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Non-canonical activity of the podosomal formin FMNL1? supports immune cell migration.

ABSTRACT: Having previously located the formin FMNL1 in macrophage podosomes, we developed an in vivo model to assess the role of FMNL1 in the migration activities of primary macrophages. Deletion of FMNL1 in mice was genetically lethal; however, targeted deletion in macrophages was achieved by employing macrophage-specific Cre. Unchallenged FMNL1-deficient mice exhibited an unexpected reduction in tissue-resident macrophages despite normal blood monocyte numbers. Upon immune stimulus, the absence of FMNL1 resulted in reduced macrophage recruitment in vivo, decreased migration in two-dimensional in vitro culture and a decrease in the number of macrophages exhibiting podosomes. Of the three described isoforms of FMNL1 - ?, ? and ? - only FMNL1? rescued macrophage migration when expressed exogenously in depleted macrophages. Surprisingly, mutation of residues in the FH2 domain of FMNL1? that disrupt barbed-end actin binding did not limit rescue of macrophage migration and podosome numbers. These observations suggest that FMNL1 contributes to macrophage migration activity by stabilizing the lifespan of podosomes without interaction of fast-growing actin termini.

SUBMITTER: Miller MR

PROVIDER: S-EPMC5450187 | biostudies-literature | 2017 May

REPOSITORIES: biostudies-literature

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Non-canonical activity of the podosomal formin FMNL1γ supports immune cell migration.

Miller Matthew R MR Miller Eric W EW Blystone Scott D SD

Journal of cell science 20170327 10

Having previously located the formin FMNL1 in macrophage podosomes, we developed an <i>in vivo</i> model to assess the role of FMNL1 in the migration activities of primary macrophages. Deletion of FMNL1 in mice was genetically lethal; however, targeted deletion in macrophages was achieved by employing macrophage-specific Cre. Unchallenged FMNL1-deficient mice exhibited an unexpected reduction in tissue-resident macrophages despite normal blood monocyte numbers. Upon immune stimulus, the absence ...[more]

PMID: 28348104

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Project description:BackgroundDisheveled-associated activator of morphogenesis 1 (DAAM1) is a formin acting downstream of Wnt signaling that is important for planar cell polarity. It has been shown to promote proper cell polarization during embryonic development in both Xenopus and Drosophila. Importantly, DAAM1 binds to Disheveled (Dvl) and thus functions downstream of the Frizzled receptors. Little is known of how DAAM1 is localized and functions in mammalian cells. We investigate here how DAAM1 affects migration and polarization of cultured cells and conclude that it plays a key role in centrosome polarity.Methodology/principal findingsUsing a specific antibody to DAAM1, we find that the protein localizes to the acto-myosin system and co-localizes with ventral myosin IIB-containing actin stress fibers. These fibers are particularly evident in the sub-nuclear region. An N-terminal region of DAAM1 is responsible for this targeting and the DAAM1(1-440) protein can interact with myosin IIB fibers independently of either F-actin or RhoA binding. We also demonstrate that DAAM1 depletion inhibits Golgi reorientation in wound healing assays. Wound-edge cells exhibit multiple protrusions characteristic of unpolarized cell migration. Finally, in U2OS cells lines stably expressing DAAM1, we observe an enhanced myosin IIB stress fiber network which opposes cell migration.Conclusions/significanceThis work highlights the importance of DAAM1 in processes underlying cell polarity and suggests that it acts in part by affecting the function of acto-myosin IIB system. It also emphasizes the importance of the N-terminal half of DAAM1. DAAM1 depletion strongly blocks centrosomal re-polarization, supporting the concept that DAAM1 signaling cooperates with the established Cdc42 associated polarity complex. These findings are also consistent with the observation that ablation of myosin IIB but not myosin IIA results in polarity defects downstream of Wnt signaling. The structure-function analysis of DAAM1 in cultured cells parallels more complex morphological events in the developing embryo.

Dataset Information

Non-canonical activity of the podosomal formin FMNL1? supports immune cell migration.

Publications

Non-canonical activity of the podosomal formin FMNL1γ supports immune cell migration.

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