Journal: EMBO Molecular Medicine

Article Title: ERK5 suppression overcomes FAK inhibitor resistance in mutant KRAS-driven non-small cell lung cancer

doi: 10.1038/s44321-024-00138-7

Figure Lengend Snippet: ( A ) Percentage of proliferation inhibition of A549, A427 and H460 cell lines treated with increasing doses of XMD8-92 or Seliciclib or in combination (top) 48 h after treatment and representative crystal violet-stained cells 72 h after drug treatment (bottom). The combination index (CI) showing the synergistic effect of combination of the 2 drugs is indicated; n = 3. ( B – D ) Relative quantification of Annexin V (AV) + Annexin V/PI (AV/PI)-positive cells by flow cytometry; n = 4 ( B ), colony forming capacity; n = 3 ( C ) and percentage of cell migration; n = 10 ( D ) of A549 cells treated with 10 µM XMD8-92 or Seliciclib or in combination, except for the colony formation assay in which cells were treated with 5 µM of each drug. ( E ) Immunoblot analysis of the indicated targets in A549 cells treated with the indicated doses of Seliciclib or XMD8-92 or in combination for 24 h. ( F ) Immunoblot analysis of the indicated targets in A549 cells treated with 10 µM XMD8-92 or 10 µM Seliciclib or in combination for 24 h. ( G ) Immunoblot analysis for the indicated targets in A549 cells transduced with shRNA control (pLKO.1 hygro + Tet-pLKO-puro) or a shRNA against CDK5 (Tet-pLKO-puro-shCDK5 + pLKO.1 hygro), or a shRNA against ERK5 (pLKO.1 hygro-shERK5 + Tet-pLKO-puro) or in combination (Tet-pLKO-puro-shCDK5 + pLKO.1 hygro-shERK5). After transduction and selection, cells were harvested for protein extraction 72 h after doxycycline (1 μg/mL) induction. ( H , I ) Relative cell number ( H ) and Annexin V (AV) + Annexin V/PI (AV/PI)-positive cell quantification by flow cytometry ( I ) in A549 cells treated as in ( G ) 72 h after doxycycline (1 μg/mL) induction; n = 3. Graphical data are mean ± SD. Statistical analyses were done using one-way ANOVA; n , number of biologically independent samples. .

Article Snippet: Human: A427 cell line , Prof. John Minna (UTSW Medical Center, Dallas, USA) , ATCC # HTB-53.

Techniques: Inhibition, Staining, Quantitative Proteomics, Flow Cytometry, Migration, Colony Assay, Western Blot, Transduction, shRNA, Control, Selection, Protein Extraction

Journal: EMBO Molecular Medicine

Article Title: ERK5 suppression overcomes FAK inhibitor resistance in mutant KRAS-driven non-small cell lung cancer

doi: 10.1038/s44321-024-00138-7

Figure Lengend Snippet: Relative quantification of cell death by flow cytometry analysis of Annexin V-Atto 633 (AV) + Annexin V/PI (AV/PI)-positive (left) and colony number (right) of A427 cells treated with XMD8-92 or Seliciclib (10 µM for apoptosis assay and 2.5 µM for colony formation each, respectively) alone or in combination; n = 3. Graphical data are mean ± SD. Statistical analyses were done using one-way ANOVA; n , number of biologically independent samples.

Article Snippet: Human: A427 cell line , Prof. John Minna (UTSW Medical Center, Dallas, USA) , ATCC # HTB-53.

Techniques: Quantitative Proteomics, Flow Cytometry, Apoptosis Assay

Journal: EMBO Molecular Medicine

Article Title: ERK5 suppression overcomes FAK inhibitor resistance in mutant KRAS-driven non-small cell lung cancer

doi: 10.1038/s44321-024-00138-7

Figure Lengend Snippet: Reagents and tools table

Article Snippet: Human: A427 cell line , Prof. John Minna (UTSW Medical Center, Dallas, USA) , ATCC # HTB-53.

Techniques: Recombinant, Plasmid Preparation, Sequencing, shRNA, Reverse Transcription Polymerase Chain Reaction, Transfection, Western Blot, In Vitro, In Vivo, cDNA Synthesis, TUNEL Assay, Software

Journal: Cell Reports

Article Title: The protease SPRTN and SUMOylation coordinate DNA-protein crosslink repair to prevent genome instability

doi: 10.1016/j.celrep.2021.110080

Figure Lengend Snippet:

Article Snippet: Rabbit anti-53BP1 , Santa Cruz Biotechnology , Cat#sc-22760; RRID: AB_2256326.

Techniques: Virus, Subcloning, Recombinant, Staining, Picogreen Assay, Proliferation Assay, Flow Cytometry, DNA Extraction, Sequencing, Luciferase, Software, Transfection, Modification, Magnetic Beads, Membrane

Journal: eLife

Article Title: A TOPBP1 allele causing male infertility uncouples XY silencing dynamics from sex body formation

doi: 10.7554/eLife.90887

Figure Lengend Snippet:

Article Snippet: Antibody , Anti-BRCA1 (rabbit polyclonal) , Proteintech , 22362-1-AP , Western blot 1:1000.

Techniques: Sequencing, Mutagenesis, Western Blot, Purification, Injection, In Situ, Software

Journal: Cell Reports

Article Title: COX2 regulates senescence secretome composition and senescence surveillance through PGE 2

doi: 10.1016/j.celrep.2021.108860

Figure Lengend Snippet:

Article Snippet: Rat anti-CD8a-168Er , Fluidigm , # 3168003B, RRID: AB_2811241.

Techniques: Recombinant, Enzyme-linked Immunosorbent Assay, Plasmid Preparation, Software, Sequencing

Journal: Cell Reports Medicine

Article Title: A dual-STING-activating nanosystem expands cancer immunotherapeutic temporal window

doi: 10.1016/j.xcrm.2024.101797

Figure Lengend Snippet: D-SAM prolongs DC activation and CTL response without T cell apoptosis or exhaustion (A–E) Flow cytometric analysis of DC and CD8 + T cell activation in MC38-OVA tumor tissues after indicated injections at 5 μg cGAMP or 50 μg PC7A correspondingly. See also Figures S10 and . (F–H) Kinetics of DC maturation and CD8 + T cell infiltration in MC38-OVA tumors upon a single injection of Adu-S100 (5 μg) or D-SAM (containing 5 μg cGAMP). (I) Apoptosis of CD8 + T cells in tumors on day 9 in (F). (J–L) CD8 + T cell infiltration and exhaustion in MC38-OVA tumor tissues upon Adu-S100 (2.5 μg, 4 times) or D-SAM (containing 5 μg cGAMP, twice) treatment. Data are represented as mean ± SEM, n = 5 mice in each group. One-way ANOVA followed by Dunnett’s test, ∗: p < 0.05, ∗∗: p < 0.01, ∗∗∗: p < 0.001.

Article Snippet: InVivoMAb anti-mouse CD8α (clone 2.43) , BioXCell , Cat: BE0061; RRID: AB_1125541.

Techniques: Activation Assay, Injection

Journal: Cell Reports Medicine

Article Title: A dual-STING-activating nanosystem expands cancer immunotherapeutic temporal window

doi: 10.1016/j.xcrm.2024.101797

Figure Lengend Snippet: D-SAM shows synergistic antitumor efficacy with αPD1 against distal, recurring, and metastatic tumors (A–C) Injected primary and untreated contralateral tumor growth in C57BL/6 mice bearing MC38 tumors after indicated treatment. See also Figure S18 . (D and E) Rechallenged MC38 tumor growth after the initial MC38 tumor cured by D-SAM or D-SAM plus αPD1. See also Figure S19 . (F) Central memory and effector memory CD8 + T cells in the spleen of (D). (G–K) Primary tumor growth and lung metastasis in BALB/c mice bearing orthotopic triple-negative 4T1 breast tumor after different treatments. See also Figure S25 . Data are represented as mean ± SEM, n ≥ 5 mice in each group. Two-way ANOVA in (B), (C), (H), and (I) and one-way ANOVA followed by Dunnett’s test in (K). ∗: p < 0.05, ∗∗: p < 0.01, ∗∗∗: p < 0.001.

Article Snippet: InVivoMAb anti-mouse CD8α (clone 2.43) , BioXCell , Cat: BE0061; RRID: AB_1125541.

Techniques: Injection

Journal: Cell Reports Medicine

Article Title: A dual-STING-activating nanosystem expands cancer immunotherapeutic temporal window

doi: 10.1016/j.xcrm.2024.101797

Figure Lengend Snippet:

Article Snippet: InVivoMAb anti-mouse CD8α (clone 2.43) , BioXCell , Cat: BE0061; RRID: AB_1125541.

Techniques: Recombinant, Isolation, cDNA Synthesis, Enzyme-linked Immunosorbent Assay, RNA Sequencing, Software, Protease Inhibitor, Lysis

Journal: EMBO Reports

Article Title: m Tert induction in p21-positive cells counteracts capillary rarefaction and pulmonary emphysema

doi: 10.1038/s44319-023-00041-1

Figure Lengend Snippet: Reagents and tools.

Article Snippet: Rabbit anti-53BP1 , Novus , NB100.

Techniques: Isolation, Recombinant, Sequencing, TRAP Assay, Modification, Blocking Assay, Plasmid Preparation, Staining, Reverse Transcription, SYBR Green Assay, Red Blood Cell Lysis, Software, Imaging, Microscopy, Irradiation, Immunodetection, Immunohistochemistry

Journal: eLife

Article Title: Polycomb repressive complex 1.1 coordinates homeostatic and emergency myelopoiesis

doi: 10.7554/eLife.83004

Figure Lengend Snippet: ( A ) Strategy for analyzing Pcgf1 Δ/Δ hematopoietic cells. Total bone marrow (BM) cells (5 × 10 6 ) from Rosa26 CreERT and Rosa26 CreERT ;Pcgf1 fl/fl were transplanted into lethally irradiated CD45.1 recipient mice. Pcgf1 was deleted by intraperitoneal injections of tamoxifen at 4 wk post-transplantation. ( B ) The proportions of Mac-1 + and/or Gr-1 + myeloid cells, B220 + B cells, and CD4 + or CD8 + T cells among CD45.2 + donor-derived hematopoietic cells in the peripheral blood (PB) from control (n = 9) and Pcgf1 Δ/Δ (n = 14) mice. ( C ) Absolute numbers of total BM cells, hematopoietic stem cells (HSCs), multipotent progenitors (MPPs), myeloid progenitors, and CLPs (ALP and BLP) in a unilateral pair of femur and tibia 4 wk after the tamoxifen injection (n = 4–5). ( D ) Cell cycle status of CD34 – LSK HSCs and CD34 + LSK MPPs assessed by Ki67 and 7-AAD staining 4 wk after the tamoxifen injection. ( E ) In vivo limiting dilution assay. Limiting numbers of BM cells (1 × 10 4 , 4 × 10 4 , 8 × 10 4 , and 2 × 10 5 ) isolated from BM of primary recipients (control and Pcgf1 Δ/Δ mice after transplantation) were transplanted into sublethally irradiated secondary recipient mice with 2 × 10 5 of competitor CD45.1 BM cells (n = 5 each). Due to the low contribution of Pcgf1 Δ/Δ HSPCs to B cells, mice with chimerism of donor myeloid and T cells more than 1% in the PB at 16 wk after transplantation were considered to be engrafted successfully, and the others were defined as non-engrafted mice. The frequencies of HSPCs that contributed to both myeloid and T cells are indicated. ( F ) Strategy for analyzing Pcgf1 Δ/Δ hematopoietic cells. Total BM cells (2 × 10 6 ) from Rosa26 CreERT and Rosa26 CreERT ;Pcgf1 fl/fl CD45.2 mice were transplanted into lethally irradiated CD45.1 recipient mice with the same number of competitor CD45.1 BM cells. Pcgf1 was deleted by intraperitoneal injections of tamoxifen at 4 wk post-transplantation. Secondary transplantation was performed using 5 × 10 6 total BM cells from primary recipients at 4 mo post-intraperitoneal injections of tamoxifen. ( G ) The chimerism of CD45.2 donor cells in PB CD45 + leukocytes, Mac-1 + and/or Gr1 + myeloid cells, B220 + B cells, and CD4 + or CD8 + T cells in control and Pcgf1 Δ/Δ mice (n = 6 each) after the tamoxifen injection. ( H ) The chimerism of CD45.2 donor-derived cells in BM 4 wk after the tamoxifen injection (n = 5). ( I ) Absolute numbers of CD45.1 and CD45.2 total BM cells, HSCs, MPPs, myeloid progenitors, and Mac-1 + mature myeloid cells in a unilateral pair of femur and tibia 4 wk after the tamoxifen injection (n = 5). Statistical significance is based on the overall number of cells. ( J ) The chimerism of CD45.2 donor-derived cells in PB in primary (n = 3 each) and secondary (n = 4–5) transplantation. Data are shown as the mean ± SEM. *p<0.05, **p<0.01, ***p<0.001 by the Student’s t -test. Each symbol is derived from an individual mouse. A representative of more than two independent experiments is shown. Figure 1—source data 1. Raw data for .

Article Snippet: Antibody , Biotin anti-mouse CD8a (53–6.7) (rat monoclonal) , Tonbo Biosciences , Cat# 30-0081; RRID :AB_2621638 , FACS (9 × 10 –3 μL/1 × 10 6 cell).

Techniques: Irradiation, Transplantation Assay, Derivative Assay, Control, Injection, Staining, In Vivo, Limiting Dilution Assay, Isolation

Journal: eLife

Article Title: Polycomb repressive complex 1.1 coordinates homeostatic and emergency myelopoiesis

doi: 10.7554/eLife.83004

Figure Lengend Snippet: ( A ) White blood cell (WBC), hemoglobin (Hb), and platelet (PLT) counts in the peripheral blood (PB) from control (n = 9) and Pcgf1 Δ/Δ (n = 14) mice. ( B ) Absolute numbers of Mac-1 + and/or Gr-1 + myeloid cells, B220 + B cells, and CD4 + or CD8 + T cells 1 and 6 mo after the tamoxifen injection. ( C ) Representative flow cytometric profiles of bone marrow (BM), spleen, and thymus of control and Pcgf1 Δ/Δ mice 1 mo after the tamoxifen injection. The percentages of gated populations over parental populations are indicated. ( D ) Absolute numbers of total cells, Mac1 + myeloid cells, Mac1 + Ly6G + neutrophiles, Mac1 + Ly6C + Ly6C – monocytes, and Mac1 + F4/80 + macrophages in the BM and spleen (n = 5 each) 1 mo after the tamoxifen injection. ( E ) Histology of the BM from control and Pcgf1 Δ/Δ mice 1 mo after the tamoxifen injection observed by hematoxylin-eosin staining. ( F ) Absolute numbers of total cells, LSK cells, and myeloid progenitors in the spleen (n = 5 each) 1 mo after the tamoxifen injection. Data are shown as the mean ± SEM. *p<0.05, **p<0.01, ***p<0.001 by the Student’s t -test. Figure 1—figure supplement 2—source data 1. Raw data for .

Article Snippet: Antibody , Biotin anti-mouse CD8a (53–6.7) (rat monoclonal) , Tonbo Biosciences , Cat# 30-0081; RRID :AB_2621638 , FACS (9 × 10 –3 μL/1 × 10 6 cell).

Techniques: Control, Injection, Staining

Journal: eLife

Article Title: Polycomb repressive complex 1.1 coordinates homeostatic and emergency myelopoiesis

doi: 10.7554/eLife.83004

Figure Lengend Snippet: ( A ) Deletion efficiency of Pcgf1 and Cebpa in TSt-4 stromal co-culture in was assessed by genomic PCR in D45.2 + hematopoietic cells treated and non-treated with 4-OHT. Floxed, floxed allele; Δ, floxed allele after the removal of targeted exons by Cre recombinase (lower). ( B ) Rescue experiments of the myeloid-biased repopulation of Pcgf1 Δ/Δ HSPCs. LSK cells from Rosa26 CreERT (control) and Rosa26 CreERT ;Pcgf1 fl/fl mice were transduced with control (GFP), Pcgf1 , or Bmi1 retrovirus, then transplanted into lethally irradiated CD45.1 recipient mice. Pcgf1 was deleted by intraperitoneal injection of tamoxifen at 4 wk post-transplantation (upper). The proportions of myeloid (Mac-1 + and/or Gr-1 + ), B cells (B220 + ), and T cells (CD4 + or CD8 + ) among CD45.2 + GFP + control (n = 4–5) and Pcgf1 Δ/Δ (n = 5) donor-derived hematopoietic cells in peripheral blood (PB) at 3 mo after transplantation are indicated (* versus GFP control) (middle). RT-qPCR analysis of Bmi1 and Pcgf1 in GFP + LSK cells purified from bone marrow (BM) of recipient mice receiving control or Pcgf1Δ/Δ LSK cells transduced with control, Bmi1 or Pcgf1 retrovirus (n = 3). Hprt1 was used to normalize the amount of input RNA (lower; n=3). Data are shown as the mean ± SEM. *p<0.05; **p<0.01; ***p<0.001 by the Student’s t -test. ( C ) Snapshots of RNA-seq signals at the Cebpe gene loci. Figure 2—figure supplement 1—source data 1. Uncropped gel images of . Figure 2—figure supplement 1—source data 2. Raw data for .

Article Snippet: Antibody , Biotin anti-mouse CD8a (53–6.7) (rat monoclonal) , Tonbo Biosciences , Cat# 30-0081; RRID :AB_2621638 , FACS (9 × 10 –3 μL/1 × 10 6 cell).

Techniques: Co-Culture Assay, Control, Transduction, Irradiation, Injection, Transplantation Assay, Derivative Assay, Quantitative RT-PCR, Purification, RNA Sequencing

Journal: eLife

Article Title: Polycomb repressive complex 1.1 coordinates homeostatic and emergency myelopoiesis

doi: 10.7554/eLife.83004

Figure Lengend Snippet: ( A ) Kaplan–Meier survival curves of control (n = 7) and Pcgf1 Δ/Δ (n = 25) mice after the tamoxifen injection. ( B ) White blood cell (WBC) and hemoglobin (Hb) in peripheral blood (PB) from control (n = 8) and moribund Pcgf1 Δ/Δ myeloproliferative neoplasms (MPN) mice (n = 6). Bars indicate median values. ( C ) Absolute numbers of total bone marrow (BM) cells and spleen weight in control (n = 8) and moribund Pcgf1 Δ/Δ MPN mice (n = 6). ( D ) The proportions of Mac-1 + and/or Gr-1 + myeloid cells, B220 + B cells, and CD4 + or CD8 + T cells in PB and BM in control (n = 8) and moribund Pcgf1 Δ/Δ MPN mice (n = 6). Data are shown as the mean ± SEM. ( E ) Representative flow cytometric profiles of PB, BM, and spleen of control and moribund Pcgf1 Δ/Δ MPN mice. The percentages of gated populations over CD45.2 + live cells are indicated. ( F ) Representative histology of BM and spleen from control and moribund Pcgf1 Δ/Δ MPN mice observed by hematoxylin-eosin staining (left). The high-power field images of BM from control and moribund Pcgf1 Δ/Δ MPN mice observed by hematoxylin-eosin staining (right). ( G ) Representative flow cytometric profiles of thymus from control mice and moribund Pcgf1 Δ/Δ T-ALL mice. ( H ) Model for the stage-specific roles of non-canonical PRC1 in hematopoietic differentiation. *p<0.05; **p<0.01; ***p<0.001 by the Student’s t -test. Each symbol is derived from an individual mouse. Figure 6—source data 1. Raw data for .

Article Snippet: Antibody , Biotin anti-mouse CD8a (53–6.7) (rat monoclonal) , Tonbo Biosciences , Cat# 30-0081; RRID :AB_2621638 , FACS (9 × 10 –3 μL/1 × 10 6 cell).

Techniques: Control, Injection, Staining, Derivative Assay

Journal: eLife

Article Title: Polycomb repressive complex 1.1 coordinates homeostatic and emergency myelopoiesis

doi: 10.7554/eLife.83004

Figure Lengend Snippet:

Article Snippet: Antibody , Biotin anti-mouse CD8a (53–6.7) (rat monoclonal) , Tonbo Biosciences , Cat# 30-0081; RRID :AB_2621638 , FACS (9 × 10 –3 μL/1 × 10 6 cell).

Techniques: Produced, Sequencing, Recombinant, Protease Inhibitor, Purification, Staining, Avidin-Biotin Assay, Blocking Assay, Plasmid Preparation, Software, Immunostaining

Journal: Cell Reports

Article Title: Genome-wide Screens Implicate Loss of Cullin Ring Ligase 3 in Persistent Proliferation and Genome Instability in TP53 -Deficient Cells

doi: 10.1016/j.celrep.2020.03.029

Figure Lengend Snippet:

Article Snippet: Anti-53BP1 Rabbit polyclonal antibody , Bethyl Laboratories , Cat#A300-272A; RRID: AB_185520.

Techniques: Recombinant, Ligation, Viability Assay, Sequencing, Plasmid Preparation, Software, Infection, Transfection, Staining

Journal: Immunity

Article Title: Differentiation and protective capacity of virus-specific CD8 + T cells suggest murine norovirus persistence in an immune-privileged enteric niche

doi: 10.1016/j.immuni.2017.09.017

Figure Lengend Snippet: (A) Sequence of epitope P1519 in the wild-type and mutant MNV strains used in this study. (B) RAW-264.7 cells were infected with the indicated MNV strains at a multiplicity of infection (MOI) of 0.1 and viral load in the culture medium was measured by qPCR at the indicated time points. Representative of 3 experiments with 2 replicates per experiment. (C) Mice were infected orally with the indicated MNV strains and P1519-specific CD8+ T cells enumerated on day 8 p.i. in the indicated tissues. (D) Summary of data from (C). Representative of 3 experiments with 5 mice per group. (E) Percent of Tet+CD8+ T cells among splenic CD44hiCD62LloCD8+ T cells responding to MNV. Error bars indicate standard error of the mean (SEM). *Unpaired t-test (p<0.05). (F) Magnitude and (G–H) cytokine production and coproduction by intestinal Tet+CD8+ T cells at day 41 p.i. with MNV-CR6WT or MNV-CR6F→Y. Colors in (H) represent number of chemokines or cytokine coproduced. (I) Shedding and (J) tissue titers of wild-type and mutant viruses measured by qPCR. Error bars show SEM. Representative of 3 experiments with 5 mice per group. (K) Granzyme B expression and (L) in vitro cell killing by Tet+CD8+ T cells induced by MNV-CW3WT or MNV-CR6F→Y. (M) P1519 sequences from stool of chronically infected mice. See also Figures S1 and S2.

Article Snippet: Depletion of CD8 + T cells was performed using anti-mouse CD8β (Lyt 3.2) antibody, clone 53-5.8 from Bio X Cell, Lebanon, NH.

Techniques: Sequencing, Mutagenesis, Infection, Expressing, In Vitro

Journal: Immunity

Article Title: Differentiation and protective capacity of virus-specific CD8 + T cells suggest murine norovirus persistence in an immune-privileged enteric niche

doi: 10.1016/j.immuni.2017.09.017

Figure Lengend Snippet: Mice were infected orally with MNV-CW3WT or MNV-CR6F→Y and at day 30 p.i. Tet+CD8+ T cells were sorted from intestinal epithelium and analyzed by microarray. (A) Genes differentially expressed by at least 2-fold in Tet+CD8+ T cells from the 2 infections (ANOVA P value <0.05). (B–C) Enrichment of gene sets defined by Gene Ontology (GO) term in Tet+CD8+ T cells. Parts A–C represent 3–4 biological replicates per group, and each biological replicate consists of pooled cells from 5 individual mice. (D) Protein expression for several differentially expressed genes from (A). Representative of 2 independent experiments with 4 mice per group.

Article Snippet: Depletion of CD8 + T cells was performed using anti-mouse CD8β (Lyt 3.2) antibody, clone 53-5.8 from Bio X Cell, Lebanon, NH.

Techniques: Infection, Microarray, Expressing

Journal: Immunity

Article Title: Differentiation and protective capacity of virus-specific CD8 + T cells suggest murine norovirus persistence in an immune-privileged enteric niche

doi: 10.1016/j.immuni.2017.09.017

Figure Lengend Snippet: (A–B) Expression of CD103 and CD69 on Tet+CD8+ T cells in the small intestine and colon at day 30 p.i. with MNV-CW3WT or MNV-CR6F→Y. (C) Summary of data from (A). Error bars show SEM. (D) Expression of CD11a and CD49d on Tet+CD8+ T cells at day 30 p.i. with MNV-CW3WT or MNV-CR6F→Y. A–D are representative of 3 experiments with 5 mice per group. (E) MNV-CW3WT-infected mice were injected i.v. with anti-CD8α to label blood accessible cells. Three minutes later, tissues were harvested and labeled (blood-accessible) and unlabeled (tissue) CD8α+ T cells analyzed. The fraction of CD8+ T cells inaccessible to i.v. staining is shown in the green gates and numbers. (F) The fraction of CD103lo Tet+CD8+ T cells among non-circulating CD8+ T cells was enumerated. Gated on CD8α−(green gate) cells from (E). E–F are representative of 2 experiments with 5 mice per group. (G–I) Enrichment for Trm cell signature genes in Tet+CD8+ Tells from MNV-CW3WT versus MNV-CR6F→Y infection at indicated times p.i. (J) Core enrichment genes for (G) and (I). (K) Viral shedding and (L) titers in mice infected with MNV-CW3WT, MNV-CR6F→Y, or both. (M) Mean fluorescence intensity of CD103 on Tet+ cells from the LP in (K). K–M are representative of 2 experiments with 5 mice per group. *Unpaired t-test (p<0.05). Error bars show SEM. See also Figure S3.

Article Snippet: Depletion of CD8 + T cells was performed using anti-mouse CD8β (Lyt 3.2) antibody, clone 53-5.8 from Bio X Cell, Lebanon, NH.

Techniques: Expressing, Infection, Injection, Labeling, Staining, Fluorescence

Journal: Immunity

Article Title: Differentiation and protective capacity of virus-specific CD8 + T cells suggest murine norovirus persistence in an immune-privileged enteric niche

doi: 10.1016/j.immuni.2017.09.017

Figure Lengend Snippet: Gene set enrichment of published (A) exhaustion and (B) effector CD8+ T cell signatures in Tet+CD8+ T cells at day 30 p.i. with MNV-CR6F→Y versus MNV-CW3WT. (C) Genes contributing simultaneously to enrichment of exhaustion and effector signatures from (A) and (B) identified by leading edge analysis. (D) Enrichment of MCMV inflationary effector genes in Tet+CD8+ T cells from day 30 MNV-CR6F→Y versus MNV-CW3WT infection. (E) MNV-CR6F→Y normalized enrichment scores for the exhaustion and effector signatures from (A–B) and (D). (F) Genes contributing to enrichment of both effector signatures from (B) and the MCMV inflationary effector signature from (D). (G) PD-1 expression and abundance of Tet+CD8+ T cells at day 60 p.i. with MNV-CW3WT or MNV-CR6F→Y. Part G is representative of 2 experiments with 5 mice per group. See also Figure S4.

Article Snippet: Depletion of CD8 + T cells was performed using anti-mouse CD8β (Lyt 3.2) antibody, clone 53-5.8 from Bio X Cell, Lebanon, NH.

Techniques: Infection, Expressing

Journal: Immunity

Article Title: Differentiation and protective capacity of virus-specific CD8 + T cells suggest murine norovirus persistence in an immune-privileged enteric niche

doi: 10.1016/j.immuni.2017.09.017

Figure Lengend Snippet: (A–C) Splenic CD8+ T cells were isolated from mice chronically infected with MNV-CR6F→Y (day 60 p.i.), labeled with CTV, and adoptively transferred into congenic recipients (9×106 CD8+ T cells/recipient). Recipients were (A) naïve, or (B–C) at the indicated time point p.i. with MNV-CW3WT or MNV-CR6F→Y. (A–C) Frequency and proliferative history (CTV dilution) of donor Tet+CD8+ T cells 5 days post adoptive transfer in the indicated tissues. (D) Summary of (A–C) showing percent Tet+ among donor CD8+ T cells. Error bars show SEM. Representative of 2 independent experiments with 4 mice per group. (E–G) Adoptive transfer was carried out as above using Thy1.1 and Thy1.2 as congenic markers. Splenic CD8+ T cells were isolated from mice chronically infected with MNV-CR6F→Y (day 30 p.i.), labeled with CTV, and adoptively transferred into congenic recipients (5×106 CD8+ T cells/recipient). Recipients were (E) naïve, or (F–G) at the indicated time point p.i. with MNV-CR6F→Y or MNV-CW3WT. In (E) and (F) some recipient groups were pretreated with anti-Thy1.1 antibody prior to transfer. Gates in green show frequency of adoptively transferred donor cells. Plots in green show Tet staining and CTV dilution of transferred donor cells. (H) Summary of (E–G) showing percent Tet+ among donor CD8+ T cells. Error bars show SEM for four mice per group. Representative of 2 independent experiments with 4 mice per group. *Unpaired t-test (p<0.05). See also Figure S5.

Article Snippet: Depletion of CD8 + T cells was performed using anti-mouse CD8β (Lyt 3.2) antibody, clone 53-5.8 from Bio X Cell, Lebanon, NH.

Techniques: Isolation, Infection, Labeling, Adoptive Transfer Assay, Staining

Journal: Immunity

Article Title: Differentiation and protective capacity of virus-specific CD8 + T cells suggest murine norovirus persistence in an immune-privileged enteric niche

doi: 10.1016/j.immuni.2017.09.017

Figure Lengend Snippet: Mice were immunized with MNV-CW3WT or MNV-CW3Y→F and 30 days later challenged with MNV-CR6F→Y or MNV-CR6WT. (A) Viral shedding and (B) tissue titers were measured in challenged mice. (C) Summary of 3 independent experiments as outlined in (A–B). (D) P1519 sequences from the proximal colon of 6 immunized mice that became chronically infected after MNV-CR6F→Y challenge (box in (B). (E) Mice were immunized with MNV-CW3WT, then challenged with MNV-CR6F→Y as above. The effect of CD8+ T cell depletion on protection against MNV-CR6F→Y was tested on the indicated days post challenge by measuring viral titers in the colon. *Unpaired t-test (p<0.05). See also Figure S6.

Article Snippet: Depletion of CD8 + T cells was performed using anti-mouse CD8β (Lyt 3.2) antibody, clone 53-5.8 from Bio X Cell, Lebanon, NH.

Techniques: Infection

Journal: Immunity

Article Title: Differentiation and protective capacity of virus-specific CD8 + T cells suggest murine norovirus persistence in an immune-privileged enteric niche

doi: 10.1016/j.immuni.2017.09.017

Figure Lengend Snippet: In vivo and in vitro detection of MNV by Tet+CD8+ T cells is inefficient after the initial week of infection.

Article Snippet: Depletion of CD8 + T cells was performed using anti-mouse CD8β (Lyt 3.2) antibody, clone 53-5.8 from Bio X Cell, Lebanon, NH.

Techniques: In Vivo, In Vitro, Infection

Journal: Immunity

Article Title: Differentiation and protective capacity of virus-specific CD8 + T cells suggest murine norovirus persistence in an immune-privileged enteric niche

doi: 10.1016/j.immuni.2017.09.017

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Depletion of CD8 + T cells was performed using anti-mouse CD8β (Lyt 3.2) antibody, clone 53-5.8 from Bio X Cell, Lebanon, NH.

Techniques: Virus, Recombinant, Saline, Reverse Transcription, Gene Expression, SYBR Green Assay, Isolation, Staining, Microarray, Plasmid Preparation, Software