Journal: Cell reports

Article Title: Viral targeting of glioblastoma stem cells with patient-specific genetic and post-translational p53 deregulations.

doi: 10.1016/j.celrep.2021.109673

Figure Lengend Snippet: Figure 1. GSC heterogeneous permissiveness to parvovirus cytotoxic infection (A) Hierarchical clustering of the GSCs, NSCs, and GBM cell lines arising from a global genome-expression array profile. (B) Heatmap of the global gene expression array showing significant change (3-fold and higher) in the level of expression of specific genes with respect to that found in NSCs. (C) Confocal IF analysis of the expression and subcellular distribution of CD133 (red) and of the NS1 protein (green) in MVMi infected neurospheres at 3 dpi; scale bar, 20 mm. (D) 3D reconstruction of the MVMp-induced damage on neurospheres’ morphology stained for the cytoplasmic nestin (green) and the nuclear NS1 (red) proteins; scale bar, 50 mm. (E) Representative experiment of GSC growth kinetic in culture, with approximate doubling times of 120 h (GS7), 80 h (GS5), and 62 h (GS8 and GS9). (F) Cell viability quantification along with infectious virus production over time in GSC cultures infected by the MVMp and MVMi strains. Representative results from three independent experiments are shown. (G) Suppression of GSC self-renewal by MVM infection measured as clonogenic potential. Colony-forming unit (CFU) values are the mean with standard errors from three experiments. *p < 0.05; **p < 0.01; ***p < 0.001, unpaired two-tailed t tests. (H) Temporal blot-analysis of NS1 protein accumulation in MVMp-infected GSCs and U373MG cultures. C, uninfected cell controls. (I) NS1 expression correlates with cell-cycle arrest at S/G2 in GSCs and in human cell lines fully permissive (NB324K) or restrictive (U87MG) to MVMi genome replication. Data of the panels were confirmed from at least two independent experiments.

Article Snippet: GBM and other established cell lines The U373MG and U87MG (purchased from the American Type Culture Collection: ATCC code HTB-17 and HTB-14, respectively), and the U251MG established human glioblastoma cell lines, were cultured in Dulbecco’s modified Eagle medium (DMEM) supplemented with 10% heat-inactivated fetal calf serum (FCS; GIBCO).

Techniques: Infection, Expressing, Gene Expression, Staining, Virus, Two Tailed Test

Journal: Cell reports

Article Title: Viral targeting of glioblastoma stem cells with patient-specific genetic and post-translational p53 deregulations.

doi: 10.1016/j.celrep.2021.109673

Figure Lengend Snippet: Figure 2. GSCs unbalance parvovirus genome expression and mis-localize capsid assembly (A) Flow cytometry of MVM capsid formation and NS1 expression in GSCs (upper) and control human cell lines (lower). Colored key boxes in the middle of the figure also apply to (D). (B) Western blot time course analysis (dpi) of VP accumulation in MVMp-infected GSCs. The VP species are marked on the right margin by respective arrows. (C) Production of assembled capsids. Cells were infected by the purified MVM strains at moi 10 pfu/cell, and assembled capsids bound to cells (0 hpi) and produced at 1 dpi (human cell lines) or at 3 dpi (GSCs) were measured in cell homogenates as hemagglutination units (HAUs) with mouse erythrocytes. Results are the mean of three experiments with standard errors. (D) Flow cytometry of MVM assembled capsid (B7-Mab) and mature DNA-filled virus (anti-2Nt antibody) production in control human U87MG and NB3243K cells (1 dpi) and GSC neurospheres (3 dpi). Representative results of at least three experiments are shown. (E) Subcellular distribution of NS1 and MVM assembled capsids (B7-Mab) visualized by IF-confocal staining in control human NB3243K and U87MG cells (1 dpi) and in GSC neurospheres (3 dpi). Phenotypes were classified in three categories: nuclear (N > C), mixed (N = C), and cytoplasmic (N < C). Scale bars, 50 mm.

Article Snippet: GBM and other established cell lines The U373MG and U87MG (purchased from the American Type Culture Collection: ATCC code HTB-17 and HTB-14, respectively), and the U251MG established human glioblastoma cell lines, were cultured in Dulbecco’s modified Eagle medium (DMEM) supplemented with 10% heat-inactivated fetal calf serum (FCS; GIBCO).

Techniques: Expressing, Flow Cytometry, Control, Western Blot, Infection, Produced, Virus, Staining

Journal: BMC Bioinformatics

Article Title: Structure and sequence analyses of Bacteroides proteins BVU_4064 and BF1687 reveal presence of two novel predominantly-beta domains, predicted to be involved in lipid and cell surface interactions

doi: 10.1186/s12859-014-0434-7

Figure Lengend Snippet: Structures of the N-terminally truncated Bacteroides proteins BVU_4064 and BF1687 (PDB codes 3kog and 3g3l, respectively). The N-terminal domain (in slate blue color) and the C-terminal domain (in orange color) of the 3kog structure show significant similarities with the corresponding domains of 3g3l structure (N and C terminal domains shown in pale cyan and wheat colors respectively). In contrast, the region connecting the domains (in green) is clearly different in the two structures: a short linker in 3kog, an extended 4-helix insertion and one extra strand that is added to the C-terminal domain in 3g3l. A histidine-rich region present at the C-terminus in both of our proteins is found ordered only in the 3kog structure (see box with text in the Figure).

Article Snippet: The crystal structures of the N-terminally truncated Bacteroides proteins BVU_4064 ( Bacteroides vulgatus strain ATCC 8482, JCSG target ID: 393242, GenBank accession: YP_001301288.1, PDB code: 3kog) and BF1687 ( Bacteroides fragilis strain NCTC 9343, JCSG target ID: 393243, Gene Bank accession: YP_211325.1, PDB code: 3g3l) have been determined to 1.85 Å and 2.2 Å resolution, using MAD and SAD phasing methods respectively as described in the section.

Techniques:

Journal: BMC Bioinformatics

Article Title: Structure and sequence analyses of Bacteroides proteins BVU_4064 and BF1687 reveal presence of two novel predominantly-beta domains, predicted to be involved in lipid and cell surface interactions

doi: 10.1186/s12859-014-0434-7

Figure Lengend Snippet: Superposition of 3kog and 3g3l structures. (A) Corresponding domains (colored in slate blue and pale cyan for N-terminal domains; orange and wheat for C-terminal domains; linker region in green) in the two structures superimpose fairly well with an overall RMSD of 3.7 Å for the 166 equivalent positions in the rigid-body alignment . (B) Stereo view of N and C terminal domains shown separately with linker regions removed to highlight the structural similarity.

Article Snippet: The crystal structures of the N-terminally truncated Bacteroides proteins BVU_4064 ( Bacteroides vulgatus strain ATCC 8482, JCSG target ID: 393242, GenBank accession: YP_001301288.1, PDB code: 3kog) and BF1687 ( Bacteroides fragilis strain NCTC 9343, JCSG target ID: 393243, Gene Bank accession: YP_211325.1, PDB code: 3g3l) have been determined to 1.85 Å and 2.2 Å resolution, using MAD and SAD phasing methods respectively as described in the section.

Techniques:

Journal: BMC Bioinformatics

Article Title: Structure and sequence analyses of Bacteroides proteins BVU_4064 and BF1687 reveal presence of two novel predominantly-beta domains, predicted to be involved in lipid and cell surface interactions

doi: 10.1186/s12859-014-0434-7

Figure Lengend Snippet: Structural similarities of the N-terminal domains. (A-F) Pre-albumin-like fold of the N-terminal domains in 3kog and 3g3l structures that is also present as a cell adhesion modules in several proteins belonging to the Transthyretin superfamily. (G) Alignment between the lipoprotein signal sequences present at the N-terminus of BVU_4064 and BF1687. The arrow points to the conserved CYS residue in the consensus sequence for the protein family PF12985.

Article Snippet: The crystal structures of the N-terminally truncated Bacteroides proteins BVU_4064 ( Bacteroides vulgatus strain ATCC 8482, JCSG target ID: 393242, GenBank accession: YP_001301288.1, PDB code: 3kog) and BF1687 ( Bacteroides fragilis strain NCTC 9343, JCSG target ID: 393243, Gene Bank accession: YP_211325.1, PDB code: 3g3l) have been determined to 1.85 Å and 2.2 Å resolution, using MAD and SAD phasing methods respectively as described in the section.

Techniques: Residue, Sequencing

Journal: BMC Bioinformatics

Article Title: Structure and sequence analyses of Bacteroides proteins BVU_4064 and BF1687 reveal presence of two novel predominantly-beta domains, predicted to be involved in lipid and cell surface interactions

doi: 10.1186/s12859-014-0434-7

Figure Lengend Snippet: Structural similarities of the C-terminal domain of 3kog and 3g3l with bacterial pore-forming toxins. The region shown in red is implicated in membrane insertion in the pore-forming toxins [epsilon toxin (PDB code: 1uyj) and aerolysin (PDB code: 1z52)] and in the hemolytic lectin (PDB code 1w3g). In both 3kog and 3g3l this region corresponds to a helical insertion.

Article Snippet: The crystal structures of the N-terminally truncated Bacteroides proteins BVU_4064 ( Bacteroides vulgatus strain ATCC 8482, JCSG target ID: 393242, GenBank accession: YP_001301288.1, PDB code: 3kog) and BF1687 ( Bacteroides fragilis strain NCTC 9343, JCSG target ID: 393243, Gene Bank accession: YP_211325.1, PDB code: 3g3l) have been determined to 1.85 Å and 2.2 Å resolution, using MAD and SAD phasing methods respectively as described in the section.

Techniques: Membrane