title: Phosphorylation on threonine 11 of β-dystrobrevin alters its interaction with kinesin heavy chain.
abstract: Dystrobrevin family members (α and β) are cytoplasmic components of the dystrophin-associated glycoprotein complex, a multimeric protein complex first isolated from skeletal muscle, which links the extracellular matrix to the actin cytoskeleton. Dystrobrevin shares high homology with the cysteine-rich and C-terminal domains of dystrophin and a common domain organization. The β-dystrobrevin isoform is restricted to nonmuscle tissues, serves as a scaffold for signaling complexes, and may participate in intracellular transport through its interaction with kinesin heavy chain. We have previously characterized the molecular determinants affecting the β-dystrobrevin-kinesin heavy chain interaction, among which is cAMP-dependent protein kinase [protein kinase A (PKA)] phosphorylation of β-dystrobrevin. In this study, we have identified β-dystrobrevin residues phosphorylated in vitro by PKA with pull-down assays, surface plasmon resonance measurements, and MS analysis. Among the identified phosphorylated residues, we demonstrated, by site-directed mutagenesis, that Thr11 is the regulatory site for the β-dystrobrevin-kinesin interaction. As dystrobrevin may function as a signaling scaffold for kinases/phosphatases, we also investigated whether β-dystrobrevin is phosphorylated in vitro by kinases other than PKA. Thr11 was phosphorylated by protein kinase C, suggesting that this represents a key residue modified by the activation of different signaling pathways.
authors.affiliation: Department of Cell Biology and Neuroscience, Istituto Superiore di Sanità, Rome, Italy.
authors.forename: Federica
authors.initials: F
authors.lastname: Fratini
authors.forename: Gianfranco
authors.initials: G
authors.lastname: Macchia
authors.forename: Paola
authors.initials: P
authors.lastname: Torreri
authors.forename: Andrea
authors.initials: A
authors.lastname: Matteucci
authors.forename: Anna Maria
authors.initials: AM
authors.lastname: Salzano
authors.forename: Marco
authors.initials: M
authors.lastname: Crescenzi
authors.forename: Pompeo
authors.initials: P
authors.lastname: Macioce
authors.forename: Tamara C
authors.initials: TC
authors.lastname: Petrucci
authors.forename: Marina
authors.initials: M
authors.lastname: Ceccarini
mesh_terms: D000595:Amino Acid Sequence; D015153:Blotting, Western; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D049029:Dystrophin-Associated Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D047468:Immunoprecipitation; D016547:Kinesins / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation / Q000235:genetics; D009479:Neuropeptides / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D020349:Surface Plasmon Resonance; D013912:Threonine / Q000235:genetics / Q000378:metabolism*
chemical_list: C111539:DTNB protein, human; D049029:Dystrophin-Associated Proteins; D009479:Neuropeptides; D013912:Threonine; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C; D016547:Kinesins