plk1 solution  (Carna Inc)

Journal: bioRxiv

Article Title: Mitochondrial Signatures Shape Phenotype Switching and Apoptosis in Response to PLK1 and RSK Inhibitors in Melanoma

doi: 10.1101/2024.06.14.599035

Figure Lengend Snippet: (A) CXCL8 transcript levels in MeWo and A-375 cells treated for 72 h with DMSO 0.1 % (v/v) or a panel of inhibitors targeting putative BI-D1870 targets. NA: non-available due to cell death. (B) Viable cell counts of melanoma cells with Presto Blue HS. (C) Representative crystal violet staining of cells treated as in A. (D) Quantification of crystal violet staining, as in C. (E,F) Immunoblotting of cell extracts from cells treated as indicated for 72 h. (G) Densitometric analysis of immunoblotting from 3 independent experiments related to E and F. (H,I) PLK1 and CXCL8 transcript levels in cells 72 h after PLK1 knockdown. (J) Immunoblotting of cell extracts from cells treated as in H-I. Numbers indicate independent experiments. (K) Densitometric analysis of immunoblotting from 3 independent experiments related to J. All data are shown as means ± SEM from 3 independent experiments. A: * p < 0.05, unpaired Student’s t -test (cell lines tested separately). D,K: * p < 0.05, paired Student’s t -test. G-I: * p < 0.05, Two-Way ANOVA with Tukey multiple comparison test.

Article Snippet: Non-targeting siRNA (Allstar, 1027280) and siRNA pools, made of four siRNAs (FlexiTube Gene Solutions, 1027416) against human PLK1 (GS5347), ABCD1 (GS215), BCL2A1 (GS597), and PRKACA (GS5566) were purchased from Qiagen (Hilden, DEU).

Techniques: Staining, Western Blot, Knockdown, Comparison

Journal: bioRxiv

Article Title: Mitochondrial Signatures Shape Phenotype Switching and Apoptosis in Response to PLK1 and RSK Inhibitors in Melanoma

doi: 10.1101/2024.06.14.599035

Figure Lengend Snippet: (A, B, C) Gene expression in MeWo and A-375 cells treated for 72 h as indicated. NA: Not available due to cell death. (D) Immunoblotting of protein extracts from MeWo cells treated as in A-C. Numbers indicate independent experiments. (E) Gene expression of differentiation markers in cells treated with siRNA against PLK1 versus non-targeting siRNA for 72 h. (F) Heat-map of Log2 fold change of specific transcripts linked to phenotype switching in MeWo cells treated with BI-D1870 (3 µM) for 72 h. Derived from RNA-Seq analyses in . (G,H) Gene expression in MeWo cells treated with BI 6727 (1 µM) or BRD7389 (10 µM) for 72 h. All data are shown as mean ± SEM from 3 independent experiments. A-C, E: * p < 0.05, Two-Way ANOVA with Tukey multiple comparison test. F: * p-adj < 0.05. G-H: * < 0.05, paired Student’s t -test.

Article Snippet: Non-targeting siRNA (Allstar, 1027280) and siRNA pools, made of four siRNAs (FlexiTube Gene Solutions, 1027416) against human PLK1 (GS5347), ABCD1 (GS215), BCL2A1 (GS597), and PRKACA (GS5566) were purchased from Qiagen (Hilden, DEU).

Techniques: Expressing, Western Blot, Derivative Assay, RNA Sequencing Assay, Comparison

Journal: bioRxiv

Article Title: Mitochondrial Signatures Shape Phenotype Switching and Apoptosis in Response to PLK1 and RSK Inhibitors in Melanoma

doi: 10.1101/2024.06.14.599035

Figure Lengend Snippet: (A) Schematic depicting the strategy to identify transcripts associated with resistance to PLK1 targeting. The six PLK inhibitors, in addition to BI-D1870, are indicated in panel E. (B) Transcripts that significantly correlate (p < 0.05) with 2 or more PLK1-directed treatments were selected for further analyses. (C) Functional gene analysis (g:Profiler, GO:BP) performed on correlating transcripts from B. Bars indicate p-adj values and circles indicate number of transcripts associated with a gene set. (D) Essentiality of transcripts encoding for mitochondrial proteins that significantly correlate with resistance to PLK targeting in melanoma cells (DepMap). (E) Heat-map representation and hierarchical clustering of Spearman correlations between the 124 mitochondrial signature transcripts and respective impact on proliferation dynamics of BI-D1870 and PLK1 targeting approaches (DepMap). (F) Functional classification of 124 mitochondrial signature transcripts associated with resistance to PLK1 targeting, according to MitoPathways3.0. (G) STRING interaction network of the 124 mitochondrial signature transcripts associated with resistance to PLK1 targeting. Nodes indicate proteins, and edges indicate interactions. Spearman’s correlation with proliferation dynamics (DepMap) is indicated by nodes’ colored borders. (H) Gene expression in MeWo cells transfected with the indicated siRNA for 48 h and treated with DMSO 0.1 % (v/v) or BI 6726 (100 nM) for 72 h. (I) Relative viable cell counts of MeWo cells treated as in H but with 10-fold dilutions of BI 6727, using Presto Blue HS. (J) Model depicting the impact of mitochondrial signatures on the response to RSK and PLK inhibitors. All data are shown as means ± SEM from 3 or more independent experiments. * p < 0.05, One-Way ANOVA with Dunnett multiple comparison test.

Article Snippet: Non-targeting siRNA (Allstar, 1027280) and siRNA pools, made of four siRNAs (FlexiTube Gene Solutions, 1027416) against human PLK1 (GS5347), ABCD1 (GS215), BCL2A1 (GS597), and PRKACA (GS5566) were purchased from Qiagen (Hilden, DEU).

Techniques: Functional Assay, Expressing, Transfection, Comparison