Anti-PLCL1 antibody [EPR11213]

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-PLCL1 antibody [EPR11213] (AB157200)

Immunofluorescence analysis of Jurkat cells, labeling PLCL1 using ab157200 at a 1/100 dilution.

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-PLCL1 antibody [EPR11213] (AB157200)

Intracellular Flow Cytometry analysis of permeabilized Jurkat cells labeling PLCL1 (red), using ab157200 at a 1/100 dilution, and negative control cells probed with a Rabbit IgG (green)

• IP

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Immunoprecipitation - Anti-PLCL1 antibody [EPR11213] (AB157200)

Western blot analysis of Immunoprecipitation pellet from Human fetal brain lysate showing PLCL1, using ab157200 at 1/10 dilution for IP.

All lanes:

Immunoprecipitation - Anti-PLCL1 antibody [EPR11213] (ab157200)

Predicted band size: 123 kDa

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• WB

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Western blot - Anti-PLCL1 antibody [EPR11213] (AB157200)

All lanes:

Western blot - Anti-PLCL1 antibody [EPR11213] (ab157200) at 1/10000 dilution

Lane 1:

Human fetal brain tissue lysate at 10 µg

Lane 2:

Human fetal kidney tissue lysate at 10 µg

Lane 3:

Human fetal lung tissue lysate at 10 µg

Lane 4:

Jurkat lysate at 10 µg

Predicted band size: 123 kDa

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• ICC

CiteAb

Immunocytochemistry - Anti-PLCL1 antibody [EPR11213] (AB157200)

Immunocytochemistry using Anti-PLCL1 antibody [EPR11213], ab157200. Publication image from Xiong, Z. et al., 2019, Adv Sci (Weinh), 31131187. Legend direct from paper.

PLCL1 repressed ccRCC progression and promoted tumor cell “slimming” in ccRCC. PLCL1‐overexpressing or PLCL1‐knockdown ccRCC cell lines were constructed by transfecting overexpressing lentivirus and shRNA. The results were plotted as the mean ± SEM of three independent experiments with at least three replicates in each independent experiment. ****, p < 0.0001, ***, p < 0.001, **, p < 0.01, *, p < 0.05. A,B) Cell growth curves of CCK8 assays for indicated cells. C) Migration and invasion assay for indicated ccRCC cells (Magnification : 200x). D) Photomicrographs of Oil Red O staining of PLCL1‐overexpressing cell lines and the negative control (Magnification : 400x & 800x). Relative TG (mmol/gprot) tested by a triglyceride assay kit, relative diameters of lipid droplets, and relative diameters of cells for ccRCC cells described above. t‐test, ****, p < 0.0001, ***, p < 0.001, *, p < 0.05. E) Representative photographs of immunofluorescence of UCP1 for cells overexpressing PLCL1 (Magnification : 600x).

• WB

CiteAb

Western blot - Anti-PLCL1 antibody [EPR11213] (AB157200)

Western Blotting using Anti-PLCL1 antibody [EPR11213], ab157200. Publication image from Xiong, Z. et al., 2019, Adv Sci (Weinh), 31131187. Legend direct from paper.

PLCL1 upregulated the expression of the lipid browning related gene UCP1 in ccRCC cells. Transcriptome sequencing was performed for A498 cells overexpressing PLCL1. A) GO enrichment for the indicated cells based on the results from sequencing. B) GSEA assays for the correlation of energy metabolism, fatty acid metabolism, and mRNA levels of PLCL1 according to the TCGA database. FDR < 25%, p < 0.05 was considered statistically significant. C) KEGG (Kyoto Encyclopedia of Genes and Genomes) enrichment top 20 for indicated cells based on the results from sequencing. D) The heatmap of cluster analysis based on sequencing results. E) Lipid browning‐related proteins were determined by western blot analysis. GAPDH was used as a loading control. F) GSEA assays for the correlation of PPAR signaling and mRNA level of PLCL1 according to TCGA database. FDR < 25%, p < 0.05 was considered as statistically significant. G) Western blot assay for the protein levels of PLCL1 and UCP1 in the PLCL1‐knockdown cells.

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• WB

CiteAb

Western blot - Anti-PLCL1 antibody [EPR11213] (AB157200)

Western Blotting using Anti-PLCL1 antibody [EPR11213], ab157200. Publication image from Xiong, Z. et al., 2019, Adv Sci (Weinh), 31131187. Legend direct from paper.

PLCL1 regulated the protein stability of UCP1 by affecting UCP1 ubiquitination levels in ccRCC. A) Relative mRNA levels of UCP1 in the cells PLCL1‐overexpressing and PLCL1‐knockdown cells. p = ns (no significance). B,C) ccRCC cells with stable PLCL1 knockdown and overexpression were treated with cycloheximide (CHX) at the indicated time points. Cells were collected, and UCP1 protein expression was analyzed by western blot. D) Cells with stable PLCL1 knockdown were treated with vehicle (DMSO), MG132 (20 x 10−6m) or chloroquine (50 x 10−6m) for 12 h. Western blotting was used to analyze the protein level of UCP1. E) GO enrichment for the indicated cells based on the results from sequencing. F,G) The cells with stable PLCL1 knockdown and overexpression were lysed and subjected to immunoprecipitation with an antibody against UCP1 and analyzed by western blotting with an anti‐ubiquitin antibody.

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• WB

CiteAb

Western blot - Anti-PLCL1 antibody [EPR11213] (AB157200)

Western Blotting using Anti-PLCL1 antibody [EPR11213], ab157200. Publication image from Xiong, Z. et al., 2019, Adv Sci (Weinh), 31131187. Legend direct from paper.

PLCL1 repressed ccRCC progression and promoted tumor cell “slimming” through UCP1‐mediated lipid browning. On the basis of PLCL1 overexpression, cells with UCP1 knocked down were constructed by transfecting UCP1 siRNA. We constructed four groups of cell lines for rescue experiments, including cell lines with PLCL1 expression lentivirus control vector and siUCP1 control, cell lines with PLCL1 expression lentivirus and siUCP1 control, cell lines with PLCL1 expression lentivirus control vector and siUCP1, and cell lines with PLCL1 expression lentivirus and siUCP1. A) Western blot assay for the protein levels of PLCL1 and UCP1 in indicated cells. B) Cell growth curves of CCK8 assays for indicated cells. ***, p < 0.001, **, p < 0.01, p = ns (no significance). C) Migration and invasion assay for indicated ccRCC cells (Magnification : 200x). t‐test, ***, p < 0.001, **, p < 0.01, *, p < 0.05, p = ns (no significance). D) Photomicrographs of Oil Red O staining of indicated cells (Magnification : 400x). Relative diameters of lipid droplets and the ccRCC cells described above.

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• WB

CiteAb

Western blot - Anti-PLCL1 antibody [EPR11213] (AB157200)

Western Blotting using Anti-PLCL1 antibody [EPR11213], ab157200. Publication image from Xiong, Z. et al., 2019, Adv Sci (Weinh), 31131187. Legend direct from paper.

PLCL1 was downregulated and predicted poor prognosis in ccRCC. A) A Venn diagram of three independent lipid‐related gene sets from the Oncomine database (https://www.oncomine.org) and the European Bioinformatics Institute (EMBL‐EBI) (https://www.ebi.ac.uk). (All gene sets are subgene sets of differentially expressed genes in ccRCC.) B) The mRNA levels of PLCL1 and PLCG2 in 533 ccRCC tissues and 72 paired tissues in ccRCC based on data from the TCGA database. (In the color scheme of the heatmap, the colder color represents the lower gene expression level, and the warmer color represents the higher gene expression level.) t‐test, p < 0.0001. C) The Kaplan–Meier curves of PLCL1 and PLCG2 in ccRCC for both overall survival (OS) and disease‐free survival (DFS). D) The ROC (receiver operating characteristic) curves of PLCL1 (AUC = 0.9642 95% CI : 0.9343 to 0.9941; p < 0.0001) and PLCG2 (AUC = 0.9466 95% CI : 0.9253 to 0.9678; p < 0.0001) in ccRCC. E) The mRNA levels of PLCL1 in 30 ccRCC tissues and adjacent nonmalignant tissues. t‐test, p < 0.0001. F) The protein levels of PLCL1 in ccRCC tissues and adjacent nonmalignant tissues (Abbreviation : N, Normal tissue; T, Tumor tissue). G) The immunohistochemistry (IHC) staining for PLCL1 in ccRCC tissues and adjacent nonmalignant tissues (Magnification : 200x & 400x). H) The mRNA and protein levels in five ccRCC cell lines (786‐0, A498, ACHN, CAKI, and OSRC) and normal cell line (293). t‐test, ****p < 0.0001.

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