The CRISPR-Cas System Is Involved in OmpR Genetic Regulation for Outer Membrane Protein Synthesis in Salmonella Typhi.

The CRISPR-Cas cluster is found in many prokaryotic genomes including those of the Enterobacteriaceae family. Salmonella enterica serovar Typhi (S. Typhi) harbors a Type I-E CRISPR-Cas locus composed of cas3, cse1, cse2, cas7, cas5, cas6e, cas1, cas2, and a CRISPR1 array. In this work, it was determined that, in the absence of cas5 or cas2, the amount of the OmpC porin decreased substantially, whereas in individual cse2, cas6e, cas1, or cas3 null mutants, the OmpF porin was not observed in an electrophoretic profile of outer membrane proteins. Furthermore, the LysR-type transcriptional regulator LeuO was unable to positively regulate the expression of the quiescent OmpS2 porin, in individual S. Typhi cse2, cas5, cas6e, cas1, cas2, and cas3 mutants. Remarkably, the expression of the master porin regulator OmpR was dependent on the Cse2, Cas5, Cas6e, Cas1, Cas2, and Cas3 proteins. Therefore, the data suggest that the CRISPR-Cas system acts hierarchically on OmpR to control the synthesis of outer membrane proteins in S. Typhi.

Principal component analysis on LC­MS/MS and 2DE­MALDI­TOF in glioblastoma cell lines reveals that mitochondria act as organelle sensors of the metabolic state in glioblastoma.

Glioblastoma is a difficult disease to diagnose. Proteomic techniques are commonly applied in biomedical research, and can be useful for early detection, making an accurate diagnosis and reducing mortality. The relevance of mitochondria in brain development and function is well known; therefore, mitochondria may influence the development of glioblastoma. The T98G (with oxidative metabolism) and U87MG (with glycolytic metabolism) cell lines are considered to be useful glioblastoma models for studying these tumors and the role of mitochondria in key aspects of this disease, such as prognosis, metastasis and apoptosis. In the present study, principal component analysis of protein abundance data identified by liquid chromatography coupled to tandem mass spectrometry (LC­MS/MS) and matrix­assisted laser desorption/ionization­time of flight mass spectrometry (MALDI­TOF) from 2D gels indicated that representative mitochondrial proteins were associated with glioblastoma. The selected proteins were organized into T98G­ and U87MG­specific protein­protein interaction networks to demonstrate the representativeness of both proteomic techniques. Gene Ontology overrepresentation analysis based on the relevant proteins revealed that mitochondrial processes were associated with metabolic changes, invasion and metastasis in glioblastoma, along with other non­mitochondrial processes, such as DNA translation, chaperone responses and autophagy. Despite the lower resolution of 2D electrophoresis, principal component analysis yielded information of comparable quality to that of LC­MS/MS. The present analysis pipeline described a specific and more complete metabolic status for each cell line, defined a clear mitochondrial performance for distinct glioblastoma tumors, and introduced a useful strategy to understand the heterogeneity of glioblastoma.

A proteomic approach of pediatric astrocytomas: MiRNAs and network insight.

Pediatric astrocytomas, a leading cause of death associated with cancer, are the most common primary central nervous system tumors found in children. Most studies of these tumors focus on adults, not on children. We examined the global protein and microRNA expression pattern by 2D SDS-PAGE, mass spectrometry (MALDI-TOF), and RT(2) miRNA PCR Array System. Proteomic studies revealed 49 proteins with changes on the expression. Interactome showed that vimentin, calreticulin, and 14-3-3 epsilon protein are hub proteins in these neoplasms. MicroRNA analyses demonstrated for the first time novel microRNAs involved in the astrocytoma biology. In conclusion, our results show that novel proteins and microRNAs with expression changes on pediatric astrocytoma could serve as biomarkers of tumor progression. BIOLOGICAL SIGNIFICANCE: Astrocytomas are tumors that progress rapidly and that invade surrounding tissues. Although some drugs have been developed to treat these neoplasms, the mortality of patients is still very high. In this study, we describe for the first time, to our knowledge, some proteins and miRNAs associated with the biology of astrocytic tumors that could be postulated as possible diagnostic or prognostic biomarkers. Altogether, our results indicate that large-scale analyses allow making a fairly accurate prediction of different cellular processes altered in astrocytic tumors.