title: PakB binds to the SH3 domain of Dictyostelium Abp1 and regulates its effects on cell polarity and early development.
abstract: Dictyostelium p21-activated kinase B (PakB) phosphorylates and activates class I myosins. PakB colocalizes with myosin I to actin-rich regions of the cell, including macropinocytic and phagocytic cups and the leading edge of migrating cells. Here we show that residues 1-180 mediate the cellular localization of PakB. Yeast two-hybrid and pull-down experiments identify two proline-rich motifs in PakB-1-180 that directly interact with the SH3 domain of Dictyostelium actin-binding protein 1 (dAbp1). dAbp1 colocalizes with PakB to actin-rich regions in the cell. The loss of dAbp1 does not affect the cellular distribution of PakB, whereas the loss of PakB causes dAbp1 to adopt a diffuse cytosolic distribution. Cosedimentation studies show that the N-terminal region of PakB (residues 1-70) binds directly to actin filaments, whereas dAbp1 exhibits only a low affinity for filamentous actin. PakB-1-180 significantly enhances the binding of dAbp1 to actin filaments. When overexpressed in PakB-null cells, dAbp1 completely blocks early development at the aggregation stage, prevents cell polarization, and significantly reduces chemotaxis rates. The inhibitory effects are abrogated by the introduction of a function-blocking mutation into the dAbp1 SH3 domain. We conclude that PakB plays a critical role in regulating the cellular functions of dAbp1, which are mediated largely by its SH3 domain.
authors.affiliation: Department of Biomedical and Molecular Sciences, Queen's University, Kingston, ON K7L 3N6, Canada.
authors.forename: Yidai
authors.initials: Y
authors.lastname: Yang
authors.forename: Marc
authors.initials: M
authors.lastname: de la Roche
authors.forename: Scott W
authors.initials: SW
authors.lastname: Crawley
authors.forename: Zhihao
authors.initials: Z
authors.lastname: Li
authors.forename: Emilia
authors.initials: E
authors.lastname: Furmaniak-Kazmierczak
authors.forename: Graham P
authors.initials: GP
authors.lastname: Côté
mesh_terms: D008841:Actin Cytoskeleton / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D016764:Cell Polarity; D002633:Chemotaxis / Q000235:genetics; D004023:Dictyostelium / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D018507:Gene Expression Regulation, Developmental; D008840:Microfilament Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009218:Myosins / Q000235:genetics / Q000378:metabolism; D054730:Protein Interaction Domains and Motifs; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction
chemical_list: D008840:Microfilament Proteins; D015800:Protozoan Proteins; D011494:Protein Kinases; C093485:110 kDa protein kinase, Dictyostelium; D009218:Myosins