Quantitative Profile of Five Murine Core Proteomes Using Label-free Functional Proteomics

Abstract

Analysis of primary animal and human tissues is key in biological and biomedical research. Comparative proteomics analysis of primary biological material would benefit from uncomplicated experimental work flows capable of evaluating an unlimited number of samples. In this report we describe the application of label-free proteomics to the quantitative analysis of five mouse core proteomes. We developed a computer program and normalization procedures that allow exploitation of the quantitative data inherent in LC-MS/MS experiments for relative and absolute quantification of proteins in complex mixtures. Important features of this approach include (i) its ability to compare an unlimited number of samples, (ii) its applicability to primary tissues and cultured cells, (iii) its straightforward work flow without chemical reaction steps, and (iv) its usefulness not only for relative quantification but also for estimation of absolute protein abundance. We applied this approach to quantitatively characterize the most abundant proteins in murine brain, heart, kidney, liver, and lung. We matched 8,800 MS/MS peptide spectra to 1,500 proteins and generated 44,000 independent data points to profile the ϳ1,000 most abundant proteins in mouse tissues. This dataset provides a quantitative profile of the fundamental proteome of a mouse, identifies the major similarities and differences between organ-specific proteomes, and serves as a paradigm of how label-free quantitative MS can be used to characterize the phenotype of mammalian primary tissues at the molecular level. Molecular & Cellular Proteomics 6: 1560 -1573, 2007.Experiments on immortalized cell lines have resulted in the generation of a vast amount of information on the biological and biochemical processes that govern the function of cultured cells. However, discerning the mechanisms by which genes control mammalian physiology in vivo may only be achieved by investigations that involve the use of animal models of which the laboratory mouse (Mus musculus) offers many advantages (1). There is a wealth of resources related to the molecular biology of mouse cells that have benefited from genomics, transcriptomics, and, more recently, proteomics projects aimed at profiling the molecular composition of murine tissues (2-4). The generation of gene-targeted mice is particularly useful in advancing our understanding of how genes control fundamental processes of mammalian physiology (e.g. Refs. 5-9).Once created, finding the phenotype of a gene-targeted strain of mice is not a trivial task (1, 10). Several years are needed to fully characterize phenotypic alterations, and subtle phenotypes often go unnoticed. Robust and high throughput methods for profiling the proteomes of primary tissues in a quantitative fashion may expedite the search for phenotypic changes in gene-targeted and other animal models. Insights gained by powerful methods for the molecular characterization of primary tissues could also direct classical physiological experiments, reduce the number of experimental an...

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