The actin-associating protein Tm5NM1 blocks mesenchymal motility without transition to amoeboid motility

Lees, Justin G.; Bach, Cuc T.; Bradbury, Peta; Paul, Andre; Gunning, Peter W.; O’Neill, Geraldine M.

doi:10.1038/onc.2010.516

Cited by 26 publications

(35 citation statements)

References 37 publications

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“…In agreement with previous studies demonstrating that tropomyosin overexpression reduces cellular motility (38)(39)(40), we observed that human Tpm1 expression significantly diminished the speed of HeLa cell migration (Fig. S4 C and D).…”

Section: Tpm1 Overexpression Partially Restores Actin Cytoskeletonsupporting

confidence: 93%

“…Tropomyosin expression increased or decreased focal adhesion area (38)(39)(40), consistent with the idea that maximal migration rates require the specific organization of the focal adhesions and actin filaments. Similarly, our findings provide evidence indicating that the tight regulation of tropomyosin and the amount of acetylated Tpm1 is necessary for maximal cellular motility (Fig.…”

Section: Tpm1 Overexpression Partially Restores Actin Cytoskeletonsupporting

confidence: 80%

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N-terminal acetylome analyses and functional insights of the N-terminal acetyltransferase NatB

Damme

Lasa²,

Polevoda

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

187

203

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Protein N-terminal acetylation (Nt-acetylation) is an important mediator of protein function, stability, sorting, and localization. Although the responsible enzymes are thought to be fairly well characterized, the lack of identified in vivo substrates, the occurrence of Nt-acetylation substrates displaying yet uncharacterized N-terminal acetyltransferase (NAT) specificities, and emerging evidence of posttranslational Nt-acetylation, necessitate the use of genetic models and quantitative proteomics. NatB, which targets Met-Glu-, Met-Asp-, and Met-Asn-starting protein N termini, is presumed to Nt-acetylate 15% of all yeast and 18% of all human proteins. We here report on the evolutionary traits of NatB from yeast to human and demonstrate that ectopically expressed hNatB in a yNatB-Δ yeast strain partially complements the natB-Δ phenotypes and partially restores the yNatB Nt-acetylome. Overall, combining quantitative N-terminomics with yeast studies and knockdown of hNatB in human cell lines, led to the unambiguous identification of 180 human and 110 yeast NatB substrates. Interestingly, these substrates included Met-Gln-N-termini, which are thus now classified as in vivo NatB substrates. We also demonstrate the requirement of hNatB activity for maintaining the structure and function of actomyosin fibers and for proper cellular migration. In addition, expression of tropomyosin-1 restored the altered focal adhesions and cellular migration defects observed in hNatB-depleted HeLa cells, indicative for the conserved link between NatB, tropomyosin, and actin cable function from yeast to human.

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Section: Tpm1 Overexpression Partially Restores Actin Cytoskeletonsupporting

confidence: 93%

Section: Tpm1 Overexpression Partially Restores Actin Cytoskeletonsupporting

confidence: 80%

N-terminal acetylome analyses and functional insights of the N-terminal acetyltransferase NatB

Damme

Lasa²,

Polevoda

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

187

203

View full text Add to dashboard Cite

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“…Cell lines were maintained in Dulbecco's modified eagle's medium (DMEM) supplemented with 10% FBS. Growth of cells in 3D collagen gels, media for live imaging, and invasion assays through 3D collagen gels was based on protocols previously reported to identify invasion plasticity (21) and as we have previously used (22). The following antibodies were used: anti-NEDD9 (ImmuQuest; clone 2G9 and Cell Signaling Technology, clone 29G), antipan-Rac (#610650 BD Bioscience), anti-HSP70 and anti-a-tubulin (Sigma-Aldrich), and horseradish peroxidase-conjugated anti-mouse and anti-rabbit (Amersham and Biorad) and anti-rat Alexa-Fluor 647 conjugate (Molecular Probes; Invitrogen).…”

Section: Cell Culture and Antibodiesmentioning

confidence: 99%

NEDD9 Regulates 3D Migratory Activity Independent of the Rac1 Morphology Switch in Glioma and Neuroblastoma

Zhong

Bach

Shum

et al. 2014

Molecular Cancer Research

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Metastasizing tumor cells must transmigrate the dense extracellular matrix that surrounds most organs. The use of three-dimensional (3D) collagen gels has revealed that many cancer cells can switch between different modes of invasion that are characterized by distinct morphologies (e.g., rounded vs. elongated). The adhesion protein NEDD9 has the potential to regulate the switch between elongated and rounded morphologies; therefore, its role was interrogated in the invasion switch of glioblastoma and neuroblastoma tumors that similarly derive from populations of neural crest cells. Interestingly, siRNA-mediated depletion of NEDD9 failed to induce cell rounding in glioma or neuroblastoma cells, contrasting the effects that have been described in other tumor model systems. Given that Rac1 GTPase has been suggested to mediate the switch between elongated and rounded invasion, the functionality of the Rac1 morphology switch was evaluated in the glioma and neuroblastoma cells. Using both dominant-negative Rac1 and Rac1-specific siRNA, the presence of this morphologic switch was confirmed in the neuroblastoma, but not in the glioma cells. However, in the absence of a morphologic change following NEDD9 depletion, a significant decrease in the cellular migration rate was observed. Thus, the data reveal that NEDD9 can regulate 3D migration speed independent of the Rac1 morphology switch.

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“…Thus, cells with altered tropomyosin expression profiles display differential actin filament stability (2,4,13). Moreover, tropomyosin isoform expression is correlated with different focal-adhesion morphology (4,14) and isoform-specific effects on cell migration (2,4,(14)(15)(16). The tropomyosin isoform Tm5NM1 is ubiquitously expressed (17), and elevated expression inhibits both 2-dimensional (2D) (2,4) and 3D (16) cell migration.…”

mentioning

confidence: 99%

“…Tm5NM1-mediated actin filament stabilization is accompanied by reduced activation of the nonreceptor kinase Src (16). Src is a major regulator of focal-adhesion disassembly (18,19), and depletion of Src family kinase activity reduces cell motility (20) and causes enlarged and stabilized focal adhesions (18,21).…”

mentioning

confidence: 99%

Tropomyosin Tm5NM1 Spatially Restricts Src Kinase Activity through Perturbation of Rab11 Vesicle Trafficking

Bach

Murray

Owen

et al. 2014

Molecular and Cellular Biology

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dIn order for cells to stop moving, they must synchronously stabilize actin filaments and their associated focal adhesions. How these two structures are coordinated in time and space is not known. We show here that the actin association protein Tm5NM1, which induces stable actin filaments, concurrently suppresses the trafficking of focal-adhesion-regulatory molecules. Using combinations of fluorescent biosensors and fluorescence recovery after photobleaching (FRAP), we demonstrate that Tm5NM1 reduces the level of delivery of Src kinase to focal adhesions, resulting in reduced phosphorylation of adhesion-resident Src substrates. Live imaging of Rab11-positive recycling endosomes that carry Src to focal adhesions reveals disruption of this pathway. We propose that tropomyosin synchronizes adhesion dynamics with the cytoskeleton by regulating actin-dependent trafficking of essential focal-adhesion molecules. Cell migration is essential for normal embryonic development, wound healing, and immune response, and migration dysregulation underpins an array of pathological conditions. Much focus is given to the initiation and speed of migration; however, of equal importance are the mechanisms by which migration is arrested. Integrin-based focal adhesions, which are bound on the external surface to the extracellular matrix and internally to the actin cytoskeleton, are key structural elements of the machinery that drives cell migration (1). While actin filament stabilization inhibits cell migration (2-6), little is known about how the associated focal adhesions are synchronously stabilized along with actin filaments. This is critical, because events that trigger focal adhesion disassembly correspondingly break the adhesion-cytoskeleton linkage (7-9), facilitating new cycles of membrane protrusion and forward movement (10). Thus, in order to stop migrating, cells must have mechanisms to synchronize actin and adhesion stability.The tropomyosins are a multi-isoform family of actin-binding proteins that display spatially and temporally restricted association with discrete actin filament populations (11). Dimers of tropomyosin associate head to tail, forming a coiled polymer that lies in the major groove of the actin filament. The association of discrete tropomyosin isoforms is thought to regulate the dynamic state of actin filaments by affecting the association of other actinregulatory proteins (12). Thus, cells with altered tropomyosin expression profiles display differential actin filament stability (2, 4, 13). Moreover, tropomyosin isoform expression is correlated with different focal-adhesion morphology (4, 14) and isoform-specific effects on cell migration (2, 4, 14-16). The tropomyosin isoform Tm5NM1 is ubiquitously expressed (17), and elevated expression inhibits both 2-dimensional (2D) (2, 4) and 3D (16) cell migration. In line with these migration effects, Tm5NM1 induces actin filament stability (13) coupled with highly stabilized focal adhesions (4).Tm5NM1-mediated actin filament stabilization is accompanied by reduce...

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The actin-associating protein Tm5NM1 blocks mesenchymal motility without transition to amoeboid motility

Cited by 26 publications

References 37 publications

N-terminal acetylome analyses and functional insights of the N-terminal acetyltransferase NatB

N-terminal acetylome analyses and functional insights of the N-terminal acetyltransferase NatB

NEDD9 Regulates 3D Migratory Activity Independent of the Rac1 Morphology Switch in Glioma and Neuroblastoma

Tropomyosin Tm5NM1 Spatially Restricts Src Kinase Activity through Perturbation of Rab11 Vesicle Trafficking

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