Abstract
Frizzled receptors have seven membrane-spanning helices and are considered as atypical G protein-coupled receptors (GPCRs). The mating response of the yeast Saccharomyces cerevisiae is mediated by a GPCR signaling system and this model organism has been used extensively in the past to study mammalian GPCR function. We show here that human Frizzled receptors (Fz1 and Fz2) can be properly targeted to the yeast plasma membrane, and that they stimulate the yeast mating pathway in the absence of added Wnt ligands, as evidenced by cell cycle arrest in G1 and reporter gene expression dependent on the mating pathway-activated FUS1 gene. Introducing intracellular portions of Frizzled receptors into the Ste2p backbone resulted in the generation of constitutively active receptor chimeras that retained mating factor responsiveness. Introducing intracellular portions of Ste2p into the Frizzled receptor backbone was found to strongly enhance mating pathway activation as compared to the native Frizzleds, likely by facilitating interaction with the yeast Galpha protein Gpa1p. Furthermore, we show reversibility of the highly penetrant G1-phase arrests exerted by the receptor chimeras by deletion of the mating pathway effector FAR1. Our data demonstrate that Frizzled receptors can functionally replace mating factor receptors in yeast and offer an experimental system to study modulators of Frizzled receptors.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Amino Acid Sequence
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Blotting, Western
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Cell Cycle / genetics
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Cell Cycle / physiology
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Cell Cycle Proteins / genetics
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Cell Cycle Proteins / metabolism
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Cyclin-Dependent Kinase Inhibitor Proteins
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Frizzled Receptors / genetics
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Frizzled Receptors / metabolism
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Frizzled Receptors / physiology*
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Humans
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Microscopy, Confocal
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Molecular Sequence Data
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Receptors, G-Protein-Coupled / genetics
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Receptors, G-Protein-Coupled / metabolism
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Receptors, G-Protein-Coupled / physiology
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Receptors, Mating Factor / genetics
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Receptors, Mating Factor / metabolism
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / metabolism
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Repressor Proteins / genetics
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Repressor Proteins / metabolism
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Saccharomyces cerevisiae / genetics
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Saccharomyces cerevisiae / physiology*
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Saccharomyces cerevisiae Proteins / genetics
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Saccharomyces cerevisiae Proteins / metabolism
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Sequence Homology, Amino Acid
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Signal Transduction / genetics
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Signal Transduction / physiology*
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Wnt Proteins / genetics
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Wnt Proteins / metabolism
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Wnt Proteins / physiology
Substances
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Cell Cycle Proteins
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Cyclin-Dependent Kinase Inhibitor Proteins
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FAR1 protein, S cerevisiae
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FZD1 protein, human
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FZD2 protein, human
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Frizzled Receptors
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Receptors, G-Protein-Coupled
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Receptors, Mating Factor
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Recombinant Fusion Proteins
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Repressor Proteins
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STE2 protein, S cerevisiae
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Saccharomyces cerevisiae Proteins
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Wnt Proteins