| Title: | Evolutionary Selection on Barrier Activity: Bar1 Is an Aspartyl Protease with Novel Substrate Specificity |
| Author(s): | Jones SK; Clarke SC; Craik CS; Bennett RJ; |
| Address: | "Department of Molecular Microbiology and Immunology, Brown University, Providence, Rhode Island, USA. Department of Pharmaceutical Chemistry, University of California-San Francisco, San Francisco, California, USA. Department of Molecular Microbiology and Immunology, Brown University, Providence, Rhode Island, USA richard_bennett@brown.edu" |
| Abstract: | "Peptide-based pheromones are used throughout the fungal kingdom for coordinating sexual responses between mating partners. Here, we address the properties and function of Bar1, an aspartyl protease that acts as a 'barrier' and antagonist to pheromone signaling in multiple species. Candida albicans Bar1 was purified and shown to exhibit preferential cleavage of native alpha pheromone over pheromones from related fungal species. This result establishes that protease substrate specificity coevolved along with changes in its pheromone target. Pheromone cleavage by Bar1 occurred between residues Thr-5 and Asn-6 in the middle of the tridecapeptide sequence. Surprisingly, proteolytic activity was independent of the amino acid residues present at the scissile bond and instead relied on residues at the C terminus of alpha pheromone. Unlike most aspartyl proteases, Bar1 also exhibited a near-neutral pH optimum and was resistant to the class-wide inhibitor pepstatin A. In addition, genetic analysis was performed on C. albicans BAR1 and demonstrated that the protease not only regulates endogenous pheromone signaling but also can limit interspecies pheromone signaling. We discuss these findings and propose that the unusual substrate specificity of Bar1 is a consequence of its coevolution with the alpha pheromone receptor Ste2 for their shared peptide target. IMPORTANCE: Pheromones are important for intraspecies communication across the tree of life. In the fungal kingdom, extracellular proteases play a key role in antagonizing pheromone signaling in multiple species. This study examines the properties and function of Candida albicans Bar1, an aspartyl protease that cleaves and thereby inactivates alpha pheromone. We demonstrate that Bar1 plays important roles in regulating both intra- and interspecies pheromone signaling. The fungal protease shows preferential activity on the endogenous pheromone, but, surprisingly, cleavage activity is dependent on amino acid residues distal to the scissile bond. We propose that the unusual substrate specificity of Bar1 is a direct result of coevolution with Ste2, the receptor for alpha pheromone, for recognition of the same peptide target. The novel specificity of Bar1 reveals the complex forces shaping the evolution of mating pathways in fungi and uncovers a protease with potentially important applications in the biotechnology industry" |
| Keywords: | Aspartic Acid Proteases/*genetics/*metabolism Candida albicans/*enzymology/genetics/physiology Hydrogen-Ion Concentration Pepstatins/metabolism Pheromones/*metabolism Protease Inhibitors/metabolism Proteolysis Signal Transduction Substrate Specificity; |
| Notes: | "MedlineJones, Stephen K Jr Clarke, Starlynn C Craik, Charles S Bennett, Richard J eng T32 GM007810/GM/NIGMS NIH HHS/ AI081704/AI/NIAID NIH HHS/ P41 GM103481/GM/NIGMS NIH HHS/ P41GM103481/GM/NIGMS NIH HHS/ AI112362/AI/NIAID NIH HHS/ R21 AI112363/AI/NIAID NIH HHS/ R21 AI112362/AI/NIAID NIH HHS/ F31DE022701/DE/NIDCR NIH HHS/ P50 GM082250/GM/NIGMS NIH HHS/ F31 DE022701/DE/NIDCR NIH HHS/ R01 AI081704/AI/NIAID NIH HHS/ Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't 2015/11/26 mBio. 2015 Nov 24; 6(6):e01604-15. doi: 10.1128/mBio.01604-15" |
