rabbit monoclonal anti phospho s6 ribosomal protein  (Cell Signaling Technology Inc)

Journal: Tumour Virus Research

Article Title: Oncolytic virus hijacks GOT1 and pyrimidinosomes to fuel pyrimidine synthesis for replication in tumor cells

doi: 10.1016/j.tvr.2026.200342

Figure Lengend Snippet: | The MAS-transported NAD + /NADH regulates CAD phosphorylation activity and NDV replication through S6K. (A) Schematic diagram showing the potential involvement of mTOR-S6K in regulating CAD phosphorylation through MAS-mediated NAD + /NADH translocation. (B-D) A549 cells were either mock-infected or infected with NDV (MOI = 1) at 6, 12, 18 and 24 hpi. Protein levels of p-S6K (T389), S6K, p-S6 (S235/236), S6, p-CAD (S1859), CAD, NP and β-actin were analyzed by WB at 12 hpi (B). Grayscale quantification of p-S6K/S6K, p-S6/S6 and p-CAD/CAD ratios from WB data (C). Intracellular NAD + /NADH ratio were measured (D). (E-H) A549 cells were infected with NDV (MOI = 1), treated with AOA, supplemented with Pyr or Asp. Protein levels of p-S6K, S6K, p-S6, S6, p-CAD, CAD, NP, and β-actin were analyzed by WB at 12 hpi (E). Grayscale quantification of p-S6K/S6K, p-S6/S6 and p-CAD/CAD ratios from WB data (F). Intracellular NAD + /NADH ratio were measured (G). Extracellular virus yields were detected (H). (I-L) A549 cells were infected with NDV (MOI = 1), treated with AOA, supplemented with Pyr, and exposed to Torin 1. Protein levels of p-S6K, S6K, p-S6, S6, p-CAD, CAD, NP, and β-actin were analyzed by WB at 12 hpi (I). Grayscale quantification of p-S6K/S6K, p-S6/S6 and p-CAD/CAD ratios from WB data (J). Intracellular NAD + /NADH ratio were measured (K). Extracellular virus yields were detected (L). (M − P) A549 cells were infected with NDV (MOI = 1), treated with AOA, and supplemented with Duro. Protein levels of p-S6K, S6K, p-S6, S6, p-CAD, CAD, NP, and β-actin were analyzed by WB at 12 hpi (M). Grayscale quantification of p-S6K/S6K, p-S6/S6 and p-CAD/CAD ratios from WB data (N). Intracellular NAD + /NADH ratio were measured (O). Extracellular virus yields were detected (P). Data are presented as means from three independent experiments, ns, p > 0.05, ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001.

Article Snippet: The antibodies used for immunoblotting in this study included monoclonal or polyclonal: anti-β-actin (Proteintech, Cat# 66009-1-Ig), anti-NP (Prepared in our laboratory), anti-S6 ribosomal protein (Cell Signaling Technology [CST], Cat# 2217), anti-phospho-S6 ribosomal protein (S235/236) (CST, Cat# 4858), anti-phospho-p70 S6 Kinase 1 (T389) (ABclonal, Cat# AP0564), anti-p70 S6 Kinase 1 (ABclonal, Cat# A2190), anti-CAD (CST, Cat# 11933), anti-phospho-CAD (S1859) (CST, Cat# 70307), anti-GOT1 (Proteintech, Cat# 14886-1-AP), anti-GOT2 (Proteintech, Cat# 14800-1-AP). anti-UMPS (Proteintech, Cat# 14830-1-AP), anti-DHODH (Proteintech, Cat# 14877-1-AP) and anti-Tom20 (Abcam, Cat# ab283317).

Techniques: Phospho-proteomics, Activity Assay, Translocation Assay, Infection, Virus

Journal: JID Innovations

Article Title: The 4E-BP1 deletion modestly mitigates mTORC1-deficient epidermal barrier defects

doi: 10.1016/j.xjidi.2026.100453

Figure Lengend Snippet: The 4E-BP1 depletion slightly rescues the skin barrier function in Rap EKO mice. ( a ) Schematic representation of the floxed Rptor locus and complete Eif4ebp1 knockout, showing PCR fragment lengths before and after recombination. Genomic DNA PCR analysis from mouse tails confirms successful recombination of the floxed Rptor region in the presence of K14-driven Cre and complete Eif4ebp1 knockout. ( b ) Western blot analysis of 4E-BP1 protein expression in back skin from indicated genotypes. ( c ) Macroscopic appearance and body weight measurements of control, Rap EKO , and dKO newborn mice at P0. ( d ) Representative toluidine blue dye penetration assay with newborn mice and the quantification of the blue stained area (n = 5). ( e ) Representative H&E-stained sections of back skin and tongue from control, Rap EKO , and dKO newborns at P0. Dashed lines indicate the basement membrane. Bar = 100 μm. Right panels show quantification of epidermal thickness and HF density (n = 5). ( f ) Left: immunohistochemical staining for p-S6 and p-4E-BP1 in back skin sections at P0. Bar = 50 μm. Right: quantitative analysis of p-S6 and p-4E-BP1 immunoreactivity (n = 5). Dashed lines indicate the basement membrane. Data are presented as mean ± SEM; each dot represents an individual mouse. Statistical significance was determined by 2-way ANOVA with multiple comparisons. ∗ P < .05, ∗∗ P < .01, ∗∗∗ P < .001, and ∗∗∗∗ P < .001. dKO denotes Rap EKO / Eif4ebp1 −/− , e denotes epidermis, and d denotes dermis. dKO, double-knockout; HF, hair follicle; K14, keratin 14; ns, not significant; p-4E-BP1, phosphorylated 4E-BP1; P0, postnatal day 0; P2, P0, postnatal day 2; p-S6, phosphorylated S6; WT, wild-type.

Article Snippet: Primary antibodies and dilutions included Ki-67 (1:200, Abcam, ab15580), K14 (1:500, Abcam, ab181595), loricrin (1:200, Invitrogen, PA5-30583), keratin 10 (1:200, Santa Cruz, SC-23877), FLG (1:250, BioLegend, #905804), phosphorylated S6 (1:200, Cell Signaling Technology, #5364s), and phosphorylated 4EBP1 (1:200, Cell Signaling Technology, #2855).

Techniques: Knock-Out, Western Blot, Expressing, Control, Staining, Membrane, Immunohistochemical staining, Double Knockout