| Cell line (Homo sapiens) |
Human osteosarcoma cells (U2OS) |
ATCC |
Cat# HTB-96, RRID:CVCL_0042
|
Authenticated through STR profiling |
| Cell line (Mus musculus)
|
Mouse embryonic fibroblasts (MEFs) |
Jiu et al., 2015
|
|
A gift from John Eriksson (University of Turku, Finland) |
| Recombinant DNA reagent |
eGFP-NMIIA (plasmid) |
Addgene, Jacobelli et al., 2009
|
RRID:Addgene_38297
|
A gift from Mathew Krummel (University of California, San Francisco) |
| Recombinant DNA reagent |
PA-GFP-Actin (plasmid) |
Addgene |
RRID:Addgene_57121
|
A gift from Michael Davidson |
| Recombinant DNA reagent |
LifeAct-mKate1.31 (plasmid) |
Addgene, Shcherbo et al., 2009
|
RRID:Addgene_54668
|
A gift from Michael Davidson |
| Recombinant DNA reagent |
mApple-NMIIA (plasmid) |
Addgene, Burnette et al., 2014
|
RRID:Addgene_54929
|
A gift from Michael Davidson |
| Recombinant DNA reagent |
Vinculin-mApple (plasmid) |
Addgene |
RRID:Addgene_54962
|
A gift from Michael Davidson |
| Recombinant DNA reagent |
α-actinin1-TagRFP-T (plasmid) |
Addgene |
RRID:Addgene_58033
|
A gift from Michael Davidson |
| Recombinant DNA reagent |
LifeAct-TagGFP2 (plasmid) |
Stefani et al., 2017
|
|
A gift from Emmanuel Lemichez (Pasteur Institute, France) |
| Recombinant DNA reagent |
Histone H2B-eGFP (plasmid) |
This paper |
|
A gift from Maria Vartiainen (University of Helsinki, Finland) |
| Recombinant DNA reagent |
Histone H2B-mCherry (plasmid) |
This paper |
|
A gift from Maria Vartiainen (University of Helsinki, Finland) |
| Recombinant DNA reagent |
eGFP-Paxillin (plasmid) |
This paper |
|
Gallus gallus paxillin ORF with T132S and M133L unattended mutations (pPL1277) |
| Recombinant DNA reagent |
miRFP670-Paxillin (plasmid) |
This paper |
|
eGFP in eGFP-Paxillin replaced with miRFP670-N1 (pPL1514) |
| Recombinant DNA reagent |
miRFP670-N1 (plasmid) |
Addgene, Shcherbakova et al., 2016
|
RRID:Addgene_79987
|
A gift from Vladislav Verkusha (Albet Einstein College of Medicine, NY) |
| Transfected construct (human) |
SMARTpoolON-TARGETplus siRNA to mDia1 |
Dharmacon |
Cat# L-010347-00-0005 |
Transfected using GeneSilencer |
| Transfected construct (human) |
SMARTpoolON-TARGETplus siRNA to mDia2 |
Dharmacon |
Cat# L-018997-00-0005
|
Transfected using GeneSilencer |
| Transfected construct (human) |
SMARTpoolON-TARGETplus siRNA to mDia3 |
Dharmacon |
Cat# L-012029-00-0005
|
Transfected using GeneSilencer |
| Transfected construct (human) |
negative (non-targeting) control siRNA |
Qiagen |
Cat# 1027281 |
Transfected using GeneSilencer |
| Antibody |
anti-vinculin (mouse monoclonal) |
Sigma Aldrich |
Cat# V9131, RRID:AB_477629
|
1:400 (IF) |
| Antibody |
anti-NMIIA (C-termini) (rabbit polyclonal) |
Biolegend |
Cat# 909801, RRID:AB_2565100
|
1:800 (IF) |
| Antibody |
anti-fibronectin (rabbit polyclonal) |
Sigma Aldrich |
Cat# F3648, RRID:AB_476976
|
1:400 (IF) a gift from Johanna Ivaska (University of Turku, Finland) |
| Antibody |
anti-Y118-paxillin (rabbit monoclonal) |
Cell Signaling Technology |
Cat# 2541, RRID:AB_2174466
|
1:50 (IF) a gift from Johanna Ivaska (University of Turku, Finland) |
| Antibody |
anti-human mDia1 (rabbit monoclonal) |
Abcam |
Cat# ab129167, RRID:AB_11143749
|
1:1000 (WB) |
| Antibody |
anti-human mDia2 (rabbit polyclonal) |
Proteintech |
Cat# 14342-1-AP, RRID:AB_2092930
|
1: 500 (WB)
|
| Antibody |
anti- human mDia3 (rabbit polyclonal) |
Sigma Aldrich |
Cat# HPA005647, RRID:AB_1078657
|
1:10,00 (WB) |
| Antibody |
anti-GAPDH (rabbit polyclonal) |
Sigma Aldrich |
Cat# G9545, RRID:AB_796208
|
1:10,000 (WB) |
| Antibody |
Anti-mouse Alexa Fluor 488 (goat polyclonal) |
Thermo Fisher Scientific |
Cat# A-11001, RRID:AB_2534069
|
1:200 (IF) |
| Antibody |
Anti-mouse Alexa Fluor 568 (goat polyclonal) |
Thermo Fisher Scientific |
Cat# A-11031, RRID:AB_144696
|
1:200 (IF) |
| Antibody |
Anti-rabbit Alexa Fluor Plus 647 (goat polyclonal) |
Thermo Fisher Scientific |
Cat# A32733, RRID:AB_2633282
|
1:400 (IF) |
| Antibody |
HRP-conjugated anti–rabbit (goat polyclonal) |
Thermo Fisher Scientific |
Cat# 32460, RRID:AB_1185567
|
1:1000 (WB) |
| Commercial assay or kit |
Lipofectamine 2000 |
Thermo Fisher Scientific |
Cat# 11668019 |
Plasmid transfection (3.5:1 DNA: Lipofectamine ratio) |
| Commercial assay or kit |
GeneSilencer |
Genlantis |
Cat# T500750 |
siRNA transfection |
| Commercial assay or kit |
jetPRIME |
Polyplus transfection |
Cat# 114–01 |
Plasmid transfection (TFM) |
| Commercial assay or kit |
NEBuilder |
New England Biolabs |
Cat# E5520S |
|
| Chemical compound, drug |
Para-amino Blebbistatin |
Optopharma |
Cat# DR-Am-89 |
50 µM working concentration |
| Chemical compound, drug |
SMIFH2 |
Sigma Aldrich |
Cat# S4826 |
12.5 µM working concentration |
| Chemical compound, drug |
CK-666 |
Sigma Aldrich |
Cat# SML0006 |
100 µM working concentration |
| Chemical compound, drug |
DMSO |
Sigma Aldrich |
Cat# D2650 |
Diluent/negative control |
| Software, algorithm |
Fiji |
Schindelin et al., 2012
|
RRID:SCR_002285
|
|
| Software, algorithm |
OriginPro |
OriginLab Corp. |
RRID:SCR_014212
|
|
| Software, algorithm |
NMII pulse quantification (macro) |
This paper. Modified from Kim and Davidson, 2011
|
|
Available at: https://github.com/UH-LMU/lmu-users/tree/master/jaakko/NMIIA_pulses
|
| Other |
Fibrinogen (from human plasma) |
Sigma Aldrich |
Cat# 341578 |
2.25 mg/ml working concentration |
| Other |
Thrombin (from human plasma) |
Sigma Aldrich |
Cat# T6884 |
1 U/ml working concentration |
| Other |
Fibronectin (human plasma) |
Sigma Aldrich |
Cat# FIBRP-RO |
10 µg/ml working concentration |
| Other |
Laminin |
Sigma Aldrich |
Cat# L2020 |
25 µg/ml working concentration |
| Other |
Collagen |
Sigma Aldrich |
Cat# C4243 |
500 µg/ml working concentration |
| Other |
poly-D-lysine |
Sigma Aldrich |
Cat# P0899 |
10 µg/ml working concentration |
| Other |
DAPI stain |
Thermo Fisher Scientific |
Cat# D1306 |
0.2 µM working concentration (IF) |
| Other |
Alexa Fluor 488-Phalloidin |
Thermo Fisher Scientific |
Cat# A12379 |
1:200 (IF) |
| Other |
Alexa Fluor 568-Phalloidin |
Thermo Fisher Scientific |
Cat# 12380 |
1:200 (IF) |
| Other |
Fluorescent microspheres. 0.2 µm diameter |
Invitrogen |
Cat# F8848 |
|
