Protéome

Résumé

Le protéome est l'ensemble des protéines exprimées dans une cellule, une partie d'une cellule (membranes, organites) ou un groupe de cellules (organe, organisme, groupe d'organismes) dans des conditions données et à un moment donné. Histoire du mot Ce terme d'origine anglo-saxonne a été inventé en 1994 par Mark Wilkins de l'Université Macquarie de Sydney. Il provient de la contraction de protéine et génome car le protéome est aux protéines ce que le génome est pour les gènes. Alors que les gènes codent des instructions, les protéines les exécutent pour assurer le fonctionnement cellulaire. Dynamique Le protéome est de nature dynamique : à la différence du génome qui est plus ou moins stable dans les cellules d'un organisme, le protéome varie temporellement et spatialement, selon les réponses de l’organisme à son environnement ou à des stimuli internes (hormonaux et chronobiologiques par exemple), ou en réponse à certains polluants (métaux lourds par exemple) L

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https://fr.wikipedia.org/wiki/Prot%C3%A9ome

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Multilayered genetic and omics dissection of mitochondrial activity in a mouse reference population

Ruedi Aebersold, Johan Auwerx, Sébastien Dubuis, Virginija Jovaisaite, Adrienne Joëlle Laurence Mottis, Evan Graehl Williams

The manner by which genotype and environment affect complex phenotypes is one of the fundamental questions in biology. In this study, we quantified the transcriptome--a subset of the metabolome--and, using targeted proteomics, quantified a subset of the liver proteome from 40 strains of the BXD mouse genetic reference population on two diverse diets. We discovered dozens of transcript, protein, and metabolite QTLs, several of which linked to metabolic phenotypes. Most prominently, Dhtkd1 was identified as a primary regulator of 2-aminoadipate, explaining variance in fasted glucose and diabetes status in both mice and humans. These integrated molecular profiles also allowed further characterization of complex pathways, particularly the mitochondrial unfolded protein response (UPR(mt)). UPR(mt) shows strikingly variant responses at the transcript and protein level that are remarkably conserved among C. elegans, mice, and humans. Overall, these examples demonstrate the value of an integrated multilayered omics approach to characterize complex metabolic phenotypes.

Cell Press2014

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With the advent of high-throughput and genome-wide screening initiatives, there is a need for improved methods for cell-based assays. Current approaches require expensive equipment, rely on large-scale culturing formats not suited for small or rare sample types, or involve tedious manual handling. Microfluidic systems could provide a solution to these limitations, since these assays are accessible, miniaturized, and automated. When coupled with high-content analysis, microfluidics has the potential to drastically increase throughput in cell biology and drug discovery. In light of these benefits, we developed 3 microfluidic approaches for mammalian cell-based assays: (1) printing of live mammalian cells into nanowell arrays, (2) a high-throughput transfection device, and (3) a module that generates complex, continuous concentration profiles. Our first technique generates high-density nanowell arrays of live mammalian cells (LMCAs) using a standard contact microarrayer. Both commonly used cell lines and primary cells cultured on the arrays are highly viable and maintain their signature phenotype, making the platform suitable for long-term stem cell differentiation studies. Our 675-well array is ~2.6x more dense than a 1,536-well microtiter plate and can be frozen and thawed, facilitating the handling, storage, and screening of large libraries of cells. LMCAs are also compatible with transfection, a technique that could enable analysis of the entire proteome in the natural cellular context. Transfection is routinely conducted on high-throughput arrays, but this setup requires manual cell culturing and precludes precise control over the cell environment. To this end, we created a microfluidic chip that streamlines cell loading and culturing and implements 280 independent transfections at up to 99% efficiency. The chip can perform co-transfections, in which the number of cells expressing each protein and the average protein expression level can be precisely tuned as a function of input DNA concentration. This platform is well-suited for optimizing synthetic gene circuits; we co-transfected four plasmids to test a histidine kinase signaling pathway and mapped the dose dependence of this network on the level of one of its constituents. The chip is readily integrated with high-content imaging, enabling the evaluation of cellular behavior and protein expression dynamics over time. To complement the biological assays that could be performed on our transfection chip, we lastly generated an accurate and automated method to manipulate molecular concentrations on chip. Our pulse-width modulation (PWM)-based microfluidic module combines up to 6 different inputs and produces arbitrary concentrations with a dynamic range of 3-5 decades. We created complex concentration profiles of 2 molecules, with each concentration independently controllable. The PWM module can execute rapid concentration changes as well as long-timescale pharmacokinetic profiles under a variety of operating conditions, making it ideal for integration with existing devices for advanced cell and pharmacokinetic studies. Taken together, the 3 microtechnologies developed in this work integrate and automate mammalian cell handling, culturing, transfection, imaging, and solution preparation. These features have far-reaching implications in fields such as synthetic biology, stem cell research, and drug development.

EPFL2017

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Observing cellular responses to perturbations is central to generating and testing hypotheses in biology. We developed a massively parallel microchemostat array capable of growing and observing 1,152 yeast-GFP strains on the single-cell level with 20 min time resolution. We measured protein abundance and localization changes in 4,085 GFP-tagged strains in response to methyl methanesulfonate and analyzed 576 GFP strains in five additional conditions for a total of more than 10,000 unique experiments, providing a systematic view of the yeast proteome in flux. We observed that processing bodies formed rapidly and synchronously in response to UV irradiation, and in conjunction with 506 deletion-GFP strains, identified four gene disruptions leading to abnormal ribonucleotide-diphosphate reductase (Rnr4) localization. Our microchemostat platform enables the large-scale interrogation of proteomes in flux and permits the concurrent observation of protein abundance, localization, cell size, and growth parameters on the single-cell level for thousands of microbial cultures in one experiment.

National Academy of Sciences2013

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EPFL2017