Dia-interacting protein modulates formin-mediated actin assembly at the cell cortex.
BACKGROUND
Mammalian Diaphanous (mDia)-related formins and the N-WASP-activated Arp2/3 complex initiate the assembly of filamentous actin. Dia-interacting protein (DIP) binds via its amino-terminal SH3 domain to the proline-rich formin homology 1 (FH1) domain of mDia1 and mDia2 and to the N-WASp proline-rich region.


RESULTS
Here, we investigated an interaction between a conserved leucine-rich region (LRR) in DIP and the mDia FH2 domain that nucleates, processively elongates, and bundles actin filaments. DIP binding to mDia2 was regulated by the same Rho-GTPase-controlled autoinhibitory mechanism modulating formin-mediated actin assembly. DIP was previously shown to interact with and stimulate N-WASp-dependent branched filament assembly via Arp2/3. Despite direct binding to both mDia1 and mDia2 FH2 domains, DIP LRR inhibited only mDia2-dependent filament assembly and bundling in vitro. DIP expression interfered with filopodia formation, consistent with a role for mDia2 in assembly of these structures. After filopodia retraction into the cell body, DIP expression induced excessive nonapoptotic membrane blebbing, a physiological process involved in both cytokinesis and amoeboid cell movement. DIP-induced blebbing was dependent on mDia2 but did not require the activities of either mDia1 or Arp2/3.


CONCLUSIONS
These observations point to a pivotal role for DIP in the control of nonbranched and branched actin-filament assembly that is mediated by Diaphanous-related formins and activators of Arp2/3, respectively. The ability of DIP to trigger blebbing also suggests a role for mDia2 in the assembly of cortical actin necessary for maintaining plasma-membrane integrity.
Eisenmann
Kathryn M
KM
Laboratory of Cell Structure and Signal Integration, Van Andel Research Institute, Grand Rapids, MI 49503, USA.
Harris
Elizabeth S
ES
Kitchen
Susan M
SM
Holman
Holly A
HA
Higgs
Henry N
HN
Alberts
Arthur S
AS
D008841:Actin Cytoskeleton / Q000378:metabolism; D051376:Actin-Related Protein 2-3 Complex / Q000378:metabolism; D000199:Actins / Q000378:metabolism*; D048868:Adaptor Proteins, Signal Transducing / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D002352:Carrier Proteins / Q000378:metabolism*; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D000080865:Formins; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D009124:Muscle Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D017434:Protein Structure, Tertiary; D011554:Pseudopodia / Q000378:metabolism / Q000648:ultrastructure; D034741:RNA, Small Interfering / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D020764:cdc42 GTP-Binding Protein / Q000378:metabolism
D051376:Actin-Related Protein 2-3 Complex; D000199:Actins; D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; C491462:DIAPH2 protein, human; D000080865:Formins; D009124:Muscle Proteins; C486062:NCKIPSD protein, human; D034741:RNA, Small Interfering; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D020764:cdc42 GTP-Binding Protein