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FHIT regulates the downstream target KDM6B by mediating its mRNA translation. A , Circos plot displaying Gene Ontology (GO) terms enriched among differentially translated genes identified by Ribo-Seq data. The “LogFC” scale represents the normalized enrichment significance of each pathway (range, 0–1). B , polysome gradient traces from AGS and FHIT-overexpressing AGS cells separated on a 10% to 50% linear sucrose gradient. Western blot analysis confirms FHIT protein expression in both cells. C , RT–qPCR analysis of KDM6B mRNA distribution across polysome fractions in AGS cells with or without FHIT overexpression. mRNA levels were normalized to spiked luciferase <t>RNA</t> and plotted as the percentage of total mRNA in each gradient fraction relative to the input. D – F , RT–qPCR analysis of KDM6B mRNA expression and Western blot detection of KDM6B protein levels in gastric cancer cells or NOC-transformed cells with or without FHIT overexpression. G , Western blot analysis of KDM6B protein levels in NOC-transformed cells (GES-1-MNU) transfected with empty vector (EV) or FHIT-overexpressing constructs after treatment with the transcriptional inhibitor ActD to block nascent mRNA synthesis. The analyses were repeated three times, and the results were expressed as mean ± SD. ns, not significant, ∗ p < 0.05, and ∗∗ p < 0.01. ActD, actinomycin D; FHIT, fragile histidine triad; KDM6B, lysine-specific demethylase 6B; MNU, N -nitroso- N -methylurea; qPCR, quantitative PCR; Ribo-Seq, ribosome <t>sequencing.</t>
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FHIT regulates the downstream target KDM6B by mediating its mRNA translation. A , Circos plot displaying Gene Ontology (GO) terms enriched among differentially translated genes identified by Ribo-Seq data. The “LogFC” scale represents the normalized enrichment significance of each pathway (range, 0–1). B , polysome gradient traces from AGS and FHIT-overexpressing AGS cells separated on a 10% to 50% linear sucrose gradient. Western blot analysis confirms FHIT protein expression in both cells. C , RT–qPCR analysis of KDM6B mRNA distribution across polysome fractions in AGS cells with or without FHIT overexpression. mRNA levels were normalized to spiked luciferase <t>RNA</t> and plotted as the percentage of total mRNA in each gradient fraction relative to the input. D – F , RT–qPCR analysis of KDM6B mRNA expression and Western blot detection of KDM6B protein levels in gastric cancer cells or NOC-transformed cells with or without FHIT overexpression. G , Western blot analysis of KDM6B protein levels in NOC-transformed cells (GES-1-MNU) transfected with empty vector (EV) or FHIT-overexpressing constructs after treatment with the transcriptional inhibitor ActD to block nascent mRNA synthesis. The analyses were repeated three times, and the results were expressed as mean ± SD. ns, not significant, ∗ p < 0.05, and ∗∗ p < 0.01. ActD, actinomycin D; FHIT, fragile histidine triad; KDM6B, lysine-specific demethylase 6B; MNU, N -nitroso- N -methylurea; qPCR, quantitative PCR; Ribo-Seq, ribosome <t>sequencing.</t>
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FHIT regulates the downstream target KDM6B by mediating its mRNA translation. A , Circos plot displaying Gene Ontology (GO) terms enriched among differentially translated genes identified by Ribo-Seq data. The “LogFC” scale represents the normalized enrichment significance of each pathway (range, 0–1). B , polysome gradient traces from AGS and FHIT-overexpressing AGS cells separated on a 10% to 50% linear sucrose gradient. Western blot analysis confirms FHIT protein expression in both cells. C , RT–qPCR analysis of KDM6B mRNA distribution across polysome fractions in AGS cells with or without FHIT overexpression. mRNA levels were normalized to spiked luciferase <t>RNA</t> and plotted as the percentage of total mRNA in each gradient fraction relative to the input. D – F , RT–qPCR analysis of KDM6B mRNA expression and Western blot detection of KDM6B protein levels in gastric cancer cells or NOC-transformed cells with or without FHIT overexpression. G , Western blot analysis of KDM6B protein levels in NOC-transformed cells (GES-1-MNU) transfected with empty vector (EV) or FHIT-overexpressing constructs after treatment with the transcriptional inhibitor ActD to block nascent mRNA synthesis. The analyses were repeated three times, and the results were expressed as mean ± SD. ns, not significant, ∗ p < 0.05, and ∗∗ p < 0.01. ActD, actinomycin D; FHIT, fragile histidine triad; KDM6B, lysine-specific demethylase 6B; MNU, N -nitroso- N -methylurea; qPCR, quantitative PCR; Ribo-Seq, ribosome <t>sequencing.</t>
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FHIT regulates the downstream target KDM6B by mediating its mRNA translation. A , Circos plot displaying Gene Ontology (GO) terms enriched among differentially translated genes identified by Ribo-Seq data. The “LogFC” scale represents the normalized enrichment significance of each pathway (range, 0–1). B , polysome gradient traces from AGS and FHIT-overexpressing AGS cells separated on a 10% to 50% linear sucrose gradient. Western blot analysis confirms FHIT protein expression in both cells. C , RT–qPCR analysis of KDM6B mRNA distribution across polysome fractions in AGS cells with or without FHIT overexpression. mRNA levels were normalized to spiked luciferase <t>RNA</t> and plotted as the percentage of total mRNA in each gradient fraction relative to the input. D – F , RT–qPCR analysis of KDM6B mRNA expression and Western blot detection of KDM6B protein levels in gastric cancer cells or NOC-transformed cells with or without FHIT overexpression. G , Western blot analysis of KDM6B protein levels in NOC-transformed cells (GES-1-MNU) transfected with empty vector (EV) or FHIT-overexpressing constructs after treatment with the transcriptional inhibitor ActD to block nascent mRNA synthesis. The analyses were repeated three times, and the results were expressed as mean ± SD. ns, not significant, ∗ p < 0.05, and ∗∗ p < 0.01. ActD, actinomycin D; FHIT, fragile histidine triad; KDM6B, lysine-specific demethylase 6B; MNU, N -nitroso- N -methylurea; qPCR, quantitative PCR; Ribo-Seq, ribosome <t>sequencing.</t>
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FHIT regulates the downstream target KDM6B by mediating its mRNA translation. A , Circos plot displaying Gene Ontology (GO) terms enriched among differentially translated genes identified by Ribo-Seq data. The “LogFC” scale represents the normalized enrichment significance of each pathway (range, 0–1). B , polysome gradient traces from AGS and FHIT-overexpressing AGS cells separated on a 10% to 50% linear sucrose gradient. Western blot analysis confirms FHIT protein expression in both cells. C , RT–qPCR analysis of KDM6B mRNA distribution across polysome fractions in AGS cells with or without FHIT overexpression. mRNA levels were normalized to spiked luciferase <t>RNA</t> and plotted as the percentage of total mRNA in each gradient fraction relative to the input. D – F , RT–qPCR analysis of KDM6B mRNA expression and Western blot detection of KDM6B protein levels in gastric cancer cells or NOC-transformed cells with or without FHIT overexpression. G , Western blot analysis of KDM6B protein levels in NOC-transformed cells (GES-1-MNU) transfected with empty vector (EV) or FHIT-overexpressing constructs after treatment with the transcriptional inhibitor ActD to block nascent mRNA synthesis. The analyses were repeated three times, and the results were expressed as mean ± SD. ns, not significant, ∗ p < 0.05, and ∗∗ p < 0.01. ActD, actinomycin D; FHIT, fragile histidine triad; KDM6B, lysine-specific demethylase 6B; MNU, N -nitroso- N -methylurea; qPCR, quantitative PCR; Ribo-Seq, ribosome <t>sequencing.</t>
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FHIT regulates the downstream target KDM6B by mediating its mRNA translation. A , Circos plot displaying Gene Ontology (GO) terms enriched among differentially translated genes identified by Ribo-Seq data. The “LogFC” scale represents the normalized enrichment significance of each pathway (range, 0–1). B , polysome gradient traces from AGS and FHIT-overexpressing AGS cells separated on a 10% to 50% linear sucrose gradient. Western blot analysis confirms FHIT protein expression in both cells. C , RT–qPCR analysis of KDM6B mRNA distribution across polysome fractions in AGS cells with or without FHIT overexpression. mRNA levels were normalized to spiked luciferase RNA and plotted as the percentage of total mRNA in each gradient fraction relative to the input. D – F , RT–qPCR analysis of KDM6B mRNA expression and Western blot detection of KDM6B protein levels in gastric cancer cells or NOC-transformed cells with or without FHIT overexpression. G , Western blot analysis of KDM6B protein levels in NOC-transformed cells (GES-1-MNU) transfected with empty vector (EV) or FHIT-overexpressing constructs after treatment with the transcriptional inhibitor ActD to block nascent mRNA synthesis. The analyses were repeated three times, and the results were expressed as mean ± SD. ns, not significant, ∗ p < 0.05, and ∗∗ p < 0.01. ActD, actinomycin D; FHIT, fragile histidine triad; KDM6B, lysine-specific demethylase 6B; MNU, N -nitroso- N -methylurea; qPCR, quantitative PCR; Ribo-Seq, ribosome sequencing.

Journal: The Journal of Biological Chemistry

Article Title: Fragile histidine triad (FHIT) suppresses gastric cancer via translational regulation of the epigenetic modifier KDM6B

doi: 10.1016/j.jbc.2026.111404

Figure Lengend Snippet: FHIT regulates the downstream target KDM6B by mediating its mRNA translation. A , Circos plot displaying Gene Ontology (GO) terms enriched among differentially translated genes identified by Ribo-Seq data. The “LogFC” scale represents the normalized enrichment significance of each pathway (range, 0–1). B , polysome gradient traces from AGS and FHIT-overexpressing AGS cells separated on a 10% to 50% linear sucrose gradient. Western blot analysis confirms FHIT protein expression in both cells. C , RT–qPCR analysis of KDM6B mRNA distribution across polysome fractions in AGS cells with or without FHIT overexpression. mRNA levels were normalized to spiked luciferase RNA and plotted as the percentage of total mRNA in each gradient fraction relative to the input. D – F , RT–qPCR analysis of KDM6B mRNA expression and Western blot detection of KDM6B protein levels in gastric cancer cells or NOC-transformed cells with or without FHIT overexpression. G , Western blot analysis of KDM6B protein levels in NOC-transformed cells (GES-1-MNU) transfected with empty vector (EV) or FHIT-overexpressing constructs after treatment with the transcriptional inhibitor ActD to block nascent mRNA synthesis. The analyses were repeated three times, and the results were expressed as mean ± SD. ns, not significant, ∗ p < 0.05, and ∗∗ p < 0.01. ActD, actinomycin D; FHIT, fragile histidine triad; KDM6B, lysine-specific demethylase 6B; MNU, N -nitroso- N -methylurea; qPCR, quantitative PCR; Ribo-Seq, ribosome sequencing.

Article Snippet: RNA transcriptome sequencing was then performed on each sample by Novogene Corporation.

Techniques: Western Blot, Expressing, Quantitative RT-PCR, Over Expression, Luciferase, Transformation Assay, Transfection, Plasmid Preparation, Construct, Blocking Assay, Real-time Polymerase Chain Reaction, Sequencing