RESUMEN
Glioblastoma multiforme (GBM) is a primary type of lethal brain tumor. Over the last two decades, temozolomide (TMZ) has remained the primary chemotherapy for GBM. However, TMZ resistance in GBM constitutes an underlying factor contributing to high rates of mortality. Despite intense efforts to understand the mechanisms of therapeutic resistance, there is currently a poor understanding of the molecular processes of drug resistance. For TMZ, several mechanisms linked to therapeutic resistance have been proposed. In the past decade, significant progress in the field of mass spectrometry-based proteomics has been made. This review article discusses the molecular drivers of GBM, within the context of TMZ resistance with a particular emphasis on the potential benefits and insights of using global proteomic techniques.
RESUMEN
A high-throughput sample preparation protocol based on the use of 96-well molecular weight cutoff (MWCO) filter plates was developed for shotgun proteomics of cell lysates. All sample preparation steps, including cell lysis, buffer exchange, protein denaturation, reduction, alkylation and proteolytic digestion are performed in a 96-well plate format, making the platform extremely well suited for processing large numbers of samples and directly compatible with functional assays for cellular proteomics. In addition, the usage of a single plate for all sample preparation steps following cell lysis reduces potential samples losses and allows for automation. The MWCO filter also enables sample concentration, thereby increasing the overall sensitivity, and implementation of washing steps involving organic solvents, for example, to remove cell membranes constituents. The optimized protocol allowed for higher throughput with improved sensitivity in terms of the number of identified cellular proteins when compared to an established protocol employing gel-filtration columns.
Asunto(s)
Métodos Analíticos de la Preparación de la Muestra/instrumentación , Células/metabolismo , Ensayos Analíticos de Alto Rendimiento/instrumentación , Ensayos Analíticos de Alto Rendimiento/métodos , Proteómica/instrumentación , Proteómica/métodos , Extractos Celulares , Cromatografía Liquida , Células HEK293 , Humanos , Espectrometría de Masas , Nanotecnología , Péptidos/metabolismo , Proteínas/metabolismoRESUMEN
The cytosolic ribosomal proteome of Arabidopsis thaliana has been studied intensively by a range of proteomics approaches and is now one of the most well characterized eukaryotic ribosomal proteomes. Plant cytosolic ribosomes are distinguished from other eukaryotic ribosomes by unique proteins, unique post-translational modifications and an abundance of ribosomal proteins for which multiple divergent paralogs are expressed and incorporated. Study of the A. thaliana ribosome has now progressed well beyond a simple cataloging of protein parts and is focused strongly on elucidating the functions of specific ribosomal proteins, their paralogous isoforms and covalent modifications. This review summarises current knowledge concerning the Arabidopsis cytosolic ribosomal proteome and highlights potentially fruitful areas of future research in this fast moving and important area.