Journal: Frontiers in cell and developmental biology

Article Title: ORF3a-Mediated Incomplete Autophagy Facilitates Severe Acute Respiratory Syndrome Coronavirus-2 Replication.

doi: 10.3389/fcell.2021.716208

Figure Lengend Snippet: FIGURE 4 | SARS-CoV ORF3a cannot induce autophagy. (A) Sequence alignment between SARS-CoV-2 ORF3a and SARS-CoV ORF3a. (B–D) SARS-CoV ORF3a does not affect autophagy. HeLa-vector, HeLa-ORF3aSARS-CoV-2, or HeLa-ORF3aSARS-CoV cell lines were treated with BafA1 (100 nM) and endogenous LC3 puncta were immunostained (B) and quantified (C). Scale bar, 15 µm. Arrow: representative autophagosomes. Mean ± SEM; n = 50; ns and ****p < 0.0001 by one-way ANOVA and Bonferroni’s post hoc test. HeLa-vector or HeLa-ORF3aSARS-CoV cell lines were treated with BafA1 (100 nM) for 4 h and the cell lysates were collected for IB with indicated antibodies (D). (E) ORF3aSARS-CoV-2 but not ORF3aSARS-CoV interacts with endogenous UVRAG. HEK293T cells were transfected with Flag-ORF3aSARS-CoV or Flag-ORF3aSARS-CoV-2 and cell lysates were collected and subjected to IP and IB with indicated antibodies at 48h post-transfection. (F–H) SARS-CoV ORF3a does not affect the formation of UVRAG complex (F), Beclin 1 complex (G), or Atg14 complex (H). HEK293T cells were co-transfected with indicated plasmids and cell lysates were collected and subjected to IP and IB with indicated antibodies at 48 h post-transfection.

Article Snippet: Primary antibodies included: mouse Flag (Sigma, #F1804), rat FlagHRP (Biolegend, #637311), mouse HA (BioLegend, #901515), rabbit SQSTM1/p62 (Cell Signaling, #39749), rabbit LC3 (Cell Signaling, #3868), rabbit Beclin-1 (Cell Signaling, #3495), rabbit UVRAG (Cell Signaling, #13115), rabbit Atg14 (Cell Signaling, #96752), rabbit Rubicon (Cell Signaling, #8465), rabbit Vps34 (Cell Signaling, #4263), rabbit Atg3 (Cell Signaling, #3415), rabbit Atg5 (Cell Signaling, #12994), mouse SARS-CoV-2 N (GeneTex, #GTX635689), and mouse GAPDH (Santa Cruz, #365062).

Techniques: Sequencing, Plasmid Preparation, Transfection

Journal: PLoS Genetics

Article Title: Beclin 1 Is Required for Neuron Viability and Regulates Endosome Pathways via the UVRAG-VPS34 Complex

doi: 10.1371/journal.pgen.1004626

Figure Lengend Snippet: A . Hela cells treated with indicated siRNA (siCon is siControl; siBec is siBeclin 1) were treated with DMSO or Bafilomycin A1 (BafA). Representative composite fluorescence plots and normalized quantification of mean fluorescence intensity of 10,000 cells were determined from three separate experiments. Mean pHrodo dextran (PE-A) fluorescence; bars represent mean +/− s.e.m. (n = 3). DMSO pHrodo dextran siCon vs. siBec one sample t-test (two-tailed) p = 0.0283; BafA pHrodo dextran siCon vs. siBec one-tailed t-test p = 0.0246. B . Mean LysoSensor (Am Cyan-A) fluorescence intensity; bars represent mean +/− s.e.m. (n = 3). LysoSensor siCon vs. siBec one sample t-test (two-tailed) p = 0.0029. C . Anti-UVRAG, -Beclin 1, -Actin and –LC3 blots of HeLa cells lysed after indicated siRNA knock-down. Asterix indicates unspecific band. D . Receptor-mediated endocytosis as measured by EGFR internalization is inhibited in Becn1 deficient MEFs and rescued with re-introduction of beclin 1. Anti-EGFR, - UVRAG, -beclin 1 and –actin blots of control or Becn1 deficient MEFs lysed after indicated treatment with EGF. Asterix indicates unspecific band. Quantification of normalized EGFR/actin from 3 separate experiments. Bars represent mean +/− s.e.m. (n = 3); p = 0.0395, 0.0208 (one-tailed t-test).

Article Snippet: Immunoprecipitation was performed with 500 μg protein from post-nuclear supernatant, using antibodies against VPS34 (Echelon), UVRAG (Bethyl), or ATG14 (MBL) overnight.

Techniques: Fluorescence, Two Tailed Test, One-tailed Test, Knockdown, Control

Journal: PLoS Genetics

Article Title: Beclin 1 Is Required for Neuron Viability and Regulates Endosome Pathways via the UVRAG-VPS34 Complex

doi: 10.1371/journal.pgen.1004626

Figure Lengend Snippet: A . Table showing results from PI(3)P ELISA. Average 450 nm absorbance, p-value of comparison to Becn1 deficient MEFs. PI(3)P concentration is determined by comparison to a standard curve. Quantification of 450 nm absorbance of control, Becn1 deficient, and Becn1 revertant MEFs (6,5,6 replicates); statistics using a one-tailed t-test p = 0.0032, 0.0336. B . Anti-beclin 1, and -actin blots of MEF cell lysates. Con is control MEFs, def is Becn1 deficient MEFs, and rev is Becn1 revertant MEFs plus beclin 1. C . Autoradiograph of 32 P-labelled PI(3)P and anti-VPS34, -beclin 1, and -actin blots after VPS34 IP of lysates from indicated MEFs with or without wortmannin (Wrtm) treatment. D . Quantification of normalized 32 P-labelled PI(3)P and 32 P-PI(3)P/VPS34 (from IP) from 3 separate experiments and VPS34/actin (from input lanes) from 9 separate experiments (including ). Con is control MEFs, def is Becn1 deficient MEFs and def+bec is Becn1 revertant MEFs. Bars represent mean +/− s.e.m. (n = 3) con vs. def p = 0.0003, p = 0.0226, and p = 0.0002 top to bottom using a one sample t-test (two-tailed). UVRAG and Atg14-associated PI(3)P production is decreased in Becn1 deficient MEFs. E . Autoradiograph of 32 P-labelled PI(3)P and anti-VPS34, -UVRAG, -beclin 1, and -actin blots after UVRAG IP of lysates from indicated MEFs with or without wortmannin (Wrtm) treatment. Con is control MEFs, def is Becn1 deficient MEFs and def+bec is Becn1 revertant MEFs. Quantification of normalized 32 P-labelled PI(3)P from 3 separate experiments. Bars represent mean +/− s.e.m. (n = 3) con vs. def p<0.0001 using a one sample t-test (two-tailed). F . Autoradiograph of 32 P-PI(3)P and anti-VPS34, -Atg14L, -beclin 1, and -actin blots after Atg14L IP of lysates from indicated MEFs with or without wortmannin (Wrtm) treatment. Quantification of normalized 32 P-labelled PI(3)P from 3 separate experiments. Bars represent mean +/− s.e.m. (n = 3) con vs. def p = 0.0065 using a one sample t-test (two-tailed).

Article Snippet: Immunoprecipitation was performed with 500 μg protein from post-nuclear supernatant, using antibodies against VPS34 (Echelon), UVRAG (Bethyl), or ATG14 (MBL) overnight.

Techniques: Enzyme-linked Immunosorbent Assay, Comparison, Concentration Assay, Control, One-tailed Test, Autoradiography, Two Tailed Test

Journal: PLoS Genetics

Article Title: Beclin 1 Is Required for Neuron Viability and Regulates Endosome Pathways via the UVRAG-VPS34 Complex

doi: 10.1371/journal.pgen.1004626

Figure Lengend Snippet: A–E . Dispersion of GFP-p40 phox in Becn1 deficient MEFs is rescued with reintroduction of beclin 1 as well as overexpression of UVRAG but not Atg14L or RUBICON. Images of Becn1 deficient MEFs cotransfected with GFP-p40 phox and AsRed ( A ), beclin 1-AsRed ( B ), Atg14L-AsRed ( C ), RUBICON-AsRed ( D ) and UVRAG-Flag and stained with anti-Flag antibody ( E ). Scale bars = 10 µm. Quantification as in E. AsRed (5,26) vs. beclin 1-AsRed (28,3) significantly different (p<0.0001); AsRed (5,26) vs. Atg14L-AsRed (3,29) not significantly different (p = 0.4741); AsRed (5,26) vs. RUBICON-AsRed (5,27) not significantly different (p = 1.0000); AsRed (5,26) vs. UVRAG-Flag (26,4) significantly different (p<0.0001) all using Fisher's exact test (two-sided). Rescue of the aberrant GFP-p40 phox distribution in Becn1 deficient MEFs by beclin 1 overexpression requires the CC domain of beclin 1. A beclin 1 mutant lacking the CC domain does not rescue the GFP-p40 phox phenotype of Becn1 deficient MEFs. Immunofluorescent images of Becn1 deficient MEFs transfected with beclin 1-myc wt ( F ) or beclin 1-myc minus its UVRAG-binding CC (beclin 1ΔCCD-myc) ( G ) then fixed and stained with anti-myc. Scalebars = 20 µm. Quantification as in E. beclin 1-myc wt (29,1) vs. beclin 1ΔCCD-myc (28,3) significantly different (p<0.0001) using Fisher's exact test (two-sided).

Article Snippet: Immunoprecipitation was performed with 500 μg protein from post-nuclear supernatant, using antibodies against VPS34 (Echelon), UVRAG (Bethyl), or ATG14 (MBL) overnight.

Techniques: Dispersion, Over Expression, Staining, Mutagenesis, Transfection, Binding Assay

Journal: OncoTargets and Therapy

Article Title:

Suppressing UVRAG Induces Radiosensitivity by Triggering Lysosomal Membrane Permeabilization in Hypopharyngeal Squamous Cell Carcinoma

doi: 10.2147/ott.s270433

Figure Lengend Snippet: Figure 2 Irradiation upregulated UVRAG in HSCC. (A) Western blot analysis of UVRAG in Fadu cells treated with or without 4 Gy irradiation. GAPDH was used as a loading control. (B) Densitometric analysis of the blots showed the ratios of UVRAG to GAPDH. (C) Hematoxylin-eosin (HE) staining and immunohistochemistry of UVRAG in HSCC tumor tissues from primary and recurrent HSCC patients who have only received radiotherapy after first surgical resection. (D) Quantification of (C). **P < 0.01; Scale bars = 50 µm.

Article Snippet: Western blot analysis was performed as previously described.8 The following antibodies were used: LC3B, P62, GAPDH, UVRAG (Cell Signaling Technology; Danvers, MA, USA).

Techniques: Irradiation, Western Blot, Control, Staining, Immunohistochemistry

Journal: OncoTargets and Therapy

Article Title:

Suppressing UVRAG Induces Radiosensitivity by Triggering Lysosomal Membrane Permeabilization in Hypopharyngeal Squamous Cell Carcinoma

doi: 10.2147/ott.s270433

Figure Lengend Snippet: Figure 4 Inhibiting UVRAG interfered autophagy in Fadu cells. (A) LC3B and p62 levels were examined by Western blot analysis in Fadu cells treated with control or UVRAG siRNA. GAPDH was used as a loading control. Rapamycin was used to increase the basal level of autophagy. (B) Densitometric analysis of the blots showed the ratios of LC3B-II and P62 to GAPDH. (C) The Autophagy Tandem Sensor RFP-GFP-LC3B Kit was used to study autophagic flux in Fadu cells treated with control or UVRAG siRNA. (D) Quantification of red and green dots in (C). **P < 0.01; size bars = 10 µm.

Article Snippet: Western blot analysis was performed as previously described.8 The following antibodies were used: LC3B, P62, GAPDH, UVRAG (Cell Signaling Technology; Danvers, MA, USA).

Techniques: Western Blot, Control

Journal: PLOS Biology

Article Title: Vps34-orchestrated lipid signaling processes regulate the transitional heterogeneity and functional adaptation of effector regulatory T cells

doi: 10.1371/journal.pbio.3003074

Figure Lengend Snippet: ( A ) Experimental schematic for ablation of Vps34 in mature Tregs during early tumorigenesis. Control or Foxp3 Cre-ERT2 Pik3c3 fl/fl mice were subcutaneously (s.c.) inoculated with MC38 colon adenocarcinoma cells (5 × 10 5 ) or B16F10 melanoma cells (2.5 × 10 5 ; see – ) in the right flank. On days 7 to 11 after tumor inoculation, i.p. injections of TAM (2 mg/mouse dissolved in corn oil) were given daily for a total of 5 injections, and tumor size was measured every other day from 7 (MC38) or 11 (B16F10) days after tumor inoculation until endpoint tumor volume (1.5 × 10 3 mm 3 ) or humane endpoint was reached. i.p., intraperitoneal; TAM, tamoxifen. Mouse image created with BioRender. ( B − D ) Control ( n = 9) or Foxp3 Cre-ERT2 Pik3c3 fl/fl ( n = 8) mice were inoculated with MC38 tumors and treated with TAM as described in ( A ). Tumor growth curves (left) and tumor weights (right) at endpoint (day 21) in control or Foxp3 Cre-ERT2 Pik3c3 fl/fl mice ( B ). Quantification of the frequency of control or Vps34-deficient TCRβ + CD4 + GFP + YFP + Tregs derived from the spleen or MC38 tumor of control or Foxp3 Cre-ERT2 Pik3c3 fl/fl mice at day 21 after tumor inoculation ( C ). Quantification of the ratio of total TCRβ + CD8 + T cells to total TCRβ + CD4 + GFP + Tregs derived from the spleen or MC38 tumor of control or Foxp3 Cre-ERT2 Pik3c3 fl/fl mice at day 21 after tumor inoculation ( D ). ( E ) Tumor growth curves (left) and tumor weights (right) at endpoint (day 23) in control or Foxp3 Cre Atg14 fl/fl mice inoculated with MC38 tumors ( n = 5 per group). ( F ) Tumor growth curves (left) and tumor weights of non-ulcerated tumors (right) at endpoint (day 25) in control ( n = 4 for tumor growth; 3 for tumor weight) or Foxp3 Cre Uvrag fl/fl ( n = 6 for tumor growth; 3 for tumor weight) mice inoculated with MC38 tumors. ( G ) Violin plots showing the activity scores of a curated Vps34-activated eTreg signature (upper; i.e., top 200 [ranked by log 2 FC] downregulated genes [log 2 FC ≤ –0.5 and FDR < 0.05] in Vps34-deficient eTregs [transitional + terminal] versus control eTregs [transitional + terminal] from single-cell transcriptome profiling as described in ; see and for details) or curated Atg14-activated eTreg signature (lower; i.e., downregulated genes [log 2 FC ≤ −0.5 and P < 0.05] in Atg14-deficient eTregs versus control eTregs from mixed BM chimera mice; see and for details) in publicly available scRNA-seq datasets of human Tregs from PBMCs, LN, or tumors in public datasets (GSE139324 ; GSE114727 ; GSE239750 as indicated). BM, bone marrow; LN, lymph node; PBMC, peripheral blood mononuclear cells. Data are shown as mean ± s.e.m. ( B – F ). Two-way ANOVA (tumor volume; B , E , F ), Welch’s t test (tumor weight; B , E, F ), two-tailed Student t test ( C , D) , or Wilcoxon rank-sum test ( G ); NS, not significant. Data are representative of 6 ( B – D ) or 2 ( E , F ) independent experiments. The numerical data underlying the graphs shown in this figure are found in ( B – F ). Genes in the Vps34-activated eTreg signature or Atg14-activated eTreg signature are shown in and were applied to publicly available gene sets as described in the figure and figure legend ( G ).

Article Snippet: Quantitative PCR analysis was performed on the QuantStudio 7 Flex System (Applied Biosystems) using the following probes: Uvrag (Mm 01273471) and Actb (Mm 00607939_s1).

Techniques: Control, Derivative Assay, Activity Assay, Two Tailed Test

Journal: Oncotarget

Article Title: Potent antitumor activity of oncolytic adenovirus expressing Beclin-1 via induction of autophagic cell death in leukemia.

doi: 10.18632/oncotarget.1018

Figure Lengend Snippet: Figure 1: Schematic diagram of adenoviral constructs and CRAd infectivity in leukemic cells. (A) Compared to wild-type Ad5, SG511-BECN contains a human telomerase reverse transcriptase (hTERT) promoter-controlled E1 expression cassette in forward orientation, an E1B 55-kDa deletion, and Beclin-1 transgene in E3 region. The fiber was modified by an Ad5/11 chimeric fiber. (B) Kusumi-1 and K562 cells were infected with 50 MOI of SG511-GFP for 24 h and then observed under a fluorescence microscope (200×) (top panel). K562 cells were treated with the indicated amount of the virus for 24 h and then collected for analysis by FACS (bottom panel). (C) Bone marrow cells obtained from a patient with AML were infected with SG511-GFP at an MOI of 50, and then cultured for 5 days in colony culture assay. Photographs were viewed under a light microscope and fluorescence microscope, respectively (400×). (D) Beclin-1 and E1A expression was determined by Western blotting of K562 cells infected with SG511-BECN at the indicated concentrations for 48 h. Actin was Western blotted for equal loading. (E) Following SG511-BECN treatment for 48 h, Beclin-1 (green) and ER (red) in K562 cells were detected by confocal immunofluorescence microscopy. Representative data of three experiments is shown.

Article Snippet: Transfection of UVRAG siRNA vector K562 cells expressing GFP-LC3 were transiently transfected with human UVRAG-siRNA, ATG5 siRNA, or ATG7 siRNA, respectively, as per the manufacturer’s instructions, using shRNA Plasmid Transfection Reagent (Santa Cruz Biotechnology, Santa Cruz, CA, USA) or Lipofectamine Transfection Reagent (Invitrogen).

Techniques: Construct, Infection, Reverse Transcription, Expressing, Modification, Fluorescence, Microscopy, Virus, Cell Culture, Light Microscopy, Western Blot, Immunofluorescence

Journal: Oncotarget

Article Title: Potent antitumor activity of oncolytic adenovirus expressing Beclin-1 via induction of autophagic cell death in leukemia.

doi: 10.18632/oncotarget.1018

Figure Lengend Snippet: Figure 2: SG511-BECN virus induced cell death involves autophagy in leukemia cells. (A) Kasumi-1 and KG-1 cells were treated with SG511-BECN at the doses indicated. Two days later, cell death was assessed by FACS analysis of annexin V- and PI staining. (B) K562 cells were treated with SG511-BECN at the indicated doses for 48 h before cell lysates were immunoblotted for the protein indicated. (C) K562 cells were incubated with SG511-BECN, or incubated with z-VAD (10 ìM) and SG511-BECN for 72 h. Then, cell viability was determined by an MTT assay. *represents P>0.05 (D) Formation of AVOs was determined after leukemic cells were infected with or without SG511, or SG511-BECN at an MOI of 50 for 48 h. (E) Formation of GFP-LC3 vacules (dots) was determined after K562 cells were stably transfected with GFP-LC3 vector and treated with 50 MOI of SG511 or SG511-BECN for 48 h. (F) K562 cells were infected with or without the indicted viruses (50 MOI) for 48 h. The cell lysates were harvested and analyzed by Western blotting using anti- LC3 and anti-p62 antibodies. (G) Kasumi-1 cells treated with SG511 and SG511-BECN, respectively and observed under TEM (×4200). There are two types of vacuole in the cytoplasm of SG511-BECN-treated cells: dense multilamellar bodies (white arrows) and inclusion bodies (black arrows). A representative of two separate experiments is shown. (H) qRT-PCR analysis monitoring p62 expression in K562 cells treated with SG511-BECN. Bars represent SD. (I) K562 cells were infected with 50 MOI SG511-BECN, alone or in combination with 20 nM BafA1. Total cell lysates were immunoblotted with anti-LC3, anti-p62, or anti-Actin antibodies, as indicated.

Article Snippet: Transfection of UVRAG siRNA vector K562 cells expressing GFP-LC3 were transiently transfected with human UVRAG-siRNA, ATG5 siRNA, or ATG7 siRNA, respectively, as per the manufacturer’s instructions, using shRNA Plasmid Transfection Reagent (Santa Cruz Biotechnology, Santa Cruz, CA, USA) or Lipofectamine Transfection Reagent (Invitrogen).

Techniques: Virus, Staining, Incubation, MTT Assay, Infection, Stable Transfection, Transfection, Plasmid Preparation, Western Blot, Quantitative RT-PCR, Expressing

Journal: Oncotarget

Article Title: Potent antitumor activity of oncolytic adenovirus expressing Beclin-1 via induction of autophagic cell death in leukemia.

doi: 10.18632/oncotarget.1018

Figure Lengend Snippet: Figure 3: Superior efficacy of SG511-BECN compared with the CRAd expressing TRAIL. (A) Leukemic cells (2×104), human MNCs, and L02 cells were treated with Ad5-BECN, SG511, or SG511-BECN at an MOI of 50 for 72 h before analysis of cell viability (MTT assay). The results represent means ± SD of three independent experiments. *P<0.0001, vs SG511, # P=0.0007, vs SG511. (B) K562 cells were treated with the indicated viruses at 50 MOI and colonies were observed on day 7 under a light microscope. (C) K562, NB4, and THP-1 cells were infected with or without SG511, SG235-TRAIL, and SG511-BECN at an MOI of 50, respectively. The cells were then plated in methylcellulose medium. After incubation for 7 days, colonies (more than 50 cells) were scored. Data represent means ± SD for separate experiments. #SG511-BECN vs SG235-TRAIL, *SG511-BECN vs SG511.

Article Snippet: Transfection of UVRAG siRNA vector K562 cells expressing GFP-LC3 were transiently transfected with human UVRAG-siRNA, ATG5 siRNA, or ATG7 siRNA, respectively, as per the manufacturer’s instructions, using shRNA Plasmid Transfection Reagent (Santa Cruz Biotechnology, Santa Cruz, CA, USA) or Lipofectamine Transfection Reagent (Invitrogen).

Techniques: Expressing, MTT Assay, Light Microscopy, Infection, Incubation

Journal: Oncotarget

Article Title: Potent antitumor activity of oncolytic adenovirus expressing Beclin-1 via induction of autophagic cell death in leukemia.

doi: 10.18632/oncotarget.1018

Figure Lengend Snippet: Figure 5: siRNA-mediated UVRAG, ATG5, or ATG7 knockdown reverses suppressive effects of SG511-BECN. (A) Expression of autophagy gene UVRAG was silenced by siRNA in K562 cells containing GFP-LC3, and LC3-positive vesicle-containing cells were determined after treatment with SG511-BECN for 48 h. UVRAG expression was detected by Western blotting of K562 cells. â-actin was used as protein loading control (insert). (B) Experimental conditions as in (A). Cell viability was assessed by an MTT assay. *Represents P<0.001 compared to control. Standard error was calculated from three independent experiments. (C) K562 cells were transfected with 100 nM ATG5 siRNA for 48 h, and then treated with SG511-BECN for 48 h. The indicated protein level was analyzed by Western blot (insert). In parallel, cell viability was assessed by an MTT assay (P=0.0019). (D) ATG7 protein level and viability of K562 cells were determined after transfection with ATG7 siRNA as indicated in (C). *represents P<0.0029.

Article Snippet: Transfection of UVRAG siRNA vector K562 cells expressing GFP-LC3 were transiently transfected with human UVRAG-siRNA, ATG5 siRNA, or ATG7 siRNA, respectively, as per the manufacturer’s instructions, using shRNA Plasmid Transfection Reagent (Santa Cruz Biotechnology, Santa Cruz, CA, USA) or Lipofectamine Transfection Reagent (Invitrogen).

Techniques: Knockdown, Expressing, Western Blot, Control, MTT Assay, Transfection

Journal: Oncotarget

Article Title: Potent antitumor activity of oncolytic adenovirus expressing Beclin-1 via induction of autophagic cell death in leukemia.

doi: 10.18632/oncotarget.1018

Figure Lengend Snippet: Figure 6: Antileukemic efficacy of SG511-BECN in established K562 tumors in vivo. Mice bearing subcutaneous K562 xenografts (n=8) were randomized to receive one of the following intratumoral injections on 5 consecutive days: PBS, Ad5, SG511, and SG511-BECN. (A) Time courses of changes in tumor volume in each group (n=7) are shown. (B) Survival curve analysis. The percentage of surviving mice was determined by monitoring the death of mice over a period of 25 days. Mice treated with SG511-BECN shows significant survival advantage over mice treated with other groups (P=0.0072). (C) Tumor tissues were obtained on day 5 after treatment with different viruses, and Beclin-1 expression and the level of LC3-II was determined by Western blotting. The expression of â-actin was used as loading control. (D) Electron microscopy images were taken of tumor tissues (3700×). Autophagic vacuoles are denoted by arrow.

Article Snippet: Transfection of UVRAG siRNA vector K562 cells expressing GFP-LC3 were transiently transfected with human UVRAG-siRNA, ATG5 siRNA, or ATG7 siRNA, respectively, as per the manufacturer’s instructions, using shRNA Plasmid Transfection Reagent (Santa Cruz Biotechnology, Santa Cruz, CA, USA) or Lipofectamine Transfection Reagent (Invitrogen).

Techniques: In Vivo, Expressing, Western Blot, Control, Electron Microscopy

Journal: International Journal of Medical Sciences

Article Title: Effects Of Oral Glutamine on Inflammatory and Autophagy Responses in Cancer Patients Treated With Abdominal Radiotherapy: A Pilot Randomized Trial

doi: 10.7150/ijms.20245

Figure Lengend Snippet: TaqMan probes and GenBank accession numbers of inflammatory and autophagic genes.

Article Snippet: UVRAG , NM_003369.3 , Hs00163433_m1.

Techniques: