title: Phosphorylation and mRNA splicing of collapsin response mediator protein-2 determine inhibition of rho-associated protein kinase (ROCK) II function in carcinoma cell migration and invasion.
abstract: The Rho-associated protein kinases (ROCK I and II) are central regulators of important cellular processes such as migration and invasion downstream of the GTP-Rho. Recently, we reported collapsin response mediator protein (CRMP)-2 as an endogenous ROCK II inhibitor. To reveal how the CRMP-2-ROCK II interaction is controlled, we further mapped the ROCK II interaction site of CRMP-2 and examined whether phosphorylation states of CRMP-2 affected the interaction. Here, we show that an N-terminal fragment of the long CRMP-2 splice variant (CRMP-2L) alone binds ROCK II and inhibits colon carcinoma cell migration and invasion. Furthermore, the interaction of CRMP-2 and ROCK II is partially regulated by glycogen synthase kinase (GSK)-3 phosphorylation of CRMP-2, downstream of PI3K. Inhibition of PI3K reduced interaction of CRMP-2 with ROCK II, an effect rescued by simultaneous inhibition of GSK3. Inhibition of PI3K also reduced colocalization of ROCK II and CRMP-2 at the cell periphery in human breast carcinoma cells. Mimicking GSK3 phosphorylation of CRMP-2 significantly reduced CRMP-2 binding of recombinant full-length and catalytic domain of ROCK II. These data implicate GSK3 in the regulation of ROCK II-CRMP-2 interactions. Using phosphorylation-mimetic and -resistant CRMP-2L constructs, it was revealed that phosphorylation of CRMP-2L negatively regulates its inhibitory function in ROCK-dependent haptotactic cell migration, as well as invasion of human colon carcinoma cells. Collectively, the presented data show that CRMP-2-dependent regulation of ROCK II activity is mediated through interaction of the CRMP-2L N terminus with the ROCK II catalytic domain as well as by GSK3-dependent phosphorylation of CRMP-2.
authors.affiliation: From the Department of Biomedical Sciences, Faculty of Health and Medical Sciences, and Biotech Research and Innovation Centre (BRIC), University of Copenhagen, Ole Maaløes Vej 5, Copenhagen N, 2200, Denmark and.
authors.forename: Marie
authors.initials: M
authors.lastname: Morgan-Fisher
authors.forename: John R
authors.initials: JR
authors.lastname: Couchman
authors.forename: Atsuko
authors.initials: A
authors.lastname: Yoneda
mesh_terms: D000818:Animals; D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D002277:Carcinoma / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D045744:Cell Line, Tumor; D002465:Cell Movement; D003110:Colonic Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D005260:Female; D038362:Glycogen Synthase Kinase 3 / Q000235:genetics / Q000378:metabolism; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D009361:Neoplasm Invasiveness; D009363:Neoplasm Proteins / Q000235:genetics / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation / Q000235:genetics; D011485:Protein Binding / Q000235:genetics; D017434:Protein Structure, Tertiary; D012326:RNA Splicing; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D012334:RNA, Neoplasm / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D054460:rho-Associated Kinases / Q000235:genetics / Q000378:metabolism*
chemical_list: D036341:Intercellular Signaling Peptides and Proteins; D009363:Neoplasm Proteins; D009419:Nerve Tissue Proteins; D012333:RNA, Messenger; D012334:RNA, Neoplasm; C095114:collapsin response mediator protein-2; C515852:ROCK2 protein, human; C580818:ROCK2 protein, rat; D054460:rho-Associated Kinases; D038362:Glycogen Synthase Kinase 3
keywords: CRMP-2 Phosphorylation; Carcinoma; Cell Invasion; Cell Migration; Collapsin Response Mediator Protein-2; Glycogen Synthase Kinase 3; Protein-serine/threonine Kinase; Rho GTPases; Rho-associated Protein Kinase II
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