Journal: bioRxiv

Article Title: Actomyosin, vimentin and LINC complex pull on osteosarcoma nuclei to deform on micropillar topography

doi: 10.1101/822445

Figure Lengend Snippet: Total protein gel images for KD of lamin A, SUN1, SUN2, nesprin 2 and vimentin used for protein normalization and corresponding immunoblots.

Article Snippet: Vimentin siRNA was used at a 40pmol concentration (sc-29522, Santa Cruz Biotechnology, USA).

Techniques: Western Blot

Journal: bioRxiv

Article Title: Actomyosin, vimentin and LINC complex pull on osteosarcoma nuclei to deform on micropillar topography

doi: 10.1101/822445

Figure Lengend Snippet: Nucleotide sequences of pairs of primers used for qPCR experiments. “F” and “R” indicate forward and reverse primers respectively. Emg1 (18S methyltransferase), Lmna (lamin A/C), Lmnb1 (lamin B1), Sun1 (Sad1 and UNC84 domain containing 1), Sun2 (Sad1 and UNC84 domain containing 2), Syne2 (nesprin 2), vim (vimentin).

Article Snippet: Vimentin siRNA was used at a 40pmol concentration (sc-29522, Santa Cruz Biotechnology, USA).

Techniques:

Journal: bioRxiv

Article Title: Actomyosin, vimentin and LINC complex pull on osteosarcoma nuclei to deform on micropillar topography

doi: 10.1101/822445

Figure Lengend Snippet: a) SaOs-2 deformation kinetics on pillars. Orthogonal images show cells expressing actin-RFP (red) and histone H2B-GFP (green) seeded on micropillars at different time points during cell adhesion (for video see supplementary movie 1). b) Quantification of nuclear deformation. (For a description of the analysis see ). The mean value is indicated by a red line. P-values compared to initial cell adhesion on pillars at 2 hours. (n=150/time point, 3 independent experiments.) c) FA localization and nuclear deformation on pillars, z-stack projection of SaOs-2 cells (actin-green; paxillin-red; nucleus DNA-blue) at different time points. Subsets represent the average number of FAs at different heights (see figure on left). (n=45/time point, 3 independent experiments). d) Immunostaining (vimentin & β-tubulin) and phalloidin (actin) labelling showing cytoskeleton elements in grey. e) Schematic summary of the cytoskeletal organization after nuclear deformation on pillars. All error bars represent ±SD.

Article Snippet: Vimentin siRNA was used at a 40pmol concentration (sc-29522, Santa Cruz Biotechnology, USA).

Techniques: Expressing, Immunostaining

Journal: bioRxiv

Article Title: Actomyosin, vimentin and LINC complex pull on osteosarcoma nuclei to deform on micropillar topography

doi: 10.1101/822445

Figure Lengend Snippet: a) Immunostaining of SaOs-2 cells on micropillars 24 hrs after drug treatments. Z stack projection of cells immunostained with actin (green) and nucleus (magenta); myosin IIA (green); vimentin (green), β-tubulin (green). b) Quantification of nuclear deformation of SaOs-2 6 hours after drug treatment (n=160/drug treatment) with mean (red line). c) RT-qPCR was used to access vimentin expression in knockdown and non-target control (non-target RNA) cells (grey) normalised to emg1 gene expression, after 48 hours. d) Western blot was used to access vimentin protein levels after 48 hrs in knockdown and control cells. e) Quantification of nuclear deformation in vimentin knockdown and non-target control cells, after 48 hours, (n=150 nuclei/siRNA treatment). All experiments were performed independently three times.

Article Snippet: Vimentin siRNA was used at a 40pmol concentration (sc-29522, Santa Cruz Biotechnology, USA).

Techniques: Immunostaining, Quantitative RT-PCR, Expressing, Western Blot

Journal: bioRxiv

Article Title: Actomyosin, vimentin and LINC complex pull on osteosarcoma nuclei to deform on micropillar topography

doi: 10.1101/822445

Figure Lengend Snippet: Actomyosin pulling force, coupled to FAs and LINC complexes, drives cell-nuclear deformation on pillar topography. In this system, vimentin IF also assist in nuclear deformation.

Article Snippet: Vimentin siRNA was used at a 40pmol concentration (sc-29522, Santa Cruz Biotechnology, USA).

Techniques: