Review



human osteosarcoma u2os  (ATCC)


Bioz Verified Symbol ATCC is a verified supplier
Bioz Manufacturer Symbol ATCC manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 99

    Structured Review

    ATCC human osteosarcoma u2os
    Human Osteosarcoma U2os, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 8550 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/human+u2os+osteosarcoma/U-2+OS/pm42881217-49-8-13
    Average 99 stars, based on 8550 article reviews
    human osteosarcoma u2os - by Bioz Stars, 2026-11
    99/100 stars

    Images

    Related Articles

    other:

    Article Title: A TRIM21-based bioPROTAC highlights the therapeutic benefit of HuR degradation.
    Article Snippet: Prior to in vitro mRNA transcription, pcGEMHE or pcDNA 3.1 constructs were linearised via PCR 5’-capped modified RNA was synthesised according to the manufacturer’s protocol using HiScribe T7 ARCA mRNA Kit (New England Biolabs) or HiScribe T7 High Yield RNA Synthesis Kit (New England Biolabs), CleanCap (Trilink) and 5- Methoxyuridine-5′-Triphosphate (5-moUTP) (40% final concentration) (Trilink) respectively.

    Article Title: A TRIM21-based bioPROTAC highlights the therapeutic benefit of HuR degradation
    Article Snippet: mRNA was purified using the MEGAclear kit (ThermoFisher Scientific) prior to quality control on the 2100 Bioanalyzer (Agilent).

    Cell Culture:

    Article Title: Identification of RNF114 as ADPr-Ub reader through non-hydrolysable ubiquitinated ADP-ribose
    Article Snippet: .. Briefly, human U2OS osteosarcoma (American Type Culture Collection (ATCC), HTB-96) cells were cultured in Dulbecco’s modified Eagle’s medium (Sigma-Aldrich) supplemented with 10% fetal bovine serum (FBS; Gibco) and penicillin-streptomycin (100 U/ml; Gibco) in a humidified atmosphere at 37 °C with 5% CO 2 . .. Cells were plated on an eight-well μ-slide glass bottom chamber slide (ibidi) and transfected with an expression plasmid for YFP-RNF114 WT, YFP-RNF114 UIM mutant (V220G,L221G,S224G), YFP-RNF114 ΔUIM (1-208), or ADPr binding mutants YFP-RNF114, Y186A, and C176A together with PaTagRFP-H2B using TransIT-LT1 Transfection Reagent (Mirus Bio), according to the manufacturer’s instructions.

    Article Title: Specificity and recognition of the ADP-ribosyl-ubiquitin modification in the DNA damage response.
    Article Snippet: .. Briefly, human U2OS osteosarcoma (American Type Culture Collection (ATCC HTB-96) WT, DTX2KO, DTX3LKO, [69] or HPF1KO [70] cells were cultured in Dulbecco’s modified Eagle’s medium (Sigma-Aldrich) supplemented with 10% fetal bovine serum (FBS; Gibco) and penicillin-streptomycin (100 U/ml; Gibco) in a humidified atmosphere at 37°C with 5% CO 2 . .. Cells were plated on an eight- well μ-slide glass bottom chamber slide (ibidi) and transfected with an expression plasmid for YFP-RNF114 WT, YFPRNF114 Znf2-3 + UIM (amino acid 140−228 of RNF114), RNF138, RNF166 or YFP-RNF166 ZnF2-3 + UIM (amino acids 149−237 of RNF166), mCherry-RNF114 or YFP-DTX2 using TransIT-LT1 Transfection Reagent (Mirus Bio), according to the manufacturer’s instructions.

    Article Title: Specificity and recognition of the ADP-ribosyl-ubiquitin modification in the DNA damage response
    Article Snippet: .. Briefly, human U2OS osteosarcoma (American Type Culture Collection (ATCC HTB-96) WT, DTX2KO, DTX3LKO, [ ] or HPF1KO [ ] cells were cultured in Dulbecco’s modified Eagle’s medium (Sigma-Aldrich) supplemented with 10% fetal bovine serum (FBS; Gibco) and penicillin-streptomycin (100 U/ml; Gibco) in a humidified atmosphere at 37°C with 5% CO 2 . .. Cells were plated on an eight-well μ-slide glass bottom chamber slide (ibidi) and transfected with an expression plasmid for YFP-RNF114 WT, YFP-RNF114 Znf2-3 + UIM (amino acid 140−228 of RNF114), RNF138, RNF166 or YFP-RNF166 ZnF2-3 + UIM (amino acids 149−237 of RNF166), mCherry-RNF114 or YFP-DTX2 using TransIT-LT1 Transfection Reagent (Mirus Bio), according to the manufacturer’s instructions.

    Article Title: Identification of RNF114 as ADPr-Ub reader through non-hydrolysable ubiquitinated ADP-ribose
    Article Snippet: .. Briefly, human U2OS osteosarcoma (American Type Culture Collection (ATCC), HTB-96) cells were cultured in Dulbecco’s modified Eagle’s medium (Sigma-Aldrich) supplemented with 10% fetal bovine serum (FBS; Gibco) and penicillin-streptomycin (100 U/ml; Gibco) in a humidified atmosphere at 37°C with 5% CO2. .. Cells were plated on an eight-well μ-slide glass bottom chamber slide (ibidi) and transfected with an expression plasmid for YFP-RNF114 WT, YFP-RNF114 UIM mutant (V220G,L221G,S224G), YFP-RNF114 ΔUIM (1-208), or YFP-RNF114 ZnF2+ZnF3 mutant C176A together with PaTagRFP-H2B using TransIT-LT1 Transfection Reagent (Mirus Bio), according to the manufacturer’s instructions.

    Modification:

    Article Title: Identification of RNF114 as ADPr-Ub reader through non-hydrolysable ubiquitinated ADP-ribose
    Article Snippet: .. Briefly, human U2OS osteosarcoma (American Type Culture Collection (ATCC), HTB-96) cells were cultured in Dulbecco’s modified Eagle’s medium (Sigma-Aldrich) supplemented with 10% fetal bovine serum (FBS; Gibco) and penicillin-streptomycin (100 U/ml; Gibco) in a humidified atmosphere at 37 °C with 5% CO 2 . .. Cells were plated on an eight-well μ-slide glass bottom chamber slide (ibidi) and transfected with an expression plasmid for YFP-RNF114 WT, YFP-RNF114 UIM mutant (V220G,L221G,S224G), YFP-RNF114 ΔUIM (1-208), or ADPr binding mutants YFP-RNF114, Y186A, and C176A together with PaTagRFP-H2B using TransIT-LT1 Transfection Reagent (Mirus Bio), according to the manufacturer’s instructions.

    Article Title: Specificity and recognition of the ADP-ribosyl-ubiquitin modification in the DNA damage response.
    Article Snippet: .. Briefly, human U2OS osteosarcoma (American Type Culture Collection (ATCC HTB-96) WT, DTX2KO, DTX3LKO, [69] or HPF1KO [70] cells were cultured in Dulbecco’s modified Eagle’s medium (Sigma-Aldrich) supplemented with 10% fetal bovine serum (FBS; Gibco) and penicillin-streptomycin (100 U/ml; Gibco) in a humidified atmosphere at 37°C with 5% CO 2 . .. Cells were plated on an eight- well μ-slide glass bottom chamber slide (ibidi) and transfected with an expression plasmid for YFP-RNF114 WT, YFPRNF114 Znf2-3 + UIM (amino acid 140−228 of RNF114), RNF138, RNF166 or YFP-RNF166 ZnF2-3 + UIM (amino acids 149−237 of RNF166), mCherry-RNF114 or YFP-DTX2 using TransIT-LT1 Transfection Reagent (Mirus Bio), according to the manufacturer’s instructions.

    Article Title: Specificity and recognition of the ADP-ribosyl-ubiquitin modification in the DNA damage response
    Article Snippet: .. Briefly, human U2OS osteosarcoma (American Type Culture Collection (ATCC HTB-96) WT, DTX2KO, DTX3LKO, [ ] or HPF1KO [ ] cells were cultured in Dulbecco’s modified Eagle’s medium (Sigma-Aldrich) supplemented with 10% fetal bovine serum (FBS; Gibco) and penicillin-streptomycin (100 U/ml; Gibco) in a humidified atmosphere at 37°C with 5% CO 2 . .. Cells were plated on an eight-well μ-slide glass bottom chamber slide (ibidi) and transfected with an expression plasmid for YFP-RNF114 WT, YFP-RNF114 Znf2-3 + UIM (amino acid 140−228 of RNF114), RNF138, RNF166 or YFP-RNF166 ZnF2-3 + UIM (amino acids 149−237 of RNF166), mCherry-RNF114 or YFP-DTX2 using TransIT-LT1 Transfection Reagent (Mirus Bio), according to the manufacturer’s instructions.

    Article Title: Identification of RNF114 as ADPr-Ub reader through non-hydrolysable ubiquitinated ADP-ribose
    Article Snippet: .. Briefly, human U2OS osteosarcoma (American Type Culture Collection (ATCC), HTB-96) cells were cultured in Dulbecco’s modified Eagle’s medium (Sigma-Aldrich) supplemented with 10% fetal bovine serum (FBS; Gibco) and penicillin-streptomycin (100 U/ml; Gibco) in a humidified atmosphere at 37°C with 5% CO2. .. Cells were plated on an eight-well μ-slide glass bottom chamber slide (ibidi) and transfected with an expression plasmid for YFP-RNF114 WT, YFP-RNF114 UIM mutant (V220G,L221G,S224G), YFP-RNF114 ΔUIM (1-208), or YFP-RNF114 ZnF2+ZnF3 mutant C176A together with PaTagRFP-H2B using TransIT-LT1 Transfection Reagent (Mirus Bio), according to the manufacturer’s instructions.

    Multiple Displacement Amplification:

    Article Title: VGLL3 modulates chemosensitivity through promoting DNA double-strand break repair
    Article Snippet: .. Human U2OS (osteosarcoma), MDA-MB-231 (triple-negative breast cancer), MCF7, SK-OV-3 (ovarian cancer), A375 (melanoma), and 293T cells were purchased from the American Type Culture Collection (Rockville, MD, USA). ..



    Similar Products

    99
    ATCC human osteosarcoma u2os
    Human Osteosarcoma U2os, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/human+u2os+osteosarcoma/U-2+OS/pm42881217-49-8-13
    Average 99 stars, based on 1 article reviews
    human osteosarcoma u2os - by Bioz Stars, 2026-11
    99/100 stars
      Buy from Supplier

    95
    ATCC human osteosarcoma u2os cells atcc htb 86
    Human Osteosarcoma U2os Cells Atcc Htb 86, supplied by ATCC, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/human+u2os+osteosarcoma/SK-ES-1/pm42881257-286-15-19
    Average 95 stars, based on 1 article reviews
    human osteosarcoma u2os cells atcc htb 86 - by Bioz Stars, 2026-11
    95/100 stars
      Buy from Supplier

    99
    ATCC u2os human osteosarcoma cell lines
    Integrated single‐cell landscape of human <t>osteosarcoma.</t> (A) Harmony‐integrated UMAP of 39,063 quality‐controlled cells colored by global cluster. (B) The same embedding colored by patient, showing cross‐patient mixing after integration. (C) Reviewed major‐cell‐type annotations displayed on the integrated UMAP; labels mark lineage centroids. (D) Relative abundance of major cell types in each patient. Colors are consistent between panels C and D.
    U2os Human Osteosarcoma Cell Lines, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/human+u2os+osteosarcoma/U-2+OS/pmc13669654-131-2-11
    Average 99 stars, based on 1 article reviews
    u2os human osteosarcoma cell lines - by Bioz Stars, 2026-11
    99/100 stars
      Buy from Supplier

    99
    ATCC male origin atcc ccl 247 u2os human osteosarcoma cells
    Integrated single‐cell landscape of human <t>osteosarcoma.</t> (A) Harmony‐integrated UMAP of 39,063 quality‐controlled cells colored by global cluster. (B) The same embedding colored by patient, showing cross‐patient mixing after integration. (C) Reviewed major‐cell‐type annotations displayed on the integrated UMAP; labels mark lineage centroids. (D) Relative abundance of major cell types in each patient. Colors are consistent between panels C and D.
    Male Origin Atcc Ccl 247 U2os Human Osteosarcoma Cells, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/human+u2os+osteosarcoma/HCT+116/pm42831021-387-82-84
    Average 99 stars, based on 1 article reviews
    male origin atcc ccl 247 u2os human osteosarcoma cells - by Bioz Stars, 2026-11
    99/100 stars
      Buy from Supplier

    99
    ATCC u2os human osteosarcoma cell line
    Integrated single‐cell landscape of human <t>osteosarcoma.</t> (A) Harmony‐integrated UMAP of 39,063 quality‐controlled cells colored by global cluster. (B) The same embedding colored by patient, showing cross‐patient mixing after integration. (C) Reviewed major‐cell‐type annotations displayed on the integrated UMAP; labels mark lineage centroids. (D) Relative abundance of major cell types in each patient. Colors are consistent between panels C and D.
    U2os Human Osteosarcoma Cell Line, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/human+u2os+osteosarcoma/U-2+OS/10__1038_slash_s41467___026___77989___w-300-10-15
    Average 99 stars, based on 1 article reviews
    u2os human osteosarcoma cell line - by Bioz Stars, 2026-11
    99/100 stars
      Buy from Supplier

    86
    Procell Inc human osteosarcoma cell lines u2os
    Integrated single‐cell landscape of human <t>osteosarcoma.</t> (A) Harmony‐integrated UMAP of 39,063 quality‐controlled cells colored by global cluster. (B) The same embedding colored by patient, showing cross‐patient mixing after integration. (C) Reviewed major‐cell‐type annotations displayed on the integrated UMAP; labels mark lineage centroids. (D) Relative abundance of major cell types in each patient. Colors are consistent between panels C and D.
    Human Osteosarcoma Cell Lines U2os, supplied by Procell Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/human+u2os+osteosarcoma/u2os/pmc13273694-40-0-12
    Average 86 stars, based on 1 article reviews
    human osteosarcoma cell lines u2os - by Bioz Stars, 2026-11
    86/100 stars
      Buy from Supplier

    86
    Servicebio Inc human osteosarcoma cell lines u2os
    Integrated single‐cell landscape of human <t>osteosarcoma.</t> (A) Harmony‐integrated UMAP of 39,063 quality‐controlled cells colored by global cluster. (B) The same embedding colored by patient, showing cross‐patient mixing after integration. (C) Reviewed major‐cell‐type annotations displayed on the integrated UMAP; labels mark lineage centroids. (D) Relative abundance of major cell types in each patient. Colors are consistent between panels C and D.
    Human Osteosarcoma Cell Lines U2os, supplied by Servicebio Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/human+u2os+osteosarcoma/1+a+cancer+cell+endometrial+hec+lines/pm42280476-57-1-21
    Average 86 stars, based on 1 article reviews
    human osteosarcoma cell lines u2os - by Bioz Stars, 2026-11
    86/100 stars
      Buy from Supplier

    99
    ATCC human u2os osteosarcoma cells
    A) Schematic for domain structures of Ect2 wild-type (Ect2-wt), N-terminally truncated active (ΔN-Ect2) and the catalytically inactive variant with PVQR->AAAA (564-567) substitutions (ΔN-Ect2-DHmut). B) Representative maximum intensity projections of 3D-SIM images of <t>U2OS</t> cells expressing EGFP control or the indicated EGFP-tagged Ect2 constructs. Upper panels show EGFP or EGFP-tagged Ect2 localization, middle panels display F-actin stained with phalloidin, and lower panels present magnified views of the regions depicted in the actin images. Scale bar: 10 µm. n = 7-15 cells. C) Quantification of mean phalloidin fluorescence intensity in wide-field images of cells expressing the indicated constructs. Cells were stained with WGA to visualize cell morphology and with phalloidin to label F-actin. Transfected cells were identified EGFP fluorescence. F-actin intensity was quantified by automated image analysis as described in the Methods. Data represent n ≥ 276 cells from 4 independent experiments. Statistical analysis was performed using one-way ANOVA followed by Tukey’s post hoc test. Bars indicate mean ± SEM. D and H) Representative TIRF images of the Rho activity sensor (mCherry-Rhotekin RBD) co-expressing either EGFP control, constitutively active Ect2 (EGFP-ΔN-Ect2) (D) or constitutively active GEF-H1 C53R (H) . Two phenotypes induced by active Ect2 are shown in (D) : (top) reduced pulsatory Rho sensor signal dynamics and (bottom) peripheral enrichment of Rho sensor signal with slow circumferential movement (white arrow). Frame rate: 3 frames/min, scale bar, 20 µm. E and F) Percent cells with peripheral Rho sensor enrichment (E) and the frequency of Rho sensor pulses in the central cell region (F) ; n ≥ 32 cells from 4 independent experiments. (G) Average pulse frequency of the Rho activity sensor signal upon co-expression of EGFP-control, active EGFP-ΔN-Ect2 or a mutant which cannot bind to active Rho (EGFP-ΔN-Ect2-RBmut); n ≥ 25 cells from 3 independent experiments. Bars indicate mean ± SEM.
    Human U2os Osteosarcoma Cells, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/human+u2os+osteosarcoma/U-2+OS/bio_rxiv__64898__2026__06__03__729549-128-0-5
    Average 99 stars, based on 1 article reviews
    human u2os osteosarcoma cells - by Bioz Stars, 2026-11
    99/100 stars
      Buy from Supplier

    99
    ATCC female human osteosarcoma cells u2os cells
    A) Schematic for domain structures of Ect2 wild-type (Ect2-wt), N-terminally truncated active (ΔN-Ect2) and the catalytically inactive variant with PVQR->AAAA (564-567) substitutions (ΔN-Ect2-DHmut). B) Representative maximum intensity projections of 3D-SIM images of <t>U2OS</t> cells expressing EGFP control or the indicated EGFP-tagged Ect2 constructs. Upper panels show EGFP or EGFP-tagged Ect2 localization, middle panels display F-actin stained with phalloidin, and lower panels present magnified views of the regions depicted in the actin images. Scale bar: 10 µm. n = 7-15 cells. C) Quantification of mean phalloidin fluorescence intensity in wide-field images of cells expressing the indicated constructs. Cells were stained with WGA to visualize cell morphology and with phalloidin to label F-actin. Transfected cells were identified EGFP fluorescence. F-actin intensity was quantified by automated image analysis as described in the Methods. Data represent n ≥ 276 cells from 4 independent experiments. Statistical analysis was performed using one-way ANOVA followed by Tukey’s post hoc test. Bars indicate mean ± SEM. D and H) Representative TIRF images of the Rho activity sensor (mCherry-Rhotekin RBD) co-expressing either EGFP control, constitutively active Ect2 (EGFP-ΔN-Ect2) (D) or constitutively active GEF-H1 C53R (H) . Two phenotypes induced by active Ect2 are shown in (D) : (top) reduced pulsatory Rho sensor signal dynamics and (bottom) peripheral enrichment of Rho sensor signal with slow circumferential movement (white arrow). Frame rate: 3 frames/min, scale bar, 20 µm. E and F) Percent cells with peripheral Rho sensor enrichment (E) and the frequency of Rho sensor pulses in the central cell region (F) ; n ≥ 32 cells from 4 independent experiments. (G) Average pulse frequency of the Rho activity sensor signal upon co-expression of EGFP-control, active EGFP-ΔN-Ect2 or a mutant which cannot bind to active Rho (EGFP-ΔN-Ect2-RBmut); n ≥ 25 cells from 3 independent experiments. Bars indicate mean ± SEM.
    Female Human Osteosarcoma Cells U2os Cells, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/human+u2os+osteosarcoma/U-2+OS/pm42214329-240-0-6
    Average 99 stars, based on 1 article reviews
    female human osteosarcoma cells u2os cells - by Bioz Stars, 2026-11
    99/100 stars
      Buy from Supplier

    Image Search Results


    Integrated single‐cell landscape of human osteosarcoma. (A) Harmony‐integrated UMAP of 39,063 quality‐controlled cells colored by global cluster. (B) The same embedding colored by patient, showing cross‐patient mixing after integration. (C) Reviewed major‐cell‐type annotations displayed on the integrated UMAP; labels mark lineage centroids. (D) Relative abundance of major cell types in each patient. Colors are consistent between panels C and D.

    Journal: International Journal of Genomics

    Article Title: Integrative Single‐Cell and Bulk Transcriptomics Identifies a Proliferative Malignant State and Supports a Functional Contribution of TROAP in Osteosarcoma

    doi: 10.1155/ijog/1407947

    Figure Lengend Snippet: Integrated single‐cell landscape of human osteosarcoma. (A) Harmony‐integrated UMAP of 39,063 quality‐controlled cells colored by global cluster. (B) The same embedding colored by patient, showing cross‐patient mixing after integration. (C) Reviewed major‐cell‐type annotations displayed on the integrated UMAP; labels mark lineage centroids. (D) Relative abundance of major cell types in each patient. Colors are consistent between panels C and D.

    Article Snippet: HOS and U2OS human osteosarcoma cell lines were obtained from the American Type Culture Collection (ATCC; Manassas, Virginia, United States; HOS, CRL‐1543; U2OS, HTB‐96).

    Techniques: Single Cell

    TROAP depletion reduces clonogenic growth, overall wound closure, migration, and invasion in osteosarcoma cells. (A) Immunoblot analysis of TROAP in HOS and U2OS cells transfected with siNC, siTROAP‐1, or siTROAP‐2; beta‐actin is the loading control. (B) Colony‐formation images and colony counts. (C) Scratch‐wound images at 0 and 24 h and overall wound closure. Because the assay was not proliferation‐matched, it is not interpreted as direct migration evidence. (D) Transwell migration and invasion images and cell counts. Data are mean + /−standard deviation from three independent biological experiments. One‐way ANOVA with Dunnett′s test compared each siTROAP group with siNC; ∗∗∗ p < 0.001.

    Journal: International Journal of Genomics

    Article Title: Integrative Single‐Cell and Bulk Transcriptomics Identifies a Proliferative Malignant State and Supports a Functional Contribution of TROAP in Osteosarcoma

    doi: 10.1155/ijog/1407947

    Figure Lengend Snippet: TROAP depletion reduces clonogenic growth, overall wound closure, migration, and invasion in osteosarcoma cells. (A) Immunoblot analysis of TROAP in HOS and U2OS cells transfected with siNC, siTROAP‐1, or siTROAP‐2; beta‐actin is the loading control. (B) Colony‐formation images and colony counts. (C) Scratch‐wound images at 0 and 24 h and overall wound closure. Because the assay was not proliferation‐matched, it is not interpreted as direct migration evidence. (D) Transwell migration and invasion images and cell counts. Data are mean + /−standard deviation from three independent biological experiments. One‐way ANOVA with Dunnett′s test compared each siTROAP group with siNC; ∗∗∗ p < 0.001.

    Article Snippet: HOS and U2OS human osteosarcoma cell lines were obtained from the American Type Culture Collection (ATCC; Manassas, Virginia, United States; HOS, CRL‐1543; U2OS, HTB‐96).

    Techniques: Migration, Western Blot, Analysis, Transfection, Control, Colony Assay, Standard Deviation

    A) Schematic for domain structures of Ect2 wild-type (Ect2-wt), N-terminally truncated active (ΔN-Ect2) and the catalytically inactive variant with PVQR->AAAA (564-567) substitutions (ΔN-Ect2-DHmut). B) Representative maximum intensity projections of 3D-SIM images of U2OS cells expressing EGFP control or the indicated EGFP-tagged Ect2 constructs. Upper panels show EGFP or EGFP-tagged Ect2 localization, middle panels display F-actin stained with phalloidin, and lower panels present magnified views of the regions depicted in the actin images. Scale bar: 10 µm. n = 7-15 cells. C) Quantification of mean phalloidin fluorescence intensity in wide-field images of cells expressing the indicated constructs. Cells were stained with WGA to visualize cell morphology and with phalloidin to label F-actin. Transfected cells were identified EGFP fluorescence. F-actin intensity was quantified by automated image analysis as described in the Methods. Data represent n ≥ 276 cells from 4 independent experiments. Statistical analysis was performed using one-way ANOVA followed by Tukey’s post hoc test. Bars indicate mean ± SEM. D and H) Representative TIRF images of the Rho activity sensor (mCherry-Rhotekin RBD) co-expressing either EGFP control, constitutively active Ect2 (EGFP-ΔN-Ect2) (D) or constitutively active GEF-H1 C53R (H) . Two phenotypes induced by active Ect2 are shown in (D) : (top) reduced pulsatory Rho sensor signal dynamics and (bottom) peripheral enrichment of Rho sensor signal with slow circumferential movement (white arrow). Frame rate: 3 frames/min, scale bar, 20 µm. E and F) Percent cells with peripheral Rho sensor enrichment (E) and the frequency of Rho sensor pulses in the central cell region (F) ; n ≥ 32 cells from 4 independent experiments. (G) Average pulse frequency of the Rho activity sensor signal upon co-expression of EGFP-control, active EGFP-ΔN-Ect2 or a mutant which cannot bind to active Rho (EGFP-ΔN-Ect2-RBmut); n ≥ 25 cells from 3 independent experiments. Bars indicate mean ± SEM.

    Journal: bioRxiv

    Article Title: Constitutive plasma membrane interaction of active Rho GEF Ect2 inhibits cortex contraction pulses

    doi: 10.64898/2026.06.03.729549

    Figure Lengend Snippet: A) Schematic for domain structures of Ect2 wild-type (Ect2-wt), N-terminally truncated active (ΔN-Ect2) and the catalytically inactive variant with PVQR->AAAA (564-567) substitutions (ΔN-Ect2-DHmut). B) Representative maximum intensity projections of 3D-SIM images of U2OS cells expressing EGFP control or the indicated EGFP-tagged Ect2 constructs. Upper panels show EGFP or EGFP-tagged Ect2 localization, middle panels display F-actin stained with phalloidin, and lower panels present magnified views of the regions depicted in the actin images. Scale bar: 10 µm. n = 7-15 cells. C) Quantification of mean phalloidin fluorescence intensity in wide-field images of cells expressing the indicated constructs. Cells were stained with WGA to visualize cell morphology and with phalloidin to label F-actin. Transfected cells were identified EGFP fluorescence. F-actin intensity was quantified by automated image analysis as described in the Methods. Data represent n ≥ 276 cells from 4 independent experiments. Statistical analysis was performed using one-way ANOVA followed by Tukey’s post hoc test. Bars indicate mean ± SEM. D and H) Representative TIRF images of the Rho activity sensor (mCherry-Rhotekin RBD) co-expressing either EGFP control, constitutively active Ect2 (EGFP-ΔN-Ect2) (D) or constitutively active GEF-H1 C53R (H) . Two phenotypes induced by active Ect2 are shown in (D) : (top) reduced pulsatory Rho sensor signal dynamics and (bottom) peripheral enrichment of Rho sensor signal with slow circumferential movement (white arrow). Frame rate: 3 frames/min, scale bar, 20 µm. E and F) Percent cells with peripheral Rho sensor enrichment (E) and the frequency of Rho sensor pulses in the central cell region (F) ; n ≥ 32 cells from 4 independent experiments. (G) Average pulse frequency of the Rho activity sensor signal upon co-expression of EGFP-control, active EGFP-ΔN-Ect2 or a mutant which cannot bind to active Rho (EGFP-ΔN-Ect2-RBmut); n ≥ 25 cells from 3 independent experiments. Bars indicate mean ± SEM.

    Article Snippet: Human U2OS osteosarcoma cells (HTB-96; ATCC) were maintained at 37 °C and 5 % CO 2 humidified atmosphere using standard cell culture techniques (DMEM + GlutaMAXTM medium, 10 % FBS, Life technologies; Gibco).

    Techniques: Variant Assay, Expressing, Control, Construct, Staining, Fluorescence, Transfection, Activity Assay, Mutagenesis

    Background-corrected average EGFP intensity in U2OS cells expressing EGFP control (black dots) or EGFP-ΔN-Ect2 (green dots), plotted against the normalized mean pulse frequency of the RBD sensor. Each dot represents a single cell. Data are from n ≥52 cells across three independent experiments.

    Journal: bioRxiv

    Article Title: Constitutive plasma membrane interaction of active Rho GEF Ect2 inhibits cortex contraction pulses

    doi: 10.64898/2026.06.03.729549

    Figure Lengend Snippet: Background-corrected average EGFP intensity in U2OS cells expressing EGFP control (black dots) or EGFP-ΔN-Ect2 (green dots), plotted against the normalized mean pulse frequency of the RBD sensor. Each dot represents a single cell. Data are from n ≥52 cells across three independent experiments.

    Article Snippet: Human U2OS osteosarcoma cells (HTB-96; ATCC) were maintained at 37 °C and 5 % CO 2 humidified atmosphere using standard cell culture techniques (DMEM + GlutaMAXTM medium, 10 % FBS, Life technologies; Gibco).

    Techniques: Expressing, Control, Single Cell

    A) Representative spinning disk confocal images of fixed U2OS cells transfected with either EGFP control, active Ect2 (EGFP-ΔN-Ect2), or a variant lacking the C-terminal PBC region (EGFP-ΔN-Ect2-ΔPBC). A schematic of the constructs is shown in . Upper panels: single central z-plane, lower panels: side views show orthogonal (x-z) projections of the z-stack along the line indicated in the corresponding upper panels. Scale bars: 10 µm (xy), 5 µm (z), n=50-56 cells from 3 independent experiments. B) Average pulse frequency of the Rho activity sensor signal (mCherry-Rhotekin-RBD) in cells expressing the indicated Ect2 variants. Red dots mark cells that generate fast high-amplitude pulses. n ≥ 51 cells from 3 independent experiments. using one-way ANOVA followed by Tukey’s post hoc test. Bars indicate mean ± SEM. C, E, G) Representative TIRF images of cells expressing the Rho activity sensor (right) and the indicated constructs (left), respectively. Frame rate: 3 frames/min, scale bar = 20 µm. D, F, H) Mean normalized intensity values of the Rho sensor signal in the corresponding cell regions (white boxes) in (C, E, F) . I) Proposed interplay between Ect2 and pulsatory Rho contraction signal network dynamics. Left: Pulsatory Rho contraction depends on a positive feedback loop in which Lbc-GEFs are recruited to the plasma membrane through binding to active Rho (right angled blue arrow). Curved red arrow points to enzymatic activation. Middle: In contrast, Ect2 is constitutively associated with the plasma membrane, independently of active Rho . Increased local concentration of Ect2 may result in a higher effective on-rate, thereby conferring a kinetic advantage that can outcompete Lbc-GEFs and thereby suppress Rho activity pulses. Curved red arrows illustrate enzymatic activation and allosteric activation. Right: Loss of the C-terminal PBC reduces Ect2 plasma membrane association, shifting its properties towards a Lbc-GEF-like phenotype, that is based on plasma membrane recruitment to active Rho, and that stimulates Rho activity pulses.

    Journal: bioRxiv

    Article Title: Constitutive plasma membrane interaction of active Rho GEF Ect2 inhibits cortex contraction pulses

    doi: 10.64898/2026.06.03.729549

    Figure Lengend Snippet: A) Representative spinning disk confocal images of fixed U2OS cells transfected with either EGFP control, active Ect2 (EGFP-ΔN-Ect2), or a variant lacking the C-terminal PBC region (EGFP-ΔN-Ect2-ΔPBC). A schematic of the constructs is shown in . Upper panels: single central z-plane, lower panels: side views show orthogonal (x-z) projections of the z-stack along the line indicated in the corresponding upper panels. Scale bars: 10 µm (xy), 5 µm (z), n=50-56 cells from 3 independent experiments. B) Average pulse frequency of the Rho activity sensor signal (mCherry-Rhotekin-RBD) in cells expressing the indicated Ect2 variants. Red dots mark cells that generate fast high-amplitude pulses. n ≥ 51 cells from 3 independent experiments. using one-way ANOVA followed by Tukey’s post hoc test. Bars indicate mean ± SEM. C, E, G) Representative TIRF images of cells expressing the Rho activity sensor (right) and the indicated constructs (left), respectively. Frame rate: 3 frames/min, scale bar = 20 µm. D, F, H) Mean normalized intensity values of the Rho sensor signal in the corresponding cell regions (white boxes) in (C, E, F) . I) Proposed interplay between Ect2 and pulsatory Rho contraction signal network dynamics. Left: Pulsatory Rho contraction depends on a positive feedback loop in which Lbc-GEFs are recruited to the plasma membrane through binding to active Rho (right angled blue arrow). Curved red arrow points to enzymatic activation. Middle: In contrast, Ect2 is constitutively associated with the plasma membrane, independently of active Rho . Increased local concentration of Ect2 may result in a higher effective on-rate, thereby conferring a kinetic advantage that can outcompete Lbc-GEFs and thereby suppress Rho activity pulses. Curved red arrows illustrate enzymatic activation and allosteric activation. Right: Loss of the C-terminal PBC reduces Ect2 plasma membrane association, shifting its properties towards a Lbc-GEF-like phenotype, that is based on plasma membrane recruitment to active Rho, and that stimulates Rho activity pulses.

    Article Snippet: Human U2OS osteosarcoma cells (HTB-96; ATCC) were maintained at 37 °C and 5 % CO 2 humidified atmosphere using standard cell culture techniques (DMEM + GlutaMAXTM medium, 10 % FBS, Life technologies; Gibco).

    Techniques: Transfection, Control, Variant Assay, Construct, Activity Assay, Expressing, Clinical Proteomics, Membrane, Binding Assay, Activation Assay, Concentration Assay