od6 ultra bandpass emission filter  (Alluxa Inc)

Journal: JCI Insight

Article Title: HIF2 α inhibits glutaminase clustering in mitochondria to sustain growth of clear cell renal cell carcinoma

doi: 10.1172/jci.insight.182711

Figure Lengend Snippet: ( A ) Rescue of GLS1 clustering by DMOG treatment is reversed by HIF2 α but not HIF1 α knockout (KO) in HUVECs. Correlation coefficient ( r ) for GLS1 (red) and COX IV (green) is shown on the right. ( B ) Western blotting analysis showing the validation of HIF1 α KO and HIF2 α KO in HUVECs. gRNAs are numbered 1–4 on top. V, vector. HIF1 α gRNA1 and HIF2 α gRNA2 were selected in the assays. ( C ) qPCR analysis showing the validation of DMOG and HIF2α transcriptional inhibitors in HUVECs. HIF1α target genes: LDHA , HK2 , ENO1 , and PDK1 . HIF2α target genes: VEGFA and DLL4 . ( D ) No reversal of the DMOG-induced GLS1 redistribution by inhibitors targeting the transcriptional activity of HIF2α. ( E ) Rescue of noQ-induced cell death by DMOG treatment in HUVECs. *** P < 0.001; **** P < 0.0001 by 1-way ANOVA followed by Bonferroni’s post hoc testing. Scale bars: 10 μm ( A and D ) and 100 μm ( E ).

Article Snippet: The following primary antibodies were used: GLS1 (Abcam, ab156876), CS (Sigma-Aldrich, SAB2702186), TIMM23 (Abcam, ab116329), DRP1 (Proteintech, 12957-1-AP), HIF1α (Novus Bio, NB100-296), HIF2α (CST, 7096s), MYC-tag (CST, 2278s), mCherry (CST, 43590), 14-3-3 (CST, 8312), and GAPDH (CST, 5174).

Techniques: Knock-Out, Western Blot, Biomarker Discovery, Plasmid Preparation, Activity Assay

Journal: JCI Insight

Article Title: HIF2 α inhibits glutaminase clustering in mitochondria to sustain growth of clear cell renal cell carcinoma

doi: 10.1172/jci.insight.182711

Figure Lengend Snippet: ( A and B ) Western blotting analysis for the validation of HIF2 α KO ( A ) and its translational inhibitor ( B ) in UMRC2 cells. ( C ) Resistance to noQ-induced GLS1 clustering in UMRC2 cells is reversed by HIF2 α KO. ( D ) Resistance to noQ-induced cell death in UMRC2 cells is reversed by treatment with an inhibitor of HIF2α translation. ( E ) Resistance to noQ-induced cell death in UMRC2 cells is reversed by HIF2 α KO. ( F ) Increased cell death in UMRC2 cells overexpressing the K320A mutant GLS1 compared with those overexpressing WT GLS. *** P < 0.001; **** P < 0.0001 by 1-way ANOVA followed by Bonferroni’s post hoc testing ( C – E ) or 2-tailed Student’s t test ( F ). Scale bars: 10 μm ( C ) and 100 μm ( D – F ).

Article Snippet: The following primary antibodies were used: GLS1 (Abcam, ab156876), CS (Sigma-Aldrich, SAB2702186), TIMM23 (Abcam, ab116329), DRP1 (Proteintech, 12957-1-AP), HIF1α (Novus Bio, NB100-296), HIF2α (CST, 7096s), MYC-tag (CST, 2278s), mCherry (CST, 43590), 14-3-3 (CST, 8312), and GAPDH (CST, 5174).

Techniques: Western Blot, Biomarker Discovery, Mutagenesis