CiteAb

Fig.1.E showing Western Blotting from the publication: Integrin-Mediated Adhesion Promotes Centrosome Separation in Early Mitosis.

Fig.4.F showing Western Blotting from the publication: Integrin-Mediated Adhesion Promotes Centrosome Separation in Early Mitosis.

Fig.6.A showing Western Blotting from the publication: Integrin-Mediated Adhesion Promotes Centrosome Separation in Early Mitosis.

Fig.1.A showing Western Blotting in a Homo sapiens (Human) sample from the publication: Cortactin Is Required for Efficient FAK, Src and Abl Tyrosine Kinase Activation and Phosphorylation of Helicobacter pylori CagA.

Fig.6.E showing Western Blotting from the publication: Uncovering the signaling landscape controlling breast cancer cell migration identifies novel metastasis driver genes.

Fig.2.B showing Western Blotting in a Homo sapiens (Human) sample from the publication: Src-mediated regulation of the PI3K pathway in advanced papillary and anaplastic thyroid cancer.

Fig.3.A showing Western Blotting in a Homo sapiens (Human) sample from the publication: Src-mediated regulation of the PI3K pathway in advanced papillary and anaplastic thyroid cancer.

Fig.6.A showing Western Blotting in a Homo sapiens (Human) sample from the publication: Src-mediated regulation of the PI3K pathway in advanced papillary and anaplastic thyroid cancer.

Fig.4.C showing Western Blotting in a Homo sapiens (Human) sample from the publication: Src-mediated regulation of the PI3K pathway in advanced papillary and anaplastic thyroid cancer.

Fig.4.C showing Western Blotting in a Homo sapiens (Human) sample from the publication: Sustained activation of the AKT/mTOR and MAP kinase pathways mediate resistance to the Src inhibitor, dasatinib, in thyroid cancer.

Fig.4.D showing Western Blotting in a Homo sapiens (Human) sample from the publication: Sustained activation of the AKT/mTOR and MAP kinase pathways mediate resistance to the Src inhibitor, dasatinib, in thyroid cancer.

Fig.3.B showing Western Blotting in a Homo sapiens (Human) sample from the publication: Sustained activation of the AKT/mTOR and MAP kinase pathways mediate resistance to the Src inhibitor, dasatinib, in thyroid cancer.

Fig.3.C showing Western Blotting in a Homo sapiens (Human) sample from the publication: Sustained activation of the AKT/mTOR and MAP kinase pathways mediate resistance to the Src inhibitor, dasatinib, in thyroid cancer.

Fig.1.D showing Western Blotting in a Homo sapiens (Human) sample from the publication: Sustained activation of the AKT/mTOR and MAP kinase pathways mediate resistance to the Src inhibitor, dasatinib, in thyroid cancer.

Fig.4.C showing Western Blotting from the publication: Dominant Suppression of β1 Integrin by Ectopic CD98-ICD Inhibits Hepatocellular Carcinoma Progression.

Fig.2.D showing Western Blotting from the publication: Dominant Suppression of β1 Integrin by Ectopic CD98-ICD Inhibits Hepatocellular Carcinoma Progression.

Fig.5.A showing Western Blotting from the publication: Apc mutation enhances PyMT-induced mammary tumorigenesis.

Fig.6.A showing Western Blotting in a Homo sapiens (Human) sample from the publication: Molecular targeting of prostate cancer cells by a triple drug combination down-regulates integrin driven adhesion processes, delays cell cycle progression and interferes with the cdk-cyclin axis.

Fig.2.D showing Western Blotting from the publication: Vicrostatin - an anti-invasive multi-integrin targeting chimeric disintegrin with tumor anti-angiogenic and pro-apoptotic activities.

Fig.4.A showing Western Blotting from the publication: Integrin-linked kinase regulates migration and proliferation of human intestinal cells under a fibronectin-dependent mechanism.

20 images found

FAK and p130Cas Modulate Stiffness-Mediated Early Transcription and Cellular Metabolism.

In Cytoskeleton (Hoboken, N.J.) on 1 March 2025 by Tumenbayar, B. I., Pham, K., et al.

Cellular metabolism is influenced by the stiffness of the extracellular matrix. Focal adhesion kinase (FAK) and its binding partner, p130Cas, transmit biomechanical signals, such as substrate stiffness, to the cell to regulate a variety of cellular responses, but their roles in early transcriptional and metabolic responses remain largely unexplored. We cultured mouse embryonic fibroblasts with or without siRNA-mediated FAK or p130Cas knockdown and assessed the early transcriptional responses of these cells to placement on soft and stiff substrates by RNA sequencing and bioinformatics analyses. Exposure to the stiff substrate altered the expression of genes important for metabolic and biosynthetic processes, and these responses were influenced by knockdown of FAK and p130Cas. Our findings reveal that FAK-p130Cas signaling mechanotransduces substrate stiffness to early transcriptional changes that alter cellular metabolism and biosynthesis.
© 2024 Wiley Periodicals LLC.

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A fibronectin gradient remodels mixed-phase mesoderm.

In Science Advances on 19 July 2024 by Zhu, M., Gu, B., et al.

Physical processes ultimately shape tissue during development. Two emerging proposals are that cells migrate toward stiffer tissue (durotaxis) and that the extent of cell rearrangements reflects tissue phase, but it is unclear whether and how these concepts are related. Here, we identify fibronectin-dependent tissue stiffness as a control variable that underlies and unifies these phenomena in vivo. In murine limb bud mesoderm, cells are either caged, move directionally, or intercalate as a function of their location along a stiffness gradient. A modified Landau phase equation that incorporates tissue stiffness accurately predicts cell diffusivity upon loss or gain of fibronectin. Fibronectin is regulated by WNT5A-YAP feedback that controls cell movements, tissue shape, and skeletal pattern. The results identify a key determinant of phase transition and show how fibronectin-dependent directional cell movement emerges in a mixed-phase environment in vivo.

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Identification of Curcumin Targets in the Brain of Epileptic Mice Using DARTS.

In ACS Omega on 28 May 2024 by Zhang, N., Lin, R., et al.

Curcumin, a compound derived from turmeric, is traditionally utilized in East Asian medicine for treating various health conditions, including epilepsy. Despite its involvement in numerous cellular signaling pathways, the specific mechanisms and targets of curcumin in epilepsy treatment have remained unclear. Our study focused on identifying the primary targets and functional pathways of curcumin in the brains of epileptic mice. Using drug affinity responsive target stabilization (DARTS) and affinity chromatography, we identified key targets in the mouse brain, revealing 232 and 70 potential curcumin targets, respectively. Bioinformatics analysis revealed a strong association of these proteins with focal adhesions and cytoskeletal components. Further experiments using DARTS, along with immunofluorescence staining and cell migration assays, confirmed curcumin's ability to regulate the dynamics of focal adhesions and influence cell migration. This study not only advances our understanding of curcumin's role in epilepsy treatment but also serves as a model for identifying therapeutic targets in neurological disorders.
© 2024 The Authors. Published by American Chemical Society.

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Bisbiguanide analogs induce mitochondrial stress to inhibit lung cancer cell invasion.

In IScience on 19 April 2024 by Knippler, C. M., Arnst, J. L., et al.

Targeting cancer metabolism to limit cellular energy and metabolite production is an attractive therapeutic approach. Here, we developed analogs of the bisbiguanide, alexidine, to target lung cancer cell metabolism and assess a structure-activity relationship (SAR). The SAR led to the identification of two analogs, AX-4 and AX-7, that limit cell growth via G1/G0 cell-cycle arrest and are tolerated in vivo with favorable pharmacokinetics. Mechanistic evaluation revealed that AX-4 and AX-7 induce potent mitochondrial defects; mitochondrial cristae were deformed and the mitochondrial membrane potential was depolarized. Additionally, cell metabolism was rewired, as indicated by reduced oxygen consumption and mitochondrial ATP production, with an increase in extracellular lactate. Importantly, AX-4 and AX-7 impacted overall cell behavior, as these compounds reduced collective cell invasion. Taken together, our study establishes a class of bisbiguanides as effective mitochondria and cell invasion disrupters, and proposes bisbiguanides as promising approaches to limiting cancer metastasis.
© 2024 The Authors.

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FAK and p130Cas modulate stiffness-mediated early transcription and cellular metabolism

Preprint on BioRxiv : the Preprint Server for Biology on 16 January 2024 by Tumenbayar, B., Tutino, V. M., et al.

SUMMARY Cellular metabolism is influenced by the stiffness of the extracellular matrix. Focal adhesion kinase (FAK) and its binding partner, p130Cas, transmit biomechanical signals about substrate stiffness to the cell to regulate a variety of cellular responses, but their roles in early transcriptional and metabolic responses remain largely unexplored. We cultured mouse embryonic fibroblasts with or without siRNA-mediated FAK or p130Cas knockdown and assessed the early transcriptional responses of these cells to placement on soft and stiff substrates by RNA sequencing and bioinformatics analyses. Exposure to the stiff ECM altered the expression of genes important for metabolic and biosynthetic processes, and these responses were influenced by knockdown of FAK and p130Cas. Our findings reveal that FAK-p130Cas signaling mechanotransduces ECM stiffness to early transcriptional changes that alter cellular metabolism and biosynthesis.

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Integrin-Mediated Adhesion Promotes Centrosome Separation in Early Mitosis.

In Cells on 16 April 2022 by Kamranvar, S. A., Gupta, D. K., et al.

Fig.1.E

Fig.1.E showing Western Blotting from the publication: Integrin-Mediated Adhesion Promotes Centrosome Separation in Early Mitosis.

Absence of integrin-mediated cell adhesion causes a delay in the early mitotic progression. (A) Experimental design illustrating how the isolated mitotic cells were used to check mitotic progression in the different culture conditions. Centrosomes, microtubules, and nucleus/chromosomes are shown ... more

Absence of integrin-mediated cell adhesion causes a delay in the early mitotic progression. (A) Experimental design illustrating how the isolated mitotic cells were used to check mitotic progression in the different culture conditions. Centrosomes, microtubules, and nucleus/chromosomes are shown in green, red, and blue, respectively. (B) Representative immunofluorescence micrographs illustrating the mitotic progression of BJ cells from early (E)- and late (L)-prometaphase to cytokinesis, the presence of divided cells (mono-nucleated cells), and the cells that failed to complete cytokinesis (bi-nucleated cells). The centrosomes and the mitotic spindle were labeled with antibodies against pericentrin (green) and α-tubulin (red) directly after nocodazole washout (0 min time-point) and after a 30, 60, and 90 min incubation period of the cells adhering to fibronectin or kept in suspension. Nuclei were stained with DAPI. Scale bar, 10 μm. (C,D) Mean (%) ± SE of the number of adherent (C) and non-adherent cells (D) in different mitotic stages determined based on centrosomes’ location, spindle formation, and nucleus status as shown in part B. The color of each bar on the graph corresponds to the different mitotic phases shown in the side color bar of part B. (E) Western blot of the mitotic cells treated as in (C,D) to monitor the phosphorylation of FAK at Tyr397 and total FAK at the described time points. less

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Integrin-Mediated Adhesion Promotes Centrosome Separation in Early Mitosis.

In Cells on 16 April 2022 by Kamranvar, S. A., Gupta, D. K., et al.

Fig.6.A

Fig.6.A showing Western Blotting from the publication: Integrin-Mediated Adhesion Promotes Centrosome Separation in Early Mitosis.

Inhibition of FAK expression reduces the activity of PLK1 and causes frequent mitotic cell death and tetraploidy. (A) Representative Western blot pictures and quantification of the bands showing the difference in the level of pPLK1 relative to the total level of PLK1 and β-actin, and pFAK in the ... more

Inhibition of FAK expression reduces the activity of PLK1 and causes frequent mitotic cell death and tetraploidy. (A) Representative Western blot pictures and quantification of the bands showing the difference in the level of pPLK1 relative to the total level of PLK1 and β-actin, and pFAK in the mitotic Tet-FAK-ON and -OFF MEF cells re-plated on fibronectin for 60 min. (B). Decrease of pPLK1 in the cells lacking FAK expression after 72 h treatment with doxycycline and 5 h after release from the double thymidine block. Representative micrographs illustrating the pPLK1 (green) signals in Tet-FAK ON and OFF cells. Scale bar, 20 μm. (C) Box plots showing the average intensity of pPLK1 in 100 cell nuclei. (D) Representative pictures of Tet-FAK OFF cells with different ploidy. The chromosomes were stained by DAPI. Scale bar, 5 μm. (E) The % of mitotic cells having diploid or tetra/polyploid chromosome sets. A total of 500 chromosome plates were analyzed for each condition of FAK ON and OFF. (F) Snapshot images from a time-lapse movie displaying the mitotic progression of a single FAK OFF cell from the beginning of mitosis (0 min time point) to cell death at the later time point. The nucleus was labeled with SiR-DNA (red). Scale bar, 50 μm. (G) Mean (%) ± SD of the number of mitotic cells that die during the early mitotic progression after rounding up. p-values less than 0.05, 0.01, 0.001, and 0.0001 were shown by two and three stars, respectively. less

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Src-mediated regulation of the PI3K pathway in advanced papillary and anaplastic thyroid cancer.

In Oncogenesis on 28 February 2018 by Beadnell, T. C., Nassar, K. W., et al.

Fig.4.C

Fig.4.C showing Western Blotting in a Homo sapiens (Human) sample from the publication: Src-mediated regulation of the PI3K pathway in advanced papillary and anaplastic thyroid cancer.

rpS6 phosphorylation is neither necessary nor sufficient for the growth inhibitory effects of the combined inhibition of Src and the MAPK pathway.a Western blot analysis was performed on whole cell lysates from the 8505C or C643 cell line expressing a doxycycline inducible P70S6K T389 ΔCT constru... more

rpS6 phosphorylation is neither necessary nor sufficient for the growth inhibitory effects of the combined inhibition of Src and the MAPK pathway.a Western blot analysis was performed on whole cell lysates from the 8505C or C643 cell line expressing a doxycycline inducible P70S6K T389 ΔCT construct. Cells were treated with or without doxycycline for 24 h prior to treatment. After 24 h the cells were treated with either DMSO, 100 nM trametinib, 100 nM dasatinib, or the combination of dasatinib and trametinib. Lysates were analyzed for the indicated antibodies. Quantification was performed using the Odyssey Image Studio V.4.0 Software. Data as means±SEM (n = 3). b Clonogenic growth was detected by crystal violet staining in the 8505C or C643 cell lines expressing the doxycycline inducible P70S6K T389 ΔCT construct. Cell lines were treated with or without doxycycline upon plating. 24 h later the cell lines were treated with either DMSO, 100 nM trametinib, 100 nM dasatinib, or the combination for 6 days. Following 6 days of treatment, the cells were released for an additional 7 days. c Western blot analysis was performed on whole cell lysates from the parental 8505C or parental C643 cell lines treated with either DMSO, 100 nM trametinib, 100 nM dasatinib, or 1 μM Everolimus for 24 h for the indicated antibodies. Quantification was performed using the Odyssey Image Studio V.4.0 Software. Data as means±SEM (n = 3). d Clonogenic growth was detected by crystal violet staining in the 8505C and C643 cell lines. Cell lines were treated for 24 h with either DMSO, 100 nM trametinib, 100 nM dasatinib, dasatinib+trametinib, or 1 µM everolimus. Colony area signal intensity was measured using Odyssey CLx imager (Li-Cor), and presented as percent fold change relative to the DMSO treated wells. Data as means±SEM (n = 3) less

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Sustained activation of the AKT/mTOR and MAP kinase pathways mediate resistance to the Src inhibitor, dasatinib, in thyroid cancer.

In Oncotarget on 28 November 2017 by Mishall, K. M., Beadnell, T. C., et al.

Fig.3.B

Fig.3.B showing Western Blotting in a Homo sapiens (Human) sample from the publication: Sustained activation of the AKT/mTOR and MAP kinase pathways mediate resistance to the Src inhibitor, dasatinib, in thyroid cancer.

The mTOR inhibitor, everolimus, inhibits cell growth and clonogenicity, but does not induce apoptosis(A) Cells were treated with DMSO or Everolimus for 72 hours, and cell number was counted using the ViCell Cell Counter. Fold changes are compared to DMSO. Three independent biological replicates w... more

The mTOR inhibitor, everolimus, inhibits cell growth and clonogenicity, but does not induce apoptosis(A) Cells were treated with DMSO or Everolimus for 72 hours, and cell number was counted using the ViCell Cell Counter. Fold changes are compared to DMSO. Three independent biological replicates were performed, and the standard error mean is displayed in the quantification graphs. p-value * = 0.05 – 0.01, φ = 0.01 – 0.001, δ = 0.001 – 0.001, Ψ < 0.0001. (B-C) BRAF-mutant (B) and RAS-mutant (C) control and DasRes cells were treated with the indicated inhibitors for 24 hours. Cell lysate was harvested and a Western blot was performed to determine changes in the AKT/mTOR pathway signaling. Control cells were treated with 100nM dasatinib, and DasRes cells were treated with 2μM dasatinib. Three independent biological replicates were performed, and representative blots for signaling proteins and loading controls are shown. The pY416 Src blot was stripped and reprobed for total Src. (D-E) BRAF-mutant (D) and Ras-mutant (E) control and DasRes cells were treated with the indicated inhibitors for 7 days, and then released for 7 days to assess colony growth after inhibitor treatment. After 2 weeks, cells were fixed and stained with crystal violet. (F) Apoptosis was measured after 24 hours of everolimus treatment by caspase 3/7 cleavage using the Caspase-Glo 3/7 kit. Fold changes were calculated by comparing treatments to DMSO. Three independent biological replicates were performed, and the standard error mean is displayed in the quantification graphs. less

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Sustained activation of the AKT/mTOR and MAP kinase pathways mediate resistance to the Src inhibitor, dasatinib, in thyroid cancer.

In Oncotarget on 28 November 2017 by Mishall, K. M., Beadnell, T. C., et al.

Fig.3.C

Fig.3.C showing Western Blotting in a Homo sapiens (Human) sample from the publication: Sustained activation of the AKT/mTOR and MAP kinase pathways mediate resistance to the Src inhibitor, dasatinib, in thyroid cancer.

The mTOR inhibitor, everolimus, inhibits cell growth and clonogenicity, but does not induce apoptosis(A) Cells were treated with DMSO or Everolimus for 72 hours, and cell number was counted using the ViCell Cell Counter. Fold changes are compared to DMSO. Three independent biological replicates w... more

The mTOR inhibitor, everolimus, inhibits cell growth and clonogenicity, but does not induce apoptosis(A) Cells were treated with DMSO or Everolimus for 72 hours, and cell number was counted using the ViCell Cell Counter. Fold changes are compared to DMSO. Three independent biological replicates were performed, and the standard error mean is displayed in the quantification graphs. p-value * = 0.05 – 0.01, φ = 0.01 – 0.001, δ = 0.001 – 0.001, Ψ < 0.0001. (B-C) BRAF-mutant (B) and RAS-mutant (C) control and DasRes cells were treated with the indicated inhibitors for 24 hours. Cell lysate was harvested and a Western blot was performed to determine changes in the AKT/mTOR pathway signaling. Control cells were treated with 100nM dasatinib, and DasRes cells were treated with 2μM dasatinib. Three independent biological replicates were performed, and representative blots for signaling proteins and loading controls are shown. The pY416 Src blot was stripped and reprobed for total Src. (D-E) BRAF-mutant (D) and Ras-mutant (E) control and DasRes cells were treated with the indicated inhibitors for 7 days, and then released for 7 days to assess colony growth after inhibitor treatment. After 2 weeks, cells were fixed and stained with crystal violet. (F) Apoptosis was measured after 24 hours of everolimus treatment by caspase 3/7 cleavage using the Caspase-Glo 3/7 kit. Fold changes were calculated by comparing treatments to DMSO. Three independent biological replicates were performed, and the standard error mean is displayed in the quantification graphs. less

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Vicrostatin - an anti-invasive multi-integrin targeting chimeric disintegrin with tumor anti-angiogenic and pro-apoptotic activities.

In PLoS One on 10 June 2010 by Minea, R. O., Helchowski, C. M., et al.

Fig.2.D

Fig.2.D showing Western Blotting from the publication: Vicrostatin - an anti-invasive multi-integrin targeting chimeric disintegrin with tumor anti-angiogenic and pro-apoptotic activities.

The expression, purification and initial characterization of VCN as an active disintegrin.Panel A – The production of Trx-VCN was assessed in different E. coli strains that were transformed, grown, induced, and processed under identical conditions. The same amount (5 µl) of cell lysates from each... more

The expression, purification and initial characterization of VCN as an active disintegrin.Panel A – The production of Trx-VCN was assessed in different E. coli strains that were transformed, grown, induced, and processed under identical conditions. The same amount (5 µl) of cell lysates from each induced strain was loaded on a precast gel and Coomassie stained. Unlike the BL21 (DE3) strain, the lysates from both AD494 (DE3) and Origami B (DE3) strains generate a unique and consistent Trx-VCN band (shown by the arrow). By employing a modified media recipe, the Origami B (DE3) Trx-VCN transformants achieve higher cell densities at the end of the induction time, generating up to 200 mg of soluble VCN per L of bacterial culture after purification. Panel B – Coomassie stained gel showing the migration of Trx-VCN before and after TEV proteolysis (lanes 2 and 3, respectively) versus C18 reverse phase-HPLC purified VCN (lane 4). Panel C– VCN and native CN exhibit an almost identical dose-dependent inhibitory effect against ADP-induced platelet aggregation when incubated with human platelet-rich plasma (with a calculated IC50 of ∼60 nM). In contrast, the rCN construct, which is also expressed as a soluble polypeptide in Origami B (DE3), shows no inhibitory activity. Panel D– The agonistic activity of VCN (FAK activation) was assessed in serum-starved non-migratory MDA-MB-435 cells kept in suspension and exposed to increasing concentrations of disintegrins for 30 min. Similar to dimeric CN, VCN is also shown to engage integrins agonistically (outside-in signaling). less

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