Shown are the averages of three independent experiments performed in quadruplicate. F. EBV miRNA expression. RNA was harvested from BJAB cells transduced with control vector (pLCE) or individual EBV miRNA expression vectors (pLCE-miR) and Mutu I cells treated with 5 ug/mL anti-IgM for 48 hrs. miRNAs were detected by qRT-PCR. Values are normalized to cellular miR-16 and reported relative to levels in Mutu I cells. By Student’s t-test, *p<0.05 or **p = 0.054. RLU = relative light units.

Article Snippet: For NF-kappaB activation, 1 x 10^5 cells per well were plated in 96-well black-well plates, stimulated for 18 hrs with 5 or 10 ug/ml anti-IgM (Sigma) as indicated, and then lysed in 1X passive lysis buffer (Promega).

Techniques: Stable Transfection, Transfection, Luciferase, Transduction, Plasmid Preparation, Activity Assay, Control, Expressing, Lysis, Quantitative RT-PCR