hcc38  (ATCC)

Journal: The Journal of Cell Biology

Article Title: Non-cell autonomous regulation of cell–cell signaling and differentiation by mitochondrial ROS

doi: 10.1083/jcb.202401084

Figure Lengend Snippet: ROS prevents exocyst binding to the membrane. (A) A diagram describing the MCF7 cell model used to examine the impact of a 30-min ROS exposure on the membrane proteome. (B) A volcano plot examining the changes in membrane proteome caused by ROS exposure (FDR < 0.05). (C) A heat map depicting the expression changes of the top 100 most differentially regulated proteins in membranes isolated from ROS-exposed cells (FDR < 0.05). (D) Protein abundances for several common membrane proteins in control and H 2 O 2 -treated cells. (E) Membrane abundances for exocyst complex components in control and H 2 O 2 -treated MCF7 cells (FDR < 0.05). (F) A heat map showing the top 100 most differentially regulated proteins in membrane fraction from 3T3 cells and 3T3 treated with H 2 O 2 . (G) A heat map representing proteomic-based measurements of the detectable subunits of the exocyst complex from membrane fractions isolated from Drosophila eggs. (H) A heat map representing proteomic-based measurements of the detectable subunits of the exocyst complex from membrane fractions isolated from NIH3T3 cells. (I) A heat map representing proteomic-based measurements of known exocyst regulators from membrane fractions isolated from MCF7 cells. (J) Western blot validation for EXOC 1 and 6 from MFC7 total cell lysate, cytosolic fractions, and purified membrane fractions. (K) Western blots examining the levels of Exoc1 and Exoc6 in membrane fractions from MCF7 cells and BT549 cells. (L) Western blots examining Exoc1, Exoc3, and Exoc6 levels in control liver and Myc-O/E hepatic tumors. Student’s t test was used for all pairwise comparisons, and one-way ANOVA was used for all experiments containing >2 sample groups. Error bars represent the standard deviation. Source data are available for this figure: .

Article Snippet: BT549 , ATCC , HTB -122.

Techniques: Binding Assay, Membrane, Expressing, Isolation, Control, Western Blot, Biomarker Discovery, Purification, Standard Deviation

Journal: The Journal of Cell Biology

Article Title: Non-cell autonomous regulation of cell–cell signaling and differentiation by mitochondrial ROS

doi: 10.1083/jcb.202401084

Figure Lengend Snippet: ROS production disrupts endosomal traffikcing and cell cell communication. (A) GO enrichment for protein domains found in proteins downregulated in membrane fractions purified from MCF7 cells exposed to ROS. (B) ROS levels from MCF7 cells and BT549 cells. (C) Quantification of the levels of EXOC1 and EXOC6 in membrane fractions from MCF7 cells and BT549 cells ( n = 3). (D) A heat map showing the abundance of RAB family GTPases in purified membrane fractions from MCF7 cells. (E) DHE staining of PHB1-RNAi and PHB1-RNAI; +RAB11-O/E ovarioles. (F) quantification of peb antibody staining fluorescence from control and PIPK-RNAi ovarioles ( n = 20). (G) volcano plot depicting the protein level differences between control 3T3cell and 3T3 cells cultured with H 2 O 2 . (H) A heat map depicting the expression of NRF2 target genes in cells adjacent to ROS-producing cells. (I) a heatmap of the gene expression abundance changes in genes associated with the term “secreted protein.” Expression levels are expressed as a ratio ROS-coculture/control. (J) A heatmap of the top 100 most significant differentially regulated genes in cells adjacent to ROS-producing cells. (K) A heatmap of the abundance of genes associated with the gene ontology term “extracellular protein” from our list of genes downregulated in cells adjacent to ROS-producing cells(Red text indicates a known role in cancer growth, metastasis, and angiogenesis). (L) ROS-Levels in Hek293 control cells, DAAO-expressing, and cells co-cultured adjacent to ROS-producing DAAO+ cells ( n = 25). (M) A Venn diagram showing the overlap of genes known to be regulated by extracellular ROS exposure (NCBI GEO: GSE227554 , H 2 O 2 exposure for 6 h) and our list of genes regulated by co-culture in contact with ROS-producing cells. (N) GO term enrichment for the 23 genes overlapping our data and the H 2 O 2 -regulated genes studied in GSE227554 . Student’s t test was used for all pairwise comparisons, and one-way ANOVA was used for all experiments containing >2 sample groups. Error bars represent the standard deviation.

Article Snippet: BT549 , ATCC , HTB -122.

Techniques: Membrane, Purification, Staining, Fluorescence, Control, Cell Culture, Expressing, Gene Expression, Co-Culture Assay, Standard Deviation

Journal: The Journal of Cell Biology

Article Title: Non-cell autonomous regulation of cell–cell signaling and differentiation by mitochondrial ROS

doi: 10.1083/jcb.202401084

Figure Lengend Snippet: ROS regulates endosomal recycling by reducing the levels of PIP2 in membranes. (A) Images of ovarioles from control and PHB-RNAi females. Top image DAPI, middle Images PIP2, bottom image LUT intensity image of PIP2 levels. (B) images of (MCF7) and (BT549) cells stained with DAPI and PIP2 antibodies (LUT intensity image of PIP2 levels). (C) Images of control (3T3) and (BT549) cells stained with PIP2 antibodies (LUT intensity image of PIP2 levels). Fluorescence levels are expressed with a Fire LUT. Arrows indicate a PIP signal in the cell body. (D) Quantification of PIP2 antibody staining levels from control and PHB-RNAi ovarioles. (E) Quantification of PIP2 antibody staining fluorescence from (MCF7) and (BT549) cells. (F) Quantification of PIP2 antibody staining fluorescence from control (3T3) and (BT549) cells. (G) Quantification of the percentage of follicles containing punctate Delta staining from three independent experiments. (Each experiment contained at least 25 egg chambers, and the total number of eggs assayed for each group was at least 85). (H and I) Delta antibody staining images from control and PIP5K-RNAi ( sktl -RNAi) ovarioles. Arrows point to Delta puncta. (J and K) peb antibody staining images from control and PIP5K-RNAi ( sktl -RNAi) ovarioles. (L) Quantification of the percentage of follicles containing abnormally low peb expression from three independent experiments. (Each experiment contained at least 25 egg chambers, and the total number of egg chambers assayed for each group was at least 85). Student’s t test was used for all pairwise comparisons, and one-way ANOVA was used for all experiments containing >2 sample groups. Error bars represent the standard deviation.

Article Snippet: BT549 , ATCC , HTB -122.

Techniques: Control, Staining, Fluorescence, Expressing, Standard Deviation

Journal: The Journal of Cell Biology

Article Title: Non-cell autonomous regulation of cell–cell signaling and differentiation by mitochondrial ROS

doi: 10.1083/jcb.202401084

Figure Lengend Snippet: Reagents

Article Snippet: BT549 , ATCC , HTB -122.

Techniques: Virus, Recombinant, Membrane, Protein Extraction, Over Expression, Software