title: Evolution of the SH3 Domain Specificity Landscape in Yeasts.
abstract: To explore the conservation of Src homology 3 (SH3) domain-mediated networks in evolution, we compared the specificity landscape of these domains among four yeast species, Saccharomyces cerevisiae, Ashbya gossypii, Candida albicans, and Schizosaccharomyces pombe, encompassing 400 million years of evolution. We first aligned and catalogued the families of SH3-containing proteins in these four species to determine the relationships between homologous domains. Then, we tagged and purified all soluble SH3 domains (82 in total) to perform a quantitative peptide assay (SPOT) for each SH3 domain. All SPOT readouts were hierarchically clustered and we observed that the organization of the SH3 specificity landscape in three distinct profile classes remains conserved across these four yeast species. We also produced a specificity profile for each SH3 domain from manually aligned top SPOT hits and compared the within-family binding motif consensus. This analysis revealed a striking example of binding motif divergence in a C. albicans Rvs167 paralog, which cannot be explained by overall SH3 sequence or interface residue divergence, and we validated this specificity change with a yeast two-hybrid (Y2H) assay. In addition, we show that position-weighted matrices (PWM) compiled from SPOT assays can be used for binding motif screening in potential binding partners and present cases where motifs are either conserved or lost among homologous SH3 interacting proteins. Finally, by comparing pairwise SH3 sequence identity to binding profile correlation we show that for ~75% of all analyzed families the SH3 specificity profile was remarkably conserved over a large evolutionary distance. Thus, a high sequence identity within an SH3 domain family predicts conserved binding specificity, whereas divergence in sequence identity often coincided with a change in binding specificity within this family. As such, our results are important for future studies aimed at unraveling complex specificity networks of peptide recognition domains in higher eukaryotes, including mammals.
authors.affiliation: EMBL/CRG Systems Biology Research Unit, Centre for Genomic Regulation-CRG, Barcelona, Spain.
authors.forename: Erik
authors.initials: E
authors.lastname: Verschueren
authors.affiliation: Department of Molecular and Cellular Genetics, UMR7156, Université de Strasbourg and CNRS, Strasbourg, France.
authors.forename: Matthias
authors.initials: M
authors.lastname: Spiess
authors.affiliation: Department of Medical Biochemistry, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands.
authors.forename: Areti
authors.initials: A
authors.lastname: Gkourtsa
authors.affiliation: Department of Medical Biochemistry, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands.
authors.forename: Teja
authors.initials: T
authors.lastname: Avula
authors.affiliation: Institut für Medizinische Immunologie, Humboldt-Universität zu Berlin, Berlin, Germany.
authors.forename: Christiane
authors.initials: C
authors.lastname: Landgraf
authors.affiliation: Department of Molecular and Cellular Genetics, UMR7156, Université de Strasbourg and CNRS, Strasbourg, France; Institut für Medizinische Immunologie, Humboldt-Universität zu Berlin, Berlin, Germany.
authors.forename: Victor Tapia
authors.initials: VT
authors.lastname: Mancilla
authors.affiliation: Department of Molecular and Cellular Genetics, UMR7156, Université de Strasbourg and CNRS, Strasbourg, France.
authors.forename: Aline
authors.initials: A
authors.lastname: Huber
authors.affiliation: Institut für Medizinische Immunologie, Humboldt-Universität zu Berlin, Berlin, Germany.
authors.forename: Rudolf
authors.initials: R
authors.lastname: Volkmer
authors.affiliation: Department of Molecular and Cellular Genetics, UMR7156, Université de Strasbourg and CNRS, Strasbourg, France.
authors.forename: Barbara
authors.initials: B
authors.lastname: Winsor
authors.affiliation: EMBL/CRG Systems Biology Research Unit, Centre for Genomic Regulation-CRG, Barcelona, Spain.
authors.forename: Luis
authors.initials: L
authors.lastname: Serrano
authors.affiliation: Department of Medical Biochemistry, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands.
authors.forename: Frans
authors.initials: F
authors.lastname: Hochstenbach
authors.affiliation: Department of Medical Biochemistry, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands.
authors.forename: Ben
authors.initials: B
authors.lastname: Distel
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D002176:Candida albicans / Q000378:metabolism; D019143:Evolution, Molecular; D005656:Fungal Proteins / Q000737:chemistry* / Q000378:metabolism; D008969:Molecular Sequence Data; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D012568:Schizosaccharomyces / Q000378:metabolism; D016415:Sequence Alignment; D020798:Two-Hybrid System Techniques; D015003:Yeasts / Q000378:metabolism*; D018909:src Homology Domains
chemical_list: D005656:Fungal Proteins