Latrunculin a

Manufactured by Bio-Techne

Sourced in United Kingdom

Latrunculin A is a potent inhibitor of actin polymerization. It binds to actin monomers, preventing their assembly into filaments. Latrunculin A is commonly used in cell biology research to study the role of the actin cytoskeleton in various cellular processes.

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Lab products found in correlation

Fetal bovine serum (fbs), by Thermo Fisher Scientific (7 mentions) Ck 666, by Merck Group (4 mentions) Fm4 64, by Thermo Fisher Scientific (4 mentions) Dimethyl sulfoxide (dmso), by Merck Group (3 mentions) Blebbistatin, by Abcam (3 mentions) Jasplakinolide, by Bio-Techne (3 mentions) Latrunculin a, by Merck Group (3 mentions) Y 27632, by Merck Group (3 mentions) Cytochalasin d, by Merck Group (3 mentions) Blebbistatin, by Merck Group (2 mentions) Nsc23766, by Bio-Techne (2 mentions) Y 27632, by Bio-Techne (2 mentions) Cytochalasin d, by Bio-Techne (2 mentions) Gm6001, by Merck Group (2 mentions) Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (2 mentions) Lipofectamine 2000 transfection reagent, by Thermo Fisher Scientific (2 mentions) Blebbistatin, by Bio-Techne (1 mentions) Nocodazole, by Merck Group (1 mentions) Fibronectin, by Merck Group (1 mentions) Anti actin, by Merck Group (1 mentions)

Spelling variants of this product

Latrunculin A (latA) (3 citations)

25 protocols using latrunculin a

Imaging and Immunoblotting of Nuclear Proteins

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For imaging the nucleus, cells were incubated with 200 ng ml⁻¹ of Hoechst 33342 (Life Technologies) or 34580 (Invitrogen) for 30 min at 37 °C and 5% CO₂.
The following antibodies were used for immunoblotting: LmnA/C (H110, Santa Cruz), anti-ArpC4 and anti-sun2 (Abcam), and anti-actin (Millipore). Antibodies were used at 1:1,000 dilution.
For immunofluorescence, we used Alexa Fluor 594-coupled phalloidin (Invitrogen; 1:400), anti-LmnAC (N18, Santa Cruz (1:50) and clone 4C11 from Sigma Aldrich (1:2,000)), anti-Lamin B1 (ab16048, Abcam (1:500)) and anti-Arp3 (Abcam; 1:200); DAPI (4,6-diamidino-2-phenylindole) for DNA staining; and secondary antibodies anti-mouse-Alexa488 and anti-Goat-Alexa488 from Jackson ImmunoResearch Laboratories were used at 1:400. Slides were mounted with custom-made Moviol.
For lymphatic vessels visualization, anti-LYVE-1 (R&D System, 1/1,000) was used.
For drug treatment CK666, latrunculin A and blebbistatin were obtained from Tocris Bioscience, Y27632 from Calbiochem, nocodazole and DMSO from Sigma-Aldrich. For silencing, Smart pool ON-TARGETplus mouse LMNA siRNA, Arpc4 siRNA and Sun1 siRNA were purchased from Thermo Fisher Scientific. Micro-channels were coated with fibronectin (Sigma) or custom-made pLL(20)-g[3.5]-PEG(2)-Rhodamin.

Thiam H.R., Vargas P., Carpi N., Crespo C.L., Raab M., Terriac E., King M.C., Jacobelli J., Alberts A.S., Stradal T., Lennon-Dumenil A.M, & Piel M. (2016). Perinuclear Arp2/3-driven actin polymerization enables nuclear deformation to facilitate cell migration through complex environments. Nature Communications, 7, 10997.

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Immunofluorescence and Western Blot Analysis

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The primary antibodies used were: a) rabbit polyclonal – NMIIA (Sigma), phospho-NMIIA (Ser1943) (Cell Signaling Technology), giantin, Man-II, and Rab6a (Abcam); b) mouse monoclonal - Rab6a (Santa Cruz Biotechnology), β-actin (Sigma), Sar1a, giantin (Abcam); c) mouse polyclonal - ST3Gal1 (Abnova). The secondary antibodies (Jackson ImmunoResearch) were: a) HRP-conjugated donkey anti-rabbit and donkey anti-mouse for Western-blotting; b) donkey anti-mouse Alexa Fluor 488 and anti-rabbit Alexa Fluor 594 for immunofluorescence. Blebbistatin (Sigma) and latrunculin A (Tocris Bioscience) were dissolved in dimethyl sulfoxide (DMSO) immediately before use. Cells treated with a corresponding concentration of DMSO served as controls. The regular working concentrations were: Blebbistatin −25 μM for up to 24 h; latrunculin A −0.2 μM for up to 24 h. Active human Rab6a full length protein was obtained from Abcam.

Petrosyan A., Casey C.A, & Cheng P.W. (2016). The role of Rab6a and phosphorylation of non-muscle myosin IIA tailpiece in alcohol-induced Golgi disorganization. Scientific Reports, 6, 31962.

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Pharmacological Inhibition of Cell Mechanics

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Pharmacological inhibitors were added to cell media 10 minutes before time-lapse microscopy experiments. The concentrations used for the inhibitors are: 100 nM Latrunculin A (Tocris Bioscience, actin polymerization inhibitor), 70 μM NSC 23766 (Tocris Bioscience, Rac 1 inhibitor), 50 μM CK 666 (Sigma, Arp 2/3 inhibitor), 20 μM ML141 (Tocris, Cdc 42 GTPase inhibitor), 20 μM SMIFH2 (Sigma, formin inhibitor), 30 μM fascin-G2 (Xcess Biosciences Inc., fascin inhibitor), 50 μM Y-27632 (Sigma, ROCK inhibitor) 25 μM ML-7 (Tocris, myosin light chain kinase inhibitor), 2 μg/ml CD29 Monoclonal Antibody TS2/16 (Life Technologies, β1-integrin activator), and 5 μg/ml monoclonal β1-integrin-blocking antibody (Abcam, P5D2). For HT-1080 cell detachment studies, cells were seeded on fast and slow-relaxing gels and allowed to spread overnight. 10 μg/ml monoclonal β1-integrin-blocking antibody was added to media and live-cell imaging started. Images were acquired every 2 minutes for 12 hours.

Adebowale K., Gong Z., Hou J.C., Wisdom K.M., Garbett D., Lee H.P., Nam S., Meyer T., Odde D.J., Shenoy V.B, & Chaudhuri O. (2021). Enhanced substrate stress relaxation promotes filopodia-mediated cell migration. Nature materials, 20(9), 1290-1299.

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Notch and Yap Signaling Inhibition

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To inhibit Notch signaling, explants were treated with DAPT (Sigma - 25 μM unless indicated), LY-411575 (Sigma - 10μM unless indicated). To inhibit Yap signaling, explants were treated with latrunculin A (Tocris – 0.5μM or 0.012μM, 0.06μM, 0.3μM in Fig. 7M)). Controls with vehicle solvent were used.

Hubaud A., Regev I., Mahadevan L, & Pourquié O. (2017). Excitable dynamics and Yap-dependent mechanical cues drive the segmentation clock. Cell, 171(3), 668-682.e11.

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Permeabilizing C. elegans Embryos for Live Imaging

Cited 2 times

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L1 stage worms were plated on perm-1(RNAi) plates and incubated at 21°C for ~60 hr, to permeabilize the embryos by preventing normal eggshell formation (Carvalho et al., 2011 (link); Olson et al., 2012 (link)). Young adult worms were picked directly into Shelton’s growth medium (SGM, see Embryo mounting and microdissection above) and dissected to release early embryos. Four-cell stage embryos were then washed three times into SGM containing various concentrations of Latrunculin A (Tocris Bioscience, #3973, stored as a 2.5 mM stock in DMSO at −80°C), as well as 10 μM FM 4-64 (Thermo Fisher Scientific #T13320). Embryos were then transferred to a drop of the same growth medium on a No. 1.5 22 × 22 mm coverslip, and mounted with the ‘hanging drop’ method (Davies et al., 2017 (link)) using a SecureSeal spacer (Electron Microscopy Sciences, #70327–9S) to avoid changing the media composition or compressing the embryos. Successful eggshell permeablization was confirmed by the presence of the FM 4–64 dye on cell membranes in the first time point of the time lapse image series.

Davies T., Kim H.X., Romano Spica N., Lesea-Pringle B.J., Dumont J., Shirasu-Hiza M, & Canman J.C. (2018). Cell-intrinsic and -extrinsic mechanisms promote cell-type-specific cytokinetic diversity. eLife, 7, e36204.

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Molecular Signaling Pathways Characterization

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All restriction enzymes were from New England Biolabs. Chemicals were obtained from SIGMA-Aldrich unless otherwise specified. APV, W7, KN62, Glycyl-H-1152, and Latrunculin A were from TOCRIS Bioscience. DNA sequencing was performed at the Johns Hopkins University School of Medicine Sequencing Facility.

Araki Y., Zeng M., Zhang M, & Huganir R.L. (2015). Rapid Dispersion of SynGAP from Synaptic Spines Triggers AMPA Receptor Insertion and Spine Enlargement during LTP. Neuron, 85(1), 173-189.

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Actin Cytoskeleton Visualization

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Early log phase cells were fixed by diluting 37% formaldehyde to a final concentration of 3.7% and incubating at 25 C for 10 minutes. Cells were subsequently pelleted (800 g for 3 min) and resuspended in PBS containing 3.7% formaldehyde and incubated for 1 hour. Cells were subsequently washed three times in PBS prior to staining with Rhodamine-conjugated phalloidin diluted in PBS containing 0.1% Tween on ice (20 Units/ml, Invitrogen) and cells were washed two times prior to imaging. For actin depolymerisation, Latrunculin A (5 µM, Tocris) was added as indicated prior to fixation and cell staining.

Kennedy M.A., Gable K., Niewola-Staszkowska K., Abreu S., Johnston A., Harris L.J., Reggiori F., Loewith R., Dunn T., Bennett S.A, & Baetz K. (2014). A Neurotoxic Glycerophosphocholine Impacts PtdIns-4, 5-Bisphosphate and TORC2 Signaling by Altering Ceramide Biosynthesis in Yeast. PLoS Genetics, 10(1), e1004010.

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Inhibition of Integrin and Cytoskeletal Dynamics

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Cells were treated with BIO 5192 (20 nM, 45 min; R&D Systems) to inhibit the VLA-4 integrin; latrunculin A (1 µM, 1 h; Tocris Bioscience) to inhibit actin polymerization; Blebb (50 µM, 10 min; MedChemExpress) to inhibit nonmuscle myosin II; Y-27632 dihydrochloride (10 µM, 1 h; Tocris Bioscience) to inhibit ROCK signaling; LY (5 µM, 1 h; MedChemExpress), BAY (Copanlisib; 64 nM, 1 h), CAL-101 (Idelalisib; 50 nM, 1 h), MLN1117 (Serabelisib; 150 nM, 1 h), and TGX-221 (85 nM, 1 h) to inhibit PI3K; and MK (5 µM, 1 h; MedChemExpress) and GSK2141795 (1 µM, 1 h) to inhibit Akt activity. Vehicle solvents were used accordingly as controls.

Hao J., Zhou H., Nemes K., Yen D., Zhao W., Bramlett C., Wang B., Lu R, & Shen K. (2021). Membrane-bound SCF and VCAM-1 synergistically regulate the morphology of hematopoietic stem cells. The Journal of Cell Biology, 220(10), e202010118.

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HIV-1 Tat Protein Perturbation Assay

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Materials were obtained from the following sources: HIV-1 Tat (Clade B, 1-86 amino acids) was acquired from Prospec Tany TechnoGene Ltd. (Rehovot, Israel); Dulbecco's modified Eagle's medium (DMEM), fetal bovine serum, horse serum, fura-2-acetomethyl ester (Fura-2-AM), and glycine were obtained from Invitrogen (Carlsbad, CA). Phalloidin-Oleate, Y27632 (Trans-4-[(1R)-1-Aminoethyl]-N-4pyridinylcyclohexanecarboxamide dihydrochloride), and Cytochalasin D (zygosporium mansonii) were acquired from EMD Millipore (Billerica, MA); Rho Inhibitor I (exoenzyme C3 transferase) was obtained from Cytoskeleton, Inc. (Denver, CO); H1152 ((S)-(+)-2-Methyl-1-[(4-methyl-5-isoquinolinyl)sulfonyl]-hexahydro-1H-1,4-diazepine dihydrochloride), latrunculin A (4-[(1R,4Z,8E,10Z,12S,15R,17R)-17-Hydroxy-5,12-dimethyl-3-oxo-2,16-dioxabicyclo[13.3.1]nonadeca-4,8,10-trien-17-yl)-2-thiazolidinone), and jasplakinolide (Cyclo[(3R)-3-(4-hydroxyphenyl)-β-alanyl-(2S,4E,6R,8S)-8-hydroxy-2,4,6-trimethyl-4-nonenoyl-L-alanyl-2-bromo-N-methyl-D-tryptophyl]) were acquired from Tocris (Bristol, UK); ML-7 (1-(5-Iodonaphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine hydrochloride) and NMDA were obtained from Sigma-Aldrich (St. Louis, MO).

Krogh K.A., Lyddon E, & Thayer S.A. (2014). HIV-1 Tat activates a RhoA signaling pathway to reduce NMDA-evoked calcium responses in hippocampal neurons via an actin-dependent mechanism. Journal of neurochemistry, 132(3), 354-366.

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Inhibitors of Cellular Mechanics in Motility

Cited 4 times

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Inhibitors were implemented in invasive morphology and/or migration assays at concentrations as follows: 10 μM GM6001 (Millipore) or 100 μM marimastat (Tocris) to broadly inhibit MMPs; 70 μM NSC23766 to inhibit Rac1 (Tocris); 10 μM Y-27632 to inhibit ROCK (Sigma); 100 μM CK-666 to inhibit Arp 2/3 (Sigma); 1 μg mL⁻¹ monoclonal β1 integrin-blocking antibody (Abcam, P5D2); 50 μM Blebbistatin (Abcam); and 2.5 μM Latrunculin-a (Tocris). Vehicle-alone controls for these inhibitors were as follows: DMSO for GM6001, marimastat, CK-666, Blebbistatin, and Latrunculin-a; deionized water for NSC23766 and Y-27632; and IgG nonspecific antibody (Sigma, I5381) for β1 integrin-blocking antibody. All inhibitor concentrations followed those used in similar studies: GM6001^{6 (link),47 (link)}, marimastat^{48 (link)}, NSC23766^{15 (link)}, Y-27632^{42 (link)}, CK-666^{49 (link)}, β1 integrin-blocking antibody^{42 (link)}, Blebbistatin^{31 (link),50 (link)}, and Latrunculin-a^{50 (link)}. Drug concentrations were also verified in-house using gelatin degradation assays and traction force microscopy experiments (described elsewhere in Methods).

Wisdom K.M., Adebowale K., Chang J., Lee J.Y., Nam S., Desai R., Rossen N.S., Rafat M., West R.B., Hodgson L, & Chaudhuri O. (2018). Matrix mechanical plasticity regulates cancer cell migration through confining microenvironments. Nature Communications, 9, 4144.

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