The scaffold protein POSH regulates axon outgrowth.
How scaffold proteins integrate signaling pathways with cytoskeletal components to drive axon outgrowth is not well understood. We report here that the multidomain scaffold protein Plenty of SH3s (POSH) regulates axon outgrowth. Reduction of POSH function by RNA interference (RNAi) enhances axon outgrowth in differentiating mouse primary cortical neurons and in neurons derived from mouse P19 cells, suggesting POSH negatively regulates axon outgrowth. Complementation analysis reveals a requirement for the third Src homology (SH) 3 domain of POSH, and we find that the actomyosin regulatory protein Shroom3 interacts with this domain of POSH. Inhibition of Shroom3 expression by RNAi leads to increased process lengths, as observed for POSH RNAi, suggesting that POSH and Shroom function together to inhibit process outgrowth. Complementation analysis and interference of protein function by dominant-negative approaches suggest that Shroom3 recruits Rho kinase to inhibit process outgrowth. Furthermore, inhibition of myosin II function reverses the POSH or Shroom3 RNAi phenotype, indicating a role for myosin II regulation as a target of the POSH-Shroom complex. Collectively, these results suggest that the molecular scaffold protein POSH assembles an inhibitory complex that links to the actin-myosin network to regulate neuronal process outgrowth.
Taylor
Jennifer
J
Department of Biological Chemistry, University of Michigan, Ann Arbor, MI 48109, USA.
Chung
Kwan-Ho
KH
Figueroa
Claudia
C
Zurawski
Jonathan
J
Dickson
Heather M
HM
Brace
E J
EJ
Avery
Adam W
AW
Turner
David L
DL
Vojtek
Anne B
AB
D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001369:Axons / Q000378:metabolism* / Q000648:ultrastructure; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology; D003598:Cytoskeletal Proteins / Q000235:genetics / Q000378:metabolism*; D005816:Genetic Complementation Test; D006801:Humans; D051379:Mice; D008840:Microfilament Proteins / Q000235:genetics / Q000378:metabolism; D009474:Neurons / Q000166:cytology / Q000502:physiology*; D024507:Nonmuscle Myosin Type IIA / Q000235:genetics / Q000378:metabolism; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D017434:Protein Structure, Tertiary; D034622:RNA Interference; D020798:Two-Hybrid System Techniques; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D003598:Cytoskeletal Proteins; D008840:Microfilament Proteins; D020033:Protein Isoforms; C111370:Sh3md2 protein, mouse; C402112:Shrm protein, mouse; D054460:rho-Associated Kinases; D024507:Nonmuscle Myosin Type IIA