EMBO/sd-character-level-ner huggingface dataset:
(E) Left: Time course of normalized EGFP signal, as assessed by TIRF microscopy, from GRAMD1 triple knockout (TKO) HeLa cells expressing EGFP-GRAM1b together with either mCherry for control or mCherry-Lact-C2 for PS masking. SMase treatment (100 mU/ml) is indicated. Right: Values of ΔF/F0 corresponding to the end of the experiment as indicated by the arrow [mean ± SEM, n = 53 cells (mCherry), n = 32 cells (mCherry-Lact-C2 OE); data are pooled from four independent experiments for each condition; two-tailed unpaired Student's t-test, **P < 0.0001].
Histological examinations of tumor types in (F) K versus KrasG12D; NanogL/L (KN) Tumor tissues collected form indicated mice analyzed using Haematoxylin Eosin (H&E) staining. Representative H&E images are shown. Scale bar, 500 μm (top), 50 μm (bottom).
(A Total PD-L1, PARP from colon (Colo-205), lung (A549), pancreatic (PANK1) lysates treated with indicated DR5 agonist antibodies named Lexa (lexatumumab), KMTR2 GAPDH is loading control
(B) Quantitation of three independent IB experiments by imageJ of non-saturated blots for TAK1, MKK7, TAB1 and TAB2 expression on phagosomes of resting M0 macrophages and M2(IL-4) macrophages. Error bars represent SEM. *p<0.01, **p<0.001 (Student's T-test)
H The biofilm formation of the indicated strains was displayed with crystal violet staining (up) and quantified with optical density measurement (down).
Top, DNA content (n/2n) of exponentially growing yeast cells. First blot: 2D gel electrophoresis of the distribution of Lk topoisomers (Lk) of the YEp13 DNA in cells quenched at 26oC and after shifting the culture to 35oC for 60 min. Second blot: 2D gel electrophoresis of the same samples upon nicking the YEp13 DNA in order to reveal the occurrence of knots (kn). (B) Experiments conducted as in (A) but in scc1-73 yeast cells. Graph: Pkn of YEp13 before and after the inactivation of cohesin.
Primary hepatocytes were obtained from 12-week-old male WT (Prom1+/+) and KO (Prom1-/-) mice or KO (Prom1-/-) hepatocytes infected with an adenovirus harboring LacZ (Ad-LacZ) or Prom1 (Ad-Prom1) for 24 hours. The molecular interaction between Prom1 and PKA regulatory subunits (B) in these hepatocytes was determined by a proximity ligation assay using anti-Prom1 and anti-PKA regulatory subunits (B), respectively. Scale bar, 20 μm.
JQ1 pre-treatment inhibits N9 microglial cell inflammation, proliferation, and migration. N9 microglial cells were pre-incubated with vehicle (DMSO), JQ1 (0.5 μM), RVX208 (30 μM), or Olinone (30 μM) for 12 h followed by stimulation with LPS (1 μg/ml) for another 2 h, and then subjected to qRT-PCR (a) for determination of inflammatory cytokine expression. Proliferation assay (b, BrdU ELISA) and migration assay (c, Transwell) were performed after incubation for additional 24 h. Quantification: mean ± SEM; n = 3 experiments; *P < 0.05, **P < 0.01 compared to vehicle + LPS
(A) Immunohistochemistry analysis of MSR1 in patient ovarian, uterus and sarcoma cancers showing tumour-associated macrophages.
Q-V Representative images of lectin stained choroidal flat mounts 7 and 14 days after laser coagulation in Cx3cr1CreER (Q, R), Ifnar1fl/fl (S, T) and Cx3cr1CreER:Ifnar1fl/fl (U, V) mice. Dashed lines indicate CNV areas and the asterisk marks the central optic nerve head. Scale bar: 200 µm.W Quantification of lectin-stained CNV areas with ImageJ software. Bars show mean ± SD. (n = 4-11 RPE/choroidal flat mounts; One-way ANOVA followed by Tukey's post-test: *p = 0.043, ***p = 0.0007).
C, D, OTUB1 suppression impairs the anchorage-independent growth of Western blot cancer cell lines. Representative images of soft-agar colonies formed by Western blot cells expressing shRNAs against OTUB1 or GFP. The number of soft agar colonies formed by cells expressing shOTUB1 compared to cells expressing shGFP. Data are presented as mean ± s.e.m. p-values were determined by two-sided t-Test, n=3.
(H) The blood vessel-specific and miR126-independent KO of EGFL7 in EGFL7fl/fl;Cdh5-CreERT2 mice prolonged the overall survival of GL261 tumor-bearing mice as compared to WT littermates (38 d in KO vs. 33 d in WT; n = 7 for KO; n = 5 for WT; log-rank test). All data are presented as mean ± SEM. Scale bars represent 5 mm
MCF10A cells were treated with either EtOH or 25uM 4HNE for 60 minutes. Isolated protein was assessed by immunoprecipitating phospho-p38 (Thr180/Tyr182) and immunoblotting for 4HNE and phospho-p38 (Thr180/Tyr182). Shown is one replicate of three independent experiments (n=3).
(B, C) Body weight of the WT and L-HRD KO mice at the age of 12 weeks. (n=5 for each group) Data information: The data are representative of three independent experiments (mean ± s.d.). *: P<0.05. **: P<0.01 by unpaired student's t test.
(C) Occupancy (% of IP-ed DNA / input DNA) of GAL4DBDCBX2, RNF2, DAXXMycHis, H3.3, and H3 proteins at 5×UAS site as measured by ChIP-qPCR of chromatin from HEK2935xUAS cells. Data are means ± SEM (n = 3).
F: Classification based on the conformation of the regulatory particle in TDP-25 inclusions compared to non-transduced control neurons (Asano et al., 2015) and poly-GA inclusions (Guo et al., 2018). GS = Ground State, SPS = Substrate Processing State. Due to the uncertainties inherent to the classification procedure, it is possible that a small fraction of particles adopted other conformations.
E. Induction of PEBP1 by EGCG or DHA in HIV-1 infected primary CD4+ T cells. Primary CD4+ T cells from healthy donors were treated with 10 μM EGCG or DHA during HIV-1 infection. Similar to Panel B, HIV-1 was isolated from the blood supernatants of patients receiving ART with a viral load of 129 copies/ml. Seventy-two hours post-treatment with 10 µM EGCG or DHA, the expression of PEBP1 was detected by qPCR.
(B) The apoptotic response in U2OS cells with expression of wild-type and mutant KLF4. U2Os cells with wildtype (pLenti-tet-on-KLF4WT) or mutant (pLenti-tet-on-KLF4YYR/AAA) KLF4 were incubated with 10 ng/ml doxycycline for 24h and then treated with 10uM doxorubicin (Dox) or cisplatin (CDDP) for additional 24h. While expression of wild-type KLF4 decreases Bax expression and inhibits drug-induced PARP1 cleavage, expression of PARylation-deficient KLF4 leads to failure in inhibiting genotoxic-induced Bax expression and PARP1 cleavage.
RIP assay was performed from HeLaS3_FH AGO2 and HeLaS3_GFP cell extract using anti-HA antibody or IgG, as negative control, followed by TERC detection by RT-qPCR. 7SK RNA was used for normalization (n=3 experimental replicates).
D. Immunoblot analysis of FOXM1 and SATB2 expression in GSCs transduced with shNT or shSATB2.
(a) Immunoblots showing active caspase-1 (casp-1) p20 compared to GAPDH loading controls in wild-type (WT), Ripk2−/− and Nod2−/− BMDCs after PR8 infection.
B Quantitative PCR of FBXO5 transcripts in FACS-sorted cells with moderate or high levels of E2F7/8 expression, after 8 hours of doxycycline treatment. To avoid bias from cell cycle defects, cells were released from HU arrest at the onset of doxycycline treatment, resulting in a strong enrichment of cells in late S or G2 of both vehicle and doxycycline-treated cells (see Fig EV3C).
Normal, si-control-transfected, or si-PTEN-transfected SGC7901 cells were treated with CDDP (1 µg/ml) and M2-si-control-exo or M2-si-CRNDE-exo. Si-control, the negative control of si-PTEN. The (B) survival rate, in SGC7901 cells. Data information: The results shown are from seven biological replicates. Data are expressed as mean ± SD. : one-way ANOVA. **P<0.01.
(A) Blue-native gel electrophoresis and immunoblots of cells expressing the wild type form of Cor1 (Cor1WT), as well as the mutants Cor1N63A, N187A, D192A (Cor1*) and Cor1N63A, N187A, D192A, V189A, Y65A, L238A, K240A (Cor1**) in wild type (WT) background as well as in cells lacking CRD1 (crd1Δ). Blots were probed with antibodies against Cor1 to visualize Complex III (CIII), as well as Cox1, to monitor Complex IV (CIV). CV: Complex V; Cor1Cor2: multimer consisting of Cor1 and Cor2
(A,B) Expression of quantitative RT-PCR+ (A) using quantitative RT-PCR of RNA extracted from cells grown either in rich medium (dark gray) or starved for 2 hours (light grey). Cells of the following genotypes were analyzed: wild-type isogenic controls (WT) and taf12-5A mutants. act1+ served as a control for normalization across samples. Values from a WT strain grown in rich medium were set at 1 to allow comparisons across culture conditions and mutant strains. Each column represents the mean value of 6 independent experiments, overlaid with individual data points and SE. Statistical significance was determined by 2-way ANOVA followed by Bonferroni's multiple comparison tests (n = 6).
Ttyh1-transduced E14.5 neural progenitors were incubated for 2 days in the presence or absence of DAPT, and samples were prepared for qPCR for Notch target genes (I)
(K) ER (-), PR (-) and HER2 (-) UCD52 patient derived tumor tissue was xenografted in breast fat pad of NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ mice, following by treatment with IgG1 or KMTR2 (50μg, 4 doses). Harvested tumors were analyzed for PD-L1, CD47 and PARP in lysates.
Metabolomics profile of GC muscle of 4-month-old HSA-Cre and control mice presented as heatmap visualization and hierarchical clustering analysis of compounds
Schematic domain overview of endophilins-A1-3, highlighting the BAR and SH3-domains.
(d) Quantification (FACS analysis) of ROS accumulation using DHE→Eth conversion of wild-type (WT) and Δspe1 cells with and without supplementation of low doses of spermidine. Data represent means ± s.e.m. (n = 4; *P 0.001).
Cell viability measured with the WST-8 assay of DHT-treated NSC97Q cells transfected with a mock plasmid or the Hes5 vector (n = 3).
Purified RPA and Rtt105-RPA complex were resolved on 15% SDS-PAGE gels and analyzed by silver staining.
(C) Determination of the interaction of MBD9 with HTA9, HTA11, PIE1, CHR11, and CHR17 by co-immunoprecipitation.
A: Primary rat hippocampal neurons were transduced with constructs expressing GFP-tagged TDP-25 (amino acids 220-414 of TDP-43) on day 5 in vitro and cultured for 8 additional days (DIV5+8). Wild-type or variant containing 8 ALS-causing mutations. Immunofluorescence shows disease-related phosphorylation at serine 403/404. Counterstain to label the neuronal cytoskeleton (MAP2) and nuclei (DAPI).
(G,H) SAGA and TFIID were tandem affinity-purified using endogenously TAP-tagged Spt7 (G) from strains containing FLAG-tagged Taf12, grown in rich medium or starved for 45 minutes. TAP-tagged Spt7 and Taf4 were eluted using the TEV protease, releasing a shorter form of each bait (Spt7-CBP or Taf4-CBP). Eluates were loaded and immuno-blotted (IB) using anti-FLAG or anti-CBP antibodies, together with 1% of either whole cell extracts (WCE) or IgG-sepharose flow-through (FT). Shown are IBs that are representative of 2 independent experiments.
B. Parametric summaries of representative sigmoidal activation and inhibition timecourses and impulse (double sigmoid) modeling of transitory inhibition. Sigmoids are summarized by half-max time (trise) and the asymptote (vinter), while impulses include a second half-max (tfall) time and final asymptote (vfinal). The strongest supported model for each timecourse is shown as a filled in line, while the alternative model is shown with a dashed line.
MARCH2+/+ (n = 8) or MARCH2-/- (n = 8) mice were intraperitoneally challenged with 1 × 106 CFU of Listeria monocytogenes. body weight changes for each group (E) are shown.
(c-d) Top: IBs for p62 and actin in control and p62 knockdown (p62 KD #325 or #788) SY5Y cells (c); and in control and p62 KO MEFs (d). Bottom: autophagic clearance of GFP-ANK1-p38 Agm in the same cells in response to serum removal.
(e, f) Survival of Drosophila during ageing without (control) and with supplementation of food at various concentrations of spermidine (as indicated). Autophagy-deficient flies (f) homozygous mutant for Atg7 (Δatg7) were compared with flies capable of autophagy (e) and heterozygous for Atg7 (control). For details of strains and additional wild-type controls, see Supplementary Information Methods and Fig. S6h, i.
F In vivo growth of Empty vector and TINCR cDNA expressing MM13 cells following intradermal injection into NSG mice. Four animals (200,000 cells/mouse) per group were used in each experiment. Tumor growth (mean±sd) was monitored weekly and tumor volume (mean±sd) measured at 4 weeks for primary tumors (originated in the site of injection, left panel) and at 8 weeks for spontaneous metastases which colonized lymph nodes (right panel). The results of two independent experiments are shown. Statistical analyses of control and TINCR-overexpressing groups from both experiments were performed using Student t -test (**: p<0.01).
Glycolytic flux of wildtype (WT) and TM6 strains strongly correlates with fructose-1,6-bisphosphate (FBP) concentration. The glycolytic flux is reported here as the flux between the metabolites fructose 6-phosphate (F6P) and FBP. Glycolytic fluxes were obtained on the basis of physiological and metabolome data, and via a novel method to estimate intracellular fluxes (Niebel et al, 2019). While on high glucose, the WT strain accomplishes a high glucose uptake rate (and thus glycolytic flux), the mutant strain (TM6) only generates a low glucose uptake (and thus glycolytic flux). On maltose, also the mutant strain achieves a high glycolytic flux, since maltose is transported by a separate transporter (Chang et al, 1989). To assess the linear correlation between the FBP concentration and the glycolytic flux across the studied conditions, we implemented Pearson's correlation analysis assisted by bootstrapping. Specifically, we used in total 53 FBP concentration measurements corresponding to six different metabolic conditions (combinations of strains and carbon sources), biological and technical replicates. We paired each of these FBP measurements with the mean and standard deviation of the model-derived glycolytic flux (A) in the corresponding metabolic condition. We assumed the normal distribution of the flux and growth rate with the given mean and standard deviation in every condition, and implemented ordinary nonparametric bootstrapping with 100,000 iterations by randomly sampling values with replacement from the 53 FBP measurements and flux or growth-rate distributions to calculate the correlation statistics.
(F) Stills of live cell imaging to follow de novo CENP-AEA reloading in RPE-1 cells harboring endogenously tagged CENP-Bmcherry. Images were taken every 15 min. White dashed circles contour nuclei prior/after mitosis and cells during mitosis based on bright field images.
H LAPTM4B partially colocalizes with PIPKIγi5. MDA‐MB‐231 cells expressing Myc‐tagged PIPKIγi5 were stained with LAPTM4B anti‐sera (red), anti‐Myc (green), and DAPI. Box is selected region for magnified view. Scale bar: 10 μm.
C Quantification of total numbers of generated cell types (BrdU+; rNSCs, IPCs + neuroblasts, astrocytes) per area in the DGs of 8 week-, 14 month-, and 21 month-old hGFAPeGFP animals. rNSCs: *** p = 0.00076, ** p = 0.0014; IPCs/neuroblasts **** p < 0.0001; astrocytes: *** p = 0.0008; * p = 0.0403; one-way ANOVA with Tukey's post hoc test. D Quantification of the percentages of generated cell types out of all generated cells (rNSCs, IPCs + neuroblasts, astrocytes) in the DG of 8 week-, 14 month-, and 21 month-old animals. rNSCs: not significant; IPCs/neuroblasts **** p < 0.0001, *** p = 0.0001; astrocytes: ** p < 0.0019, **** p < 0.0001; one-way ANOVA with Tukey's post hoc test. Data information: All data are represented as mean ± SEM; number of experimental animals (indicated by red dots): (C,D) n = 5 in 8 week-old mice, n = 6 in 14 month-old mice, n = 5 in 21 month-old mice
C. Filmstrip showing distribution of OCT4 to daughter cells during cell division.
g, PDAC-free survival of autophagy-proficient (blue) and Atg5-deficient (red) KrasG12D/+ mice. d, g, Median survival ± s.d. and male/female (m/f) ratio are provided. 'n/a', no animals succumbed to PDAC. A log-rank test (Mantel-Cox) was used for statistics. a, f, Scale bars: 500 µm (a), 50 µm (f). *P  0.05, **P  0.01; NS, not significant.
B) Volcano plot showing log2 fold change of the number of endogenous siRNAs targeting each C. elegans gene in zsp-1(gg682) vs WT animals on x-axis and log10 (adjusted p-value) of changes on the y-axis. Dots in red indicate siRNAs pools targeting this gene were significantly different in zsp-1(gg682) and WT animals. Dashed lines indicate log2(fold change)=-1 or 1. Adjusted P-value was calculated using Benjamini Hochberg method in DEseq2.
(E) Wild-type (control) and GRAMD1 TKO (TKO) HeLa cells that stably expressed either EGFP or EGFP-tagged GRAMD1b constructs as indicated [wild-type (WT), R189W mutant (R189W), R189W/R191A mutant (RW/RA)], were cultured in the medium supplemented with 10% lipoprotein-deficient serum (LPDS) and mevastatin (50 µM) for 16 hours and then treated with SMase (100 mU/ml) for 3 hours at 37°C. Top: Lysates of the cells were processed for SDS-PAGE and IB with anti-SREBP-2 and anti-actin antibodies. Arrows indicate precursor (P) and cleaved (C) forms of SREBP-2. Bottom: The response rate was obtained by normalizing the ratio of the band intensity of the cleaved SREBP-2 over the total band intensity of cleaved and precursor forms of SREBP-2 from the cells with SMase treatment by the one from the cells without SMase treatment for each condition. Note that the suppression of SREBP-2 cleavage is attenuated in GRAMD1 TKO HeLa cells compared to wild-type control HeLa cells. Note also the rescue by expression of wild-type EGFP-GRAMD1b (WT) but not by mutant versions of EGFP-GRAMD1b [R189W, R189W/R191A (RW/RA)] [mean ± SEM, n = 4 lysates (independent experiments) for each condition; Dunnett's multiple comparisons test, **P = 0.0035 (Control + EGFP vs. TKO + EGFP), **P < 0.0001 (TKO + EGFP vs. TKO + WT), n.s. denotes not significant].
(E) Statistical analysis of IMA ratio at 4 weeks and 8 weeks post Ad-Cre treatment (n=5 per group). Results were shown as mean ± SEM.
(A) A cartoon representation of a crystal structure of the COP1 WD40 domain (grey) in complex with the TRIB1 tail (cyan) (PDB: 5IGQ). Point mutations introduced in the WD40 domain are shown in red.
J, hIGFREO had similar whole-body and femur length as WT (n=7-9 mice per group).
(b-e) Normalized ratio between luciferase activity bound to beads and present in lysates. The indicated purified GST fusion proteins coupled to beads were incubated with lysates of 293ET cells expressing Nap1, Sintbad or Tank, each fused to luciferase (b)
D Heatmap shows expression of putative LRH-1 target genes in this study's transcriptome profiles. Colors indicate log2 fold change of marker genes in the naïve versus primed hPSCs, with a range from dark blue for lower expression to dark red for higher expression. E Heatmap shows expression of putative RARγ target genes in this study's transcriptome profiles. Colors indicate log2 fold change of marker genes in the naïve versus primed hPSCs, with a range from dark blue for lower expression to dark red for higher expression.
MARCH2+/+ HEK293T cells harboring empty vector or MARCH2-/- HEK293T cells harboring empty vector, Flag-tagged MARCH2, or MARCH2CCH (a mutant harboring mutations in residues within the catalytic domain: C64S, C67S, and H90Q) were infected with VSV-GFP (MOI = 0.05). Viral replication was assessed under a fluorescence microscope (H)
(B) Western blot analysis of RC subunits in MEFs upon 48 hours treatment with actinonin, doxycycline and chloramphenicol (+ antibiotic), followed by a 48 hours recovery period (- antibiotic). During both periods, proteins are isolated at 12h, 24h and 48h. Antibodies against subunits of C I (NDUFB6), C III (UQCRC1), C IV (COXI), MRPS35 and MRPL37 were used as indicated.
D LM2-4luc produced the highest levels of VEGF and IL-8, but lower levels of IL-6, compared to other TC types; ***P < 0.001 and *P < 0.05 (unpaired t-tests).
d. Box plots display the distribution of the cell membrane derived PIN2-GFP fluorescence intensity (FI) values (in arbitrary units, a.u.) in roots transferred to ammonium ((grey, epidermis (ep) and red, cortex (co) and to nitrate (blue, epidermis (ep) and green, cortex (co)). 5 cells per roots were analyzed in at least 9 roots per genotype per treatment. The statistical significance was evaluated with ANOVA at p<0.05. The box chart components are defined as; box (25%-75%), central band (median line), central box (mean) and the range is within 1.5IQR.
B, C At day 0 (B, before tumor injection) or at day 26 (C, after tumor injection), surface PD-1 expression in CD44hi CD8+ or CD44hi CD4+T cells, and the percentages of CD44hi CD8+T cells or Foxp3+ CD4+ cells were analyzed (mean ± SD, n ≥ 5 mice).
G. PEBP1 gene deletion after Lv-PEBP1-sg1 knockout in the primary CD4+ T cells. The PCR products of PEBP1 were cloned and then sequenced. PEBP1-sg1 target gene sequences are shown in red letters. Dashes indicate deleted bases relative to the wild-type sequence.
PMA-treated THP-1 shSAMHD1 cells transduced for 24 h with HIV-1 GFP ΔCA-SP1 viruses (0.1 U RT/ml red line, 0.5 U RT/ml blue line). E) CXCL-10 protein in supernatants from (C, D) (ELISA).
F. WT MEFs were untreated, treated with 10 μg/ml FAC for 12h or treated with 10 μg/ml FAC, 10 μM E64d and 10 μg/ml pepstatin A (pep A) for 12h. The cells were immunostained with anti-NCOA4 and anti-LAMP1. Scale bar, 10 μm. Quantitative data are shown as the means ± SD of three biological replicates. At least 30 cells were quantified in each replicate. P=0.033 (Welch Two Sample t-test).
(C) Old APPPS1 tissue was incubated with conditioned media collected from cultured WT primary microglial cells (old alone + MG CM, grey bar) or with unconditioned slice culture media as a control (old alone + Ctr, white bar). Quantitative analysis of core-only plaques at 14 DIV reveals enhanced amyloid plaque clearance upon incubation of the old APPPS1 tissue with conditioned media collected from cultured primary microglia. The values are expressed as percentages of core-only plaques from the total number of amyloid plaques and represent mean ± SEM from 3 independent experiments, each experiment including 2 independent slice culture dishes (*P < 0.05).
H&amp;amp;E staining shows neovascular tufts (arrows). Representative of five retinae per group in two independent experiments. Scale bar, 50 μm.
D Immunostaining of HeLa Kyoto cells stably expressing PCDH7‐LAP‐tagged BAC transgenes (green) probed for tubulin (red) and DNA (blue). Top panel shows mitotic cell those were treated with 5 μM STC for 10 h, and bottom panel shows interphase cells. Scale bar, 10 μm.
A Cytokine release of 3 days differentiated THP-1 cells of indicated genotypes upon TLR2 blockade (T2.5) and challenge with TLR ligands, heat inactivated (hi) or viable bacteria.
C FACS plots showing DNA content on the x-axis (propidium iodide) of HeLa cells with stable expression of wild-type or KEN-mutant E2F7, after 24 hours of doxycycline. Thresholds for EGFP positivity were set by applying the same gate to vehicle- and doxycycline treated cells.
A. Individual hESCs have the potential to generate another stem cell through self-renewal or to differentiate into a more lineage-specific cell type.
(B) Autophagy in Foxg1 Cre MCL‐1Δ/Δ mice. Brain sections from E15.5 animals with the indicated genotypes were stained for LC3 (green). Confocal images were taken using × 20 (bottom panel) or × 63 (top and middle panels) objectives. Scale bars=50 μm.
(A) Larval NSCs of control (ctrl; yw) and pr-set7716/Df(3R) ED5644 (716/Df) at 0h, 24h and 48h ALH were stained for Pr-set7, Dpn and Mira. Yellow dotted boxes, zoomed-in area. Data information: White arrows point to NSCs. Scale bars 15 μm (A).
(a) Survival rate of wild-type mice (WT) and indicated knockout mice after intranasal infection with 750 plaque forming units (PFU) of the PR8 virus.