The Unconventional Myosin, Myo2p, Is a Calmodulin Target at Sites of Cell Growth in Saccharomyces cerevisiae

• Published in: The Journal of cell biology 1994-02, Vol.124 (3), p.315-323
• Main Authors: Brockerhoff, Susan E., Stevens, Richard C., Davis, Trisha N.
• Format: Article
• Language: English
• Subjects: Alleles; Antibodies; Base Sequence; Biological and medical sciences; Calcium - metabolism; Calmodulin - genetics; Calmodulin - metabolism; Carrier Proteins - genetics; Carrier Proteins - metabolism; Cell cycle; Cell cycle, cell proliferation; Cell Division; Cell growth; Cell physiology; Cells; Cellular biology; Cytoskeletal Proteins; Egtazic Acid - pharmacology; Fluorescent Antibody Technique; Fundamental and applied biological sciences. Psychology; Fungal Proteins - genetics; Fungal Proteins - metabolism; Genetic mutation; Molecular and cellular biology; Molecular Sequence Data; Mother cells; Mutation; Myosin Heavy Chains; Myosin Type II; Myosin Type V; Neurons; Phenotypes; Physical growth; Plasmids; Recombinant Fusion Proteins - metabolism; Saccharomyces cerevisiae; Saccharomyces cerevisiae - cytology; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae - growth & development; Saccharomyces cerevisiae - metabolism; Saccharomyces cerevisiae Proteins; Schizosaccharomyces pombe Proteins; Yeasts
• ISSN: 0021-9525; 1540-8140
• DOI: 10.1083/jcb.124.3.315

Myo2p is an unconventional myosin required for polarized growth in Saccharomyces cerevisiae. Four lines of evidence suggest that (a) Myo2p is a target of calmodulin at sites of cell growth, and (b) the interaction between Myo2p and calmodulin is Ca2+ independent. First, as assessed by indirect immunofluorescence, the distributions of Myo2p and calmodulin are nearly indistinguishable throughout the cell cycle. Second, a genetic analysis indicates that mutations in CMD1 show allele-specific synthetic lethality with the myo2-66 conditional mutation. Mutations that inactivate the Ca2+-binding sites of calmodulin have little or no effect on strains carrying myo2-66, whereas an allele with a mutation outside the Ca2+-binding sites dramatically increases the severity of the phenotype conferred by myo2-66. Third, Myo2p coimmunoprecipitates with calmodulin in the presence of Ca2+ or EGTA. Finally, we used a modified gel overlay assay to demonstrate direct interaction between calmodulin and fusion proteins containing portions of Myo2p. Calmodulin binds specifically to the region of Myo2p containing six tandem repeats of a motif called an IQ site. Binding occurs in either Ca2+ or EGTA, and only two sites are required to observe binding.