Quantitative proteomic analysis of ribosomal protein L35b mutant of Saccharomyces cerevisiae.
Recent studies have revealed that in higher eukaryotes, several ribosomal proteins are involved in some pathological events or developmental defects, indicating that ribosomal proteins perform unconventional functions other than protein biosynthesis. To obtain an insight into the novel roles of ribosomal proteins, we aimed to analyze the changes in proteome expression in ribosomal protein mutants by using Saccharomyces cerevisiae as a model system. We introduced the rpl35bDelta mutation into the 4159 green fluorescent protein (GFP)-tagged yeast strains by using the synthetic genetic array (SGA) method, and performed quantitative proteomic analysis by using a multilabel microplate reader and flow cytometer. We identified 22 upregulated and 20 downregulated proteins in the rpl35bDelta mutant. These proteins were primarily classified into the Gene Ontology (GO) categories of cellular biosynthetic process, translation, protein or nucleotide metabolic process, cell wall organization and biogenesis, and hyperosmotic response. We also investigated the correlation between the mRNA and protein levels of the identified proteins. Our results show that a ribosomal protein mutation can lead to perturbation in the expression of several proteins, including some other ribosomal proteins. Furthermore, our approach of combining a library of GFP-tagged yeast strains and the SGA method provides an effective and highly sensitive method for dynamic analysis of the effects of various mutations on proteome expression.
Song
Yong Bhum
YB
School of Biological Sciences, and Research Center for Functional Cellulomics, Institute of Microbiology, Seoul National University, Seoul, Republic of Korea.
Jhun
Min A
MA
Park
Taesung
T
Huh
Won-Ki
WK
D015536:Down-Regulation; D017353:Gene Deletion; D005800:Genes, Fungal; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D016133:Polymerase Chain Reaction; D040901:Proteomics; D012331:RNA, Fungal / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012269:Ribosomal Proteins / Q000235:genetics* / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics* / Q000378:metabolism*; D015854:Up-Regulation
D012331:RNA, Fungal; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D012269:Ribosomal Proteins; D029701:Saccharomyces cerevisiae Proteins; D049452:Green Fluorescent Proteins