The structure of SAV1646 from Staphylococcus aureus belonging to a new `ribosome-associated' subfamily of bacterial proteins.

The crystal structure of the SAV1646 protein from the pathogenic microorganism Staphylococcus aureus has been determined at 1.7 Šresolution. The 106-amino-acid protein forms a two-layer sandwich with α/ß topology. The protein molecules associate as dimers in the crystal and in solution, with the monomers related by a pseudo-twofold rotation axis. A sequence-homology search identified the protein as a member of a new subfamily of yet uncharacterized bacterial `ribosome-associated' proteins with at least 13 members to date. A detailed analysis of the crystal protein structure along with the genomic structure of the operon containing the sav1646 gene allowed a tentative functional model of this protein to be proposed. The SAV1646 dimer is assumed to form a complex with ribosomal proteins L21 and L27 which could help to complete the assembly of the large subunit of the ribosome.

Markers of survival and metastatic potential in childhood CNS primitive neuro-ectodermal brain tumours: an integrative genomic analysis.

BACKGROUND: Childhood CNS primitive neuro-ectodermal brain tumours (PNETs) are very aggressive brain tumours for which the molecular features and best treatment approaches are unknown. We assessed a large cohort of these rare tumours to identify molecular markers to enhance clinical management of this disease. METHODS: We obtained 142 primary hemispheric CNS PNET samples from 20 institutions in nine countries and examined transcriptional profiles for a subset of 51 samples and copy number profiles for a subset of 77 samples. We used clustering, gene, and pathway enrichment analyses to identify tumour subgroups and group-specific molecular markers, and applied immunohistochemical and gene-expression analyses to validate and assess the clinical significance of the subgroup markers. FINDINGS: We identified three molecular subgroups of CNS PNETs that were distinguished by primitive neural (group 1), oligoneural (group 2), and mesenchymal lineage (group 3) gene-expression signatures with differential expression of cell-lineage markers LIN28 and OLIG2. Patients with group 1 tumours were most often female (male:female ratio 0·61 for group 1 vs 1·25 for group 2 and 1·63 for group 3; p=0·043 [group 1 vs groups 2 and 3]), youngest (median age at diagnosis 2·9 years [95% CI 2·4-5·2] for group 1 vs 7·9 years [6·0-9·7] for group 2 and 5·9 years [4·9-7·8] for group 3; p=0·005), and had poorest survival (median survival 0·8 years [95% CI 0·5-1·2] in group 1, 1·8 years [1·4-2·3] in group 2 and 4·3 years [0·8-7·8] in group 3; p=0·019). Patients with group 3 tumours had the highest incidence of metastases at diagnosis (no distant metastasis:metastasis ratio 0·90 for group 3 vs 2·80 for group 1 and 5·67 for group 2; p=0·037). INTERPRETATION: LIN28 and OLIG2 are promising diagnostic and prognostic molecular markers for CNS PNET that warrant further assessment in prospective clinical trials. FUNDING: Canadian Institute of Health Research, Brainchild/SickKids Foundation, and the Samantha Dickson Brain Tumour Trust.

Thrombospondin-1 mimetics are promising novel therapeutics for MYC-associated medulloblastoma.

BACKGROUND: Medulloblastoma (MB) comprises four subtypes of which group 3 MB are the most aggressive. Although overall survival for MB has improved, the outcome of group 3 MB remains dismal. C-MYC (MYC) amplification or MYC overexpression which characterizes group 3 MB is a strong negative prognostic factor and is frequently associated with metastases and relapses. We previously reported that MYC expression alone promotes highly aggressive MB phenotypes, in part via repression of thrombospondin-1 (TSP-1), a potent tumor suppressor. METHODS: In this study, we examined the potential role of TSP-1 and TSP-1 peptidomimetic ABT-898 in MYC-amplified human MB cell lines and two distinct murine models of MYC-driven group 3 MBs. RESULTS: We found that TSP-1 reconstitution diminished metastases and prolonged survival in orthotopic xenografts and promoted chemo- and radio-sensitivity via AKT signaling. Furthermore, we demonstrate that ABT-898 can recapitulate the effects of TSP-1 expression in MB cells in vitro and specifically induced apoptosis in murine group 3 MB tumor cells. CONCLUSION: Our data underscore the importance of TSP-1 as a critical tumor suppressor in MB and highlight TSP-1 peptidomimetics as promising novel therapeutics for the most lethal subtype of MB.

Crystal structure of Staphylococcus aureus Zn-glyoxalase I: new subfamily of glyoxalase I family.

The crystal structures of protein SA0856 from Staphylococcus aureus in its apo-form and in complex with a Zn2+-ion have been presented. The 152 amino acid protein consists of two similar domains with α + ß topology. In both crystalline state and in solution, the protein forms a dimer with monomers related by a twofold pseudo-symmetry rotation axis. A sequence homology search identified the protein as a member of the structural family Glyoxalase I. We have shown that the enzyme possesses glyoxalase I activity in the presence of Zn2+, Mg2+, Ni2+, and Co2+, in this order of preference. Sequence and structure comparisons revealed that human glyoxalase I should be assigned to a subfamily A, while S. aureus glyoxalase I represents a new subfamily B, which includes also proteins from other bacteria. Both subfamilies have a similar protein chain fold but rather diverse sequences. The active sites of human and staphylococcus glyoxalases I are also different: the former contains one Zn-ion per chain; the latter incorporates two of these ions. In the active site of SA0856, the first Zn-ion is well coordinated by His58, Glu60 from basic molecule and Glu40*, His44* from adjacent symmetry-related molecule. The second Zn3-ion is coordinated only by residue His143 from protein molecule and one acetate ion. We suggest that only single Zn1-ion plays the role of catalytic center. The newly found differences between the two subfamilies could guide the design of new drugs against S. aureus, an important pathogenic micro-organism.

Apoptosis of viral-infected airway epithelial cells limit viral production and is altered by corticosteroid exposure.

BACKGROUND: Effects of respiratory viral infection on airway epithelium include airway hyper-responsiveness and inflammation. Both features may contribute to the development of asthma. Excessive damage and loss of epithelial cells are characteristic in asthma and may result from viral infection. OBJECTIVE: To investigate apoptosis in Adenoviral-infected Guinea pigs and determine the role of death receptor and ligand expression in the airway epithelial response to limit viral infection. METHODS: Animal models included both an Acute and a Chronic Adeno-infection with ovalbumin-induced airway inflammation with/without corticosteroid treatment. Isolated airway epithelial cells were cultured to study viral production after infection under similar conditions. Immunohistochemistry, western blots and viral DNA detection were used to assess apoptosis, death receptor and TRAIL expression and viral release. RESULTS: In vivo and in vitro Adeno-infection demonstrated different apoptotic and death receptors (DR) 4 and 5 expression in response to corticosteroid exposure. In the Acute Adeno-infection model, apoptosis and DR4/5 expression was coordinated and were time-dependent. However, in vitro Acute viral infection in the presence of corticosteroids demonstrated delayed apoptosis and prolonged viral particle production. This reduction in apoptosis in Adeno-infected epithelial cells by corticosteroids exposure induced a prolonged virus production via both DR4 and TRAIL protein suppression. In the Chronic model where animals were ovalbumin-sensitized/challenged and were treated with corticosteroids, apoptosis was reduced relative to adenovirus-infected or corticosteroid alone. CONCLUSION: Our data suggests that apoptosis of infected cells limits viral production and may be mediated by DR4/5 and TRAIL expression. In the Acute model of Adeno-infection, corticosteroid exposure may prolong viral particle production by altering this apoptotic response of the infected cells. This results from decreased DR4 and TRAIL expression. In the Chronic model treated with corticosteroids, a similar decreased apoptosis was observed. This data suggests that DR and TRAIL modulation by corticosteroids may be important in viral infection of airway epithelium. The prolonged virus release in the setting of corticosteroids may result from reduced apoptosis and suppressed DR4/TRAIL expression by the infected cells.

Integrated (epi)-Genomic Analyses Identify Subgroup-Specific Therapeutic Targets in CNS Rhabdoid Tumors.

We recently reported that atypical teratoid rhabdoid tumors (ATRTs) comprise at least two transcriptional subtypes with different clinical outcomes; however, the mechanisms underlying therapeutic heterogeneity remained unclear. In this study, we analyzed 191 primary ATRTs and 10 ATRT cell lines to define the genomic and epigenomic landscape of ATRTs and identify subgroup-specific therapeutic targets. We found ATRTs segregated into three epigenetic subgroups with distinct genomic profiles, SMARCB1 genotypes, and chromatin landscape that correlated with differential cellular responses to a panel of signaling and epigenetic inhibitors. Significantly, we discovered that differential methylation of a PDGFRB-associated enhancer confers specific sensitivity of group 2 ATRT cells to dasatinib and nilotinib, and suggest that these are promising therapies for this highly lethal ATRT subtype.

Carbon nanotube compared with carbon black: effects on bacterial survival against grazing by ciliates and antimicrobial treatments.

The ingestion and digestion of Escherichia coli by the ciliated protozoan, Tetrahymena thermophila, was investigated after an initial exposure to either water-soluble single-walled carbon nanotubes (SWNT) or to carbon black (CB). Both SWNT and CB were internalised and visible in food vacuoles of ciliates. When presented with E. coli expressing green-fluorescent protein (GFP), these ciliates internalised bacteria as well. However, ciliates that had first internalised SWNT but not CB subsequently externalised or egested vesicle-like structures with fluorescent bacteria inside. These egested bacteria were viable and less susceptible than planktonic E. coli to killing either by the antibiotic, chloramphenicol or the disinfectant, glutaraldehyde. These results suggest that SWNT can alter the intracellular trafficking of vesicles within ciliates, leading to bacterial prey being packaged externally and protected for a time from environmental killing, which could have implications for sewage treatment and for public health.