Identification of FHOD1-binding proteins and mechanisms of FHOD1-regulated actin dynamics.
Formin homology-2-domain containing protein 1 (FHOD1) regulates gene transcription, actin-cytoskeleton structure, and cell migration. To gain insight into the mechanisms by which FHOD1 mediates these diverse activities, a yeast-two-hybrid screen was performed to identify FHOD1-binding proteins. Three proteins specifically interacted with the carboxy-terminal two-thirds of FHOD1, which includes the FH1, FH2, and diaphanous activating domains (DAD). The newly identified FHOD1-binding proteins are protein kinase C binding protein 1 (PRKCBP1), cyclophilin B, and an isoform of WASP-interacting SH3-domain protein/diaphanous-interacting protein 1 (WISH/DIP1), named WISH-B. The proline-rich FH1 domain of FHOD1 was sufficient to interact with the central portion of PRKCP1 and full-length cyclophilin B. The FH1 domain also interacted with full-length WISH-B, but the extreme amino-terminus was sufficient to associate with WISH-B as well. WISH-B altered the solubility of FHOD1 in vitro and a truncation mutant containing the amino-terminal 227 residues of WISH-B disrupted FHOD1-induced stress fibers. WISH-B did not affect FHOD1-induced gene transcription through the serum response factor (SRF) recognition site on the skeletal alpha actin promoter (SkA). However, stabilization of F-actin prevented FHOD1 dependent activation of this promoter in presence of high, but not low serum concentrations. Thus, the identification of a new FHOD1-binding protein provides insight into the mechanisms by which FHOD1 regulates actin polymerization and transcription.
Westendorf
Jennifer J
JJ
The Cancer Center and Department of Orthopaedic Surgery, University of Minnesota, Minneapolis, Minnesota, USA. weste047@umn.edu
Koka
Sreenivas
S
D000199:Actins / Q000235:genetics*; D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism*; D002460:Cell Line; D021983:Cyclophilins / Q000378:metabolism; D005326:Fetal Proteins / Q000235:genetics / Q000378:metabolism*; D000080865:Formins; D006801:Humans; D051379:Mice; D009124:Muscle Proteins / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D019696:Peptidylprolyl Isomerase; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D025941:Protein Interaction Mapping; D017434:Protein Structure, Tertiary; D000075370:Receptors for Activated C Kinase; D011956:Receptors, Cell Surface / Q000378:metabolism; D020218:Response Elements / Q000235:genetics; D022502:Stress Fibers / Q000378:metabolism* / Q000648:ultrastructure; D015533:Transcriptional Activation / Q000235:genetics; D025521:Tumor Suppressor Proteins; D020798:Two-Hybrid System Techniques
D000199:Actins; D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; C120021:FHOD1 protein, human; D005326:Fetal Proteins; D000080865:Formins; D009124:Muscle Proteins; C426229:Nckipsd protein, mouse; D009687:Nuclear Proteins; D000075370:Receptors for Activated C Kinase; D011956:Receptors, Cell Surface; D025521:Tumor Suppressor Proteins; C000617312:ZMYND8 protein, human; C081786:cyclophilin B; D021983:Cyclophilins; D019696:Peptidylprolyl Isomerase