Journal: Molecular Biomedicine

Article Title: Fat mass and obesity-associated protein (FTO) mediated m 6 A modification of circFAM192A promoted gastric cancer proliferation by suppressing SLC7A5 decay

doi: 10.1186/s43556-024-00172-4

Figure Lengend Snippet: FTO was upregulated in GC and predicted a poor prognosis. a ELISA assays showed the m 6 A level in gastric cancer tissues and normal tissues; b QPCR assays displayed the FTO mRNA level in 51 pairs of GC samples from patients; c The chart exhibited the level of FTO protein in GC samples; d Representative IHC images of paired GC samples with FTO antibody, the scale bar is 20 μm; e The expression pattern of FTO in STAD, ANT and normal tissues with data from TCGA and GTEx database; f The OS (left) and PFS (right) curve of GC patients with different FTO expression drawn with data from TCGA. Quantitative data from three independent experiments are shown as the mean ± SD (error bars). * P < 0.05, ** P < 0.01, *** P < 0.001 (Student’s t-test). STAD: stomach adenocarcinoma; ANT:adjacent normal tissues

Article Snippet: Primary Antibodies included mouse anti-FTO (1:1000, Abcam, USA), mouse anti-SLC7A5 (1:50, santa curz, USA), rabbit anti-YTHDC2 (1:1000, proteintech, China), mouse anti-YTHDF2 (1:1000, proteintech), rabbit anti-mTOR (1:1000, CST), rabbit anti-p-mTOR mouse (1:1000, CST), rabbit anti-p-p70 (1:1000, CST), antiGAPDH (1:20000, Beyotime).

Techniques: Enzyme-linked Immunosorbent Assay, Expressing

Journal: Molecular Biomedicine

Article Title: Fat mass and obesity-associated protein (FTO) mediated m 6 A modification of circFAM192A promoted gastric cancer proliferation by suppressing SLC7A5 decay

doi: 10.1186/s43556-024-00172-4

Figure Lengend Snippet: FTO regulated circFAM192A expression through the m 6 A dependent manner. a The expression of pre-mRNA and linear FAM192A upon FTO knockout; b The expression of pre-mRNA and linear FAM192A upon FTO overexpression; c Pull down assays exhibited the quantity of FTO protein captured by circFAM192A targeting probes and negative control probes; d CircFAM192A captured by anti-FTO antibody in RIP assays; e Drawing of luciferase plasmids with different mutant binding site; f The relative luciferase activity in HEK-293 T cells transfected with plasmids containing different mutant m 6 A sites; g The m. 6 A abundance in circFAM192A upon FTO knockout or overexpression; h The stability of circFAM192A under the condition of FTO knockout (left) and overexpression (right) in ActD treatment assays; i QPCR assays showed the level of circFAM192A when knocking down YTHDF1/2/3, YTHDC2; j The enrichment between circFAM192A and anti-YTHDF2 antibody in RIP assays; k RIP assays showed the level of circFAM192A captured by anti-YTHDF2 antibody after knocking out or ovexpressing FTO; l Pull down assays showed the level of YTHDF2 or YTHDC2 captured by circFAM192A targeting probes and negative probes; m QPCR assays showed the level of circFAM192A when knocking down POP1. Quantitative data from three independent experiments are shown as the mean ± SD (error bars). * P < 0.05, ** P < 0.01, *** P < 0.001 (Student’s t-test)

Article Snippet: Primary Antibodies included mouse anti-FTO (1:1000, Abcam, USA), mouse anti-SLC7A5 (1:50, santa curz, USA), rabbit anti-YTHDC2 (1:1000, proteintech, China), mouse anti-YTHDF2 (1:1000, proteintech), rabbit anti-mTOR (1:1000, CST), rabbit anti-p-mTOR mouse (1:1000, CST), rabbit anti-p-p70 (1:1000, CST), antiGAPDH (1:20000, Beyotime).

Techniques: Expressing, Knock-Out, Over Expression, Negative Control, Luciferase, Mutagenesis, Binding Assay, Activity Assay, Transfection

Journal: Molecular Biomedicine

Article Title: Fat mass and obesity-associated protein (FTO) mediated m 6 A modification of circFAM192A promoted gastric cancer proliferation by suppressing SLC7A5 decay

doi: 10.1186/s43556-024-00172-4

Figure Lengend Snippet: FTO promoted GC proliferation through the circFAM192A/SLC7A5 axis. a - c CCK8 ( a ), EdU assays ( b ) and colony formation ( c ) assays showed the growth ability of controlled, circFAM192A overexpressed, circFAM192A overexpressed but SLC7A5 silenced MGC803 and AGS cells; d WB assays revealed the alteration of key molecules in mTOR signaling pathway in controlled, FTO overexpressed, FTO overexpressed but circFAM192A silenced MGC803 and AGS cells; e WB assays revealed the alteration of key molecules in mTOR signaling pathway in controlled, circFAM192A overexpressed, circFAM192A overexpressed but SLC7A5 silenced MGC803 and AGS cells; f The level of SLC7A5 in tumors with FTO overexpressed and cirFAM192A overexpressed in IHC assays (scale bar, 20 μm); g The expression of SLC7A5 in tumors with FTO knockout and cirFAM192A knockdown in IHC assays (scale bar, 20 μm). Quantitative data from three independent experiments are shown as the mean ± SD (error bars). * P < 0.05, ** P < 0.01, *** P < 0.001 (Student’s t-test)

Article Snippet: Primary Antibodies included mouse anti-FTO (1:1000, Abcam, USA), mouse anti-SLC7A5 (1:50, santa curz, USA), rabbit anti-YTHDC2 (1:1000, proteintech, China), mouse anti-YTHDF2 (1:1000, proteintech), rabbit anti-mTOR (1:1000, CST), rabbit anti-p-mTOR mouse (1:1000, CST), rabbit anti-p-p70 (1:1000, CST), antiGAPDH (1:20000, Beyotime).

Techniques: Expressing, Knock-Out

Journal: Molecular Biomedicine

Article Title: Fat mass and obesity-associated protein (FTO) mediated m 6 A modification of circFAM192A promoted gastric cancer proliferation by suppressing SLC7A5 decay

doi: 10.1186/s43556-024-00172-4

Figure Lengend Snippet: FB23-2 inhibited GC proliferation in vitro and in vivo. a The inhibitory effect of FB23-2 with different concentration on MGC803 and AGS cells; (b-d) CCK8 ( b ), colony formation ( c ) and EdU assays ( d ) showed the growth ability of controlled and FB23-2 treated cells (10 μM on MGC803 and 2 μM on AGS); e Tumors from mice injected with DMSO and FB23-2 (2 mg/kg); f The growth rate of GC cells in vivo after FB23-2 or DMSO treatment; g The expression of circFAM192A in FB23-2 or DMSO treated tumors; h WB assays showed the expression of SLC7A5, p-p70 and p-mTOR in FB23-2 or DMSO treated tumors; i Schematic of the action of FTO regulating gastric cancer proliferation. Quantitative data from three independent experiments are shown as the mean ± SD (error bars). * P < 0.05, ** P < 0.01, *** P < 0.001 (Student’s t-test)

Article Snippet: Primary Antibodies included mouse anti-FTO (1:1000, Abcam, USA), mouse anti-SLC7A5 (1:50, santa curz, USA), rabbit anti-YTHDC2 (1:1000, proteintech, China), mouse anti-YTHDF2 (1:1000, proteintech), rabbit anti-mTOR (1:1000, CST), rabbit anti-p-mTOR mouse (1:1000, CST), rabbit anti-p-p70 (1:1000, CST), antiGAPDH (1:20000, Beyotime).

Techniques: In Vitro, In Vivo, Concentration Assay, Injection, Expressing