Journal: Access Microbiology

Article Title: Identification of NpdA as the protein forming the surface layer in Paracidovorax citrulli and evidence of its occurrence as a surface layer protein in diverse genera of the Betaproteobacteria and Gammaproteobacteria

doi: 10.1099/acmi.0.000685.v3

Figure Lengend Snippet: Cultures examined in the S-layer studies

Article Snippet: Paracidovorax anthurii , DSM-16745 , DSMZ.

Techniques:

Journal: Access Microbiology

Article Title: Identification of NpdA as the protein forming the surface layer in Paracidovorax citrulli and evidence of its occurrence as a surface layer protein in diverse genera of the Betaproteobacteria and Gammaproteobacteria

doi: 10.1099/acmi.0.000685.v3

Figure Lengend Snippet: Betaproteobacteria possessing an npdA ortholog*

Article Snippet: Paracidovorax anthurii , DSM-16745 , DSMZ.

Techniques:

Journal: FASEB journal : official publication of the Federation of American Societies for Experimental Biology

Article Title: IPF Pathogenesis is Dependent upon TGFβ Induction of IGF-1

doi: 10.1096/fj.201901719RR

Figure Lengend Snippet: Summary of antibodies used for Western blot and immunoprecipitation.

Article Snippet: p-SGK1 , sc-16745-R , WB , 1:1,000 , Santa Cruz.

Techniques: Western Blot, Immunoprecipitation

Journal: FASEB journal : official publication of the Federation of American Societies for Experimental Biology

Article Title: IPF Pathogenesis is Dependent upon TGFβ Induction of IGF-1

doi: 10.1096/fj.201901719RR

Figure Lengend Snippet: (A) Top: RT-qPCR of Igf-1 gene expression in AKR-2B cells pre-treated for 2 hours with the indicated TGFβ-pathway inhibitors (10 μM SB431542, TGFβ Type-I Receptor (T1R) inhibitor; 20 μM LY294002, PI3K inhibitor; 300 nM MK2206, AKT inhibitor; 10 μM U0126, MEK inhibitor; 10 nM Rapamycin, mTOR inhibitor) or vehicle (0.1% DMSO), followed by stimulation with TGFβ for 12 hours (n=4). Bottom: Western blot of IGF-1 in AKR-2B cells as treated for qPCR and controls show the functionality of inhibitors on target protein activation following 3 (p-ERK/ERK) or 6 hours (p-SMAD3/SMAD3, p-AKT/AKT, p-S6K/S6K, p-SGK1/SGK1) treatment (n=3). (B) Top: RT-qPCR of Igf-1 expression in AKR-2B cells with shRNA-mediated stable knockdown of SMAD2 (sh-SMAD2), SMAD3 (sh-SMAD3), or nontargeting (sh-NT) after 12 hours TGFβ stimulation (n=3). Bottom: Western blot of IGF-1 in AKR-2B KD cells as treated for qPCR and controls confirming the level of knockdown of SMAD2, SMAD3, and p-SMAD2 or p-SMAD3 at 6 hours with TGFβ (+) or vehicle (−) (n=3). (C) Top: RT-qPCR of Igf-1 gene expression in AKR-2B cells with shRNA-mediated stable knockdown of mTOR (sh-mTOR) or NT (sh-NT) following 12 hours TGFβ stimulation (n=4). Bottom: Western blot of IGF-1 in AKR-2B KD cells as treated for qPCR and controls confirm the level of mTOR KD at 6 hours in the presence or absence of TGFβ (n=3). (A-C) Relative fold gene expression by TGFβ compared to control is indicated at the top of the vehicle or NT bars. Data are presented as means −/+ Standard Error of the Mean (SEM) for the number of biological replicates indicated (n). Statistical significance was determined after computing single factor ANOVA and/or unpaired two-tailed Student’s t-test (p<0.05 (*), p<0.01 (**), p<0.005 (***), p<0.001 (****). Results demonstrate that TGFβ-induction of Igf-1 transcription and translation requires SMAD2 & mTOR signaling.

Article Snippet: p-SGK1 , sc-16745-R , WB , 1:1,000 , Santa Cruz.

Techniques: Quantitative RT-PCR, Gene Expression, Western Blot, Activation Assay, Expressing, shRNA, Knockdown, Control, Two Tailed Test

Journal: The Journal of Clinical Investigation

Article Title: RABL6A inhibits tumor-suppressive PP2A/AKT signaling to drive pancreatic neuroendocrine tumor growth

doi: 10.1172/JCI123049

Figure Lengend Snippet: (A) Schematic of the kinases and phosphatases that regulate AKT-S473 phosphorylation. P, phosphorylation; arrows, activating events; perpendicular bars, inhibiting events. (B) Western blots of BON-1 control (CON) and RABL6A knockdown (KD1 and KD2) cells showing selective loss of pAKT-S473 in RABL6A-depleted cells versus moderately increased phosphorylation of other mTORC2 substrates, SGK1 and PKCα. GAPDH was the loading control. (C) Relative phosphorylation of mTORC2 substrates in BON-1 CON, KD1, and KD2 cells was quantified by ImageJ. Data represent the mean ± SD from 3 independent experiments (*P < 0.05 and **P < 0.001 compared with CON, 2-way ANOVA and adjusted for multiple comparisons using the Bonferroni method). (D) Western blots of pAKT-S473 and pAKT-T308 following in vitro phosphatase assays using phosphorylated HA-tagged Myr-AKT as substrate to which the indicated amounts (μg) of BON-1 CON, KD1, or KD2 lysates were added. As controls, buffer (–) or the general phosphatase inhibitor potassium fluoride (KF), was added to substrate prior to the phosphatase reaction. (E) Relative levels of pAKT-S473, normalized to total HA-Myr-Akt, were quantified by ImageJ analysis of blots from 3 or more experiments in which 320 μg cell lysate was tested. *P < 0.005 KD (KD1 and KD2) versus CON, 2-way ANOVA adjusted for multiple comparisons using the Bonferroni test. (F) Western blots of pAKT-S473, AKT, and RABL6A following okadaic acid (OA) treatment (100 nM, 20 hours) in BON-1 CON and KD cells showing significant restoration of pAKT-S473 by PP2A inhibition. Vinculin was loading control. (G) Relative phosphorylation of AKT-S473 was quantified from 3 experiments. *P < 0.005 compared with untreated counterparts, 2-way ANOVA adjusted for multiple comparisons using the Bonferroni method. Western blots in B, D, and F are representative of 3 or more experiments.

Article Snippet: Antibodies against p-SGK1-S422 (no. SC-16745), p-PKCα-S657 (no. SC-12356), Nek2 (no. SC-55601), and CIP2A (no. SC-80659) were purchased from Santa Cruz.

Techniques: Western Blot, In Vitro, Inhibition