Yeast Interspecies Comparative Proteomics Reveals Divergence in Expression Profiles and Provides Insights into Proteome Resource Allocation and Evolutionary Roles of Gene Duplication

Abstract

Omics analysis is a versatile approach for understanding the conservation and diversity of molecular systems across multiple taxa. In this study, we compared the proteome expression profiles of four yeast species (Saccharomyces cerevisiae, Saccharomyces mikatae, Kluyveromyces waltii, and Kluyveromyces lactis) grown on glucose-or glycerol-containing media. Conserved expression changes across all species were observed only for a small proportion of all proteins differentially expressed between the two growth conditions. Two Kluyveromyces species, both of which exhibited a high growth rate on glycerol, a nonfermentative carbon source, showed distinct species-specific expression profiles. In K. waltii grown on glycerol, proteins involved in the glyoxylate cycle and gluconeogenesis were expressed in high abundance. In K. lactis grown on glycerol, the expression of glycolytic and ethanol metabolic enzymes was unexpectedly low, whereas proteins involved in cytoplasmic translation, including ribosomal proteins and elongation factors, were highly expressed. These marked differences in the types of predominantly expressed proteins suggest that K. lactis optimizes the balance of proteome resource allocation between metabolism and protein synthesis giving priority to cellular growth. In S. cerevisiae, about 450 duplicate gene pairs were retained after whole-genome duplication. Intriguingly, we found that in the case of duplicates with conserved sequences, the total abundance of proteins encoded by a duplicate pair in S. cerevisiae was similar to that of protein encoded by nonduplicated ortholog in Kluyveromyces yeast. Genome-wide analysis of multiple organisms is one of the most useful approaches to uncover both the conservation and divergence of biological processes with respect to evolutionary history. Transcriptomic studies using DNA microarrays or deep-sequencing techniques have not only characterized the expression profiles of cell-type and -state specific genes involved in many biological processes, the have also revealed the expression patterns of genes involved in a variety among different species (1). Proteomic analysis, for which mass spectrometry is one of the most powerful and widely used analytical techniques (2), can provide much more direct evidence of both conservation and divergence of expression profiles across multiple species.Interspecies comparative proteomic studies with mass spectrometry have been hitherto carried out in primates (3, 4), yeasts (5), and a variety of other model organisms (6 -9). Abundance of the core proteome, for which orthologs have been identified in all organisms investigated, are well conserved across a wide range model organisms, including yeasts, plants, worms, flies, and humans (7). The profiles of orthologous protein abundance appear to be more conserved than profiles of transcript abundance in chimpanzees and humans (4), worms and flies (6), and many other prokaryotes and eukaryotes (8), suggesting that there is more selection pressure on protein expression than transcript e...

Proteomics analysis for asymmetric inheritance of preexisting proteins between mother and daughter cells in budding yeast

“…Protein extraction from yeast cells, digestion of proteins into tryptic peptides and their fractionation via isoelectronic focusing were performed according to our previous report (Kito et al 2016). Fractionated peptides were analyzed with a LC-MS/MS system consisting of a LTQ-Orbitrap mass spectrometer (Thermo Fisher Scientific) and a DiNa nano LC (KYA Technologies, Tokyo, Japan).…”

A strategy for absolute proteome quantification with mass spectrometry by hierarchical use of peptide‐concatenated standards

“…The absolute abundances measured by our strategy in which multiple PCSs are hierarchically used were compared to protein abundances provided by label‐free methods using mass spectrometric analyses. Relative copy numbers of expressed proteins calculated by identification frequency of peptide ions (PSM counts) in our large‐scale mass spectrometric analysis (see also Supporting Materials and methods) for budding yeast, which was cultured in our laboratory in identical conditions, showed significantly overall high correlation with the absolute abundances determined by the strategy used here (Spearman correlation coefficient = 0.79∼0.80, Fig. A and Supporting Information Fig.…”

“…(A) Comparison to the relative copy number calculated by the label‐free method. The relative copy numbers (ppm, fraction in total protein copy number) determined in our large‐scale analysis based on the identification frequency of peptide ions (PSM counts) (see also Supporting Materials and methods) was plotted against the absolute protein abundances determined by PCS in this study. (B) Comparison to the protein abundance in a public database (PaxDB ) integrated from many quantitative datasets mainly determined by label‐free methods.…”

“…For the design of artificial sequences suited for ID‐tag peptides, we explored the amino acid residues that affect spectral peak intensities in a large‐scale proteomic dataset, including 24 LC‐MS/MS analyses for 24 fractions of tryptic peptides separated by isoelectric focusing using the OFFGEL fractionator system (see also Supporting Materials and methods). As the numbers of identified peptides and their detected frequencies were relatively high for peptide ions with double‐charged state and within a range of peptide length from ten to 15 (Supporting Information Fig.…”

Estimating the protein burden limit of yeast cells by measuring the expression limits of glycolytic proteins

“…Proteins of interest were excised from the gels and digested using trypsin. The tryptic peptides were analyzed by LC-MS/MS consisting of an LTQ-Orbitrap mass spectrometer (ThermoFisher) and a DiNa nano LC (KYA Technologies) system according to the method described previously ( Kito et al, 2016 ). The peptide mixture was separated with reverse-phase chromatography.…”