Journal: Cancer letters

Article Title: Glutamine transporter SLC38A3 promotes breast cancer metastasis via Gsk3β/β-catenin/EMT pathway

doi: 10.1016/j.canlet.2024.216653

Figure Lengend Snippet: ( A ) Heatmap of Log normalized mRNA levels of key genes involved in glutamine metabolism in a panel of breast cancer cell lines. Each cell line is shown with three biological replicates. The Log normalized mRNA levels are shown in different colors from green to red with increasing gene expression. ( B ) Protein expression level of SLC38A3 in a panel of breast cancer cell lines. GAPDH was used as a control to confirm equal loading of protein. ( C ) Distant metastasis-free survival rates of breast cancer patients with low (n=1388) or high (n=1377) expression levels of SLC38A3. The survival curve was obtained from KM-plotter.

Article Snippet: The shift in molecular weight of SLC38A3 protein is consistent with the protein validation of SLC38A3 in ‘The Human Protein Atlas’ and could be due to the glycosylation of SLC38A3 protein ( 23 , 31 – 33 ).

Techniques: Gene Expression, Expressing, Control

Journal: Cancer letters

Article Title: Glutamine transporter SLC38A3 promotes breast cancer metastasis via Gsk3β/β-catenin/EMT pathway

doi: 10.1016/j.canlet.2024.216653

Figure Lengend Snippet: ( A ) Quantitative RT-PCR showing mRNA levels of SLC38A3 in MDA-MB-468 and SUM159PT cell lines treated with SLC38A3 siRNA or scrambled siRNA. ( B ) Protein expression level of SLC38A3 in MDA-MB-468 and SUM159PT cell lines treated with SLC38A3 siRNA or scrambled siRNA. GAPDH was used as a control to confirm equal loading of protein. ( C ) Cellular content of glutamine and some associated metabolites in MDA-MB-468 and SUM159PT cell lines transfected with SLC38A3 siRNA or scrambled siRNA quantified by using high-resolution 1 H MRS. ( D ) Cell viability of MDA-MB-468 and SUM159PT cell lines treated with SLC38A3 siRNA or scrambled siRNA. ( E ) Mean values of DCFDA intensity in MDA-MB-468 and SUM159PT cell lines treated with SLC38A3 siRNA or scrambled siRNA. ( F ) Mean values of MitoSOX intensity in MDA-MB-468 and SUM159PT cell lines treated with SLC38A3 siRNA or scrambled siRNA. ( G ) Percentage of Annexin V-positive cells of MDA-MB-468 and SUM159PT treated with SLC38A3 siRNA or scrambled siRNA. All data are shown as mean values ± S.D. of three independent experiments. *p<0.05, **

Article Snippet: The shift in molecular weight of SLC38A3 protein is consistent with the protein validation of SLC38A3 in ‘The Human Protein Atlas’ and could be due to the glycosylation of SLC38A3 protein ( 23 , 31 – 33 ).

Techniques: Quantitative RT-PCR, Expressing, Control, Transfection

Journal: Cancer letters

Article Title: Glutamine transporter SLC38A3 promotes breast cancer metastasis via Gsk3β/β-catenin/EMT pathway

doi: 10.1016/j.canlet.2024.216653

Figure Lengend Snippet: ( A ) Representative images and ( B ) numbers of migrated and invaded cells of MDA-MB-468 and SUM159PT cell lines treated with SLC38A3 siRNA or scrambled siRNA. Cells were counted using Image J. ( C ) Protein expression levels of SLC38A3, N-cadherin, E-cadherin, vimentin, phospho-Gsk3β (ser9), Gsk3β, phospho-β-catenin (ser33/37/Thr41) and β-catenin in MDA-MB-468 and SUM159PT cell lines treated with SLC38A3 siRNA or scrambled siRNA. GAPDH was used as loading control. ( D ) Quantitative RT-PCR showing mRNA levels of CDH1, CDH2, VIM, SNAI1, SNAI2, ZEB1, ZEB2, TWIST1 and TWIST2 in MDA-MB-468 and SUM159PT cell lines treated with SLC38A3 siRNA or scrambled siRNA. All data are shown as mean values ± S.D. of three independent experiments. *p<0.05, **

Article Snippet: The shift in molecular weight of SLC38A3 protein is consistent with the protein validation of SLC38A3 in ‘The Human Protein Atlas’ and could be due to the glycosylation of SLC38A3 protein ( 23 , 31 – 33 ).

Techniques: Migration, Expressing, Control, Quantitative RT-PCR

Journal: Cancer letters

Article Title: Glutamine transporter SLC38A3 promotes breast cancer metastasis via Gsk3β/β-catenin/EMT pathway

doi: 10.1016/j.canlet.2024.216653

Figure Lengend Snippet: ( A ) Quantitative RT-PCR showing mRNA levels of SLC38A3 in stably SLC38A3-Myc- and empty vector (EV)- expressing MDA-MB-231 and SUM159PT cell lines. ( B ) Protein expression levels of SLC38A3 and Myc-tag in stably SLC38A3-Myc- and EV-expressing MDA-MB-231 and SUM159PT cell lines. GAPDH was used as a control to confirm equal loading of protein. ( C ) Cell viability of stably SLC38A3-Myc- and EV-expressing MDA-MB-231 and SUM159PT cell lines. ( D ) Cellular content of glutamine and associated metabolites in stably SLC38A3-Myc- and EV-expressing MDA-MB-231 and SUM159PT cell lines measured with high-resolution 1 H MRS. ( E ) Representative images of migrated and invaded cells of stably SLC38A3-Myc-and EV-expressing MDA-MB-231 and SUM159PT cell lines. ( F ) Number of migrated and invaded cells of stable SLC38A3-Myc- and EV-expressing MDA-MB-231 and SUM159PT cell lines. Cells were counted using Image J. ( G ) Protein expression levels of SLC38A3, N-cadherin, E-cadherin, vimentin, phospho-Gsk3β (ser9), Gsk3β, phospho-β-catenin (ser33/37/Thr41) and β-catenin in stably SLC38A3-Myc- and EV-expressing MDA-MB-231 and SUM159PT cell lines. GAPDH was used as loading control. ( H ) Quantitative RT-PCR showing mRNA levels of CDH1, CDH2, VIM, SNAI1, SNAI2, ZEB1, ZEB2, TWIST1 and TWIST2 in stable SLC38A3-Myc- and EV-expressing MDA-MB-231 and SUM159PT cell lines. All data are shown as mean values ± S.D. of three independent experiments. *p<0.05, **

Article Snippet: The shift in molecular weight of SLC38A3 protein is consistent with the protein validation of SLC38A3 in ‘The Human Protein Atlas’ and could be due to the glycosylation of SLC38A3 protein ( 23 , 31 – 33 ).

Techniques: Over Expression, Migration, Quantitative RT-PCR, Stable Transfection, Plasmid Preparation, Expressing, Control

Journal: Cancer letters

Article Title: Glutamine transporter SLC38A3 promotes breast cancer metastasis via Gsk3β/β-catenin/EMT pathway

doi: 10.1016/j.canlet.2024.216653

Figure Lengend Snippet: ( A ) Tumor growth curves of MDA-MB-231 negative control (NC), MDA-MB-231 shSLC38A3#4, SUM159PT NC and SUM159PT shSLC38A3#4 xenografts. Tumor volume analysis is shown as mean value ± S.D. of eight independent experiments each. ( B ) Tumor weight analysis of all tumors shown in (A). Data are shown as mean values ± S.D. of eight independent experiments each. ( C ) Real-time PCR showing mRNA levels of SLC38A3 in tumor tissues of above xenografts. Data are shown as mean values ± S.D. of eight independent experiments each. ( D ) High-resolution 1 H MRS quantification of cellular contents of glutamine and its associated metabolites in tumor tissues of all primary tumors in our study. Data are shown as mean values ± S.D. of five independent experiments each. *p<0.05, **

Article Snippet: The shift in molecular weight of SLC38A3 protein is consistent with the protein validation of SLC38A3 in ‘The Human Protein Atlas’ and could be due to the glycosylation of SLC38A3 protein ( 23 , 31 – 33 ).

Techniques: Negative Control, Real-time Polymerase Chain Reaction

Journal: Cancer letters

Article Title: Glutamine transporter SLC38A3 promotes breast cancer metastasis via Gsk3β/β-catenin/EMT pathway

doi: 10.1016/j.canlet.2024.216653

Figure Lengend Snippet: ( A ) Representative images of spontaneous lung metastases in mice growing primary orthotopic MDA-MB-231 NC versus MDA-MB-231 shSLC38A3#4 tumors. ( B ) Average number of lung nodules per microscopy slide in lungs from MDA-MB-231 NC versus MDA-MB-231 shSLC38A3#4 inoculated mice. ( C ) Representative images of spontaneous liver metastases in mice growing primary orthotopic MDA-MB-231 NC versus MDA-MB-231 shSLC38A3#4 tumors. ( D ) Average number of liver nodules per microscopy slide in MDA-MB-231 NC versus MDA-MB-231 shSLC38A3#4 inoculated mice. ( E ) Protein expression levels of SLC38A3, N-cadherin, E-cadherin, and vimentin in MDA-MB-231 NC versus MDA-MB-231 shSLC38A3#4 xenografts and SUM159PT NC versus SUM159PT shSLC38A3#4 xenografts. β-actin was used as loading control. All quantitative data are shown as mean values ± S.D. of 5 independent experiments. *p<0.05, **

Article Snippet: The shift in molecular weight of SLC38A3 protein is consistent with the protein validation of SLC38A3 in ‘The Human Protein Atlas’ and could be due to the glycosylation of SLC38A3 protein ( 23 , 31 – 33 ).

Techniques: Microscopy, Expressing, Control

Journal: Cancer letters

Article Title: Glutamine transporter SLC38A3 promotes breast cancer metastasis via Gsk3β/β-catenin/EMT pathway

doi: 10.1016/j.canlet.2024.216653

Figure Lengend Snippet: SLC38A3 increases the influx and metabolism of neutral amino acids including glutamine and promotes GSH production in breast cancer, which activates mTOR/Gsk3β and β-catenin signaling in breast cancer. The activation of Gsk3β and β-catenin upregulates EMT-inducing transcription factors, which promotes metastasis of breast cancer.

Article Snippet: The shift in molecular weight of SLC38A3 protein is consistent with the protein validation of SLC38A3 in ‘The Human Protein Atlas’ and could be due to the glycosylation of SLC38A3 protein ( 23 , 31 – 33 ).

Techniques: Activation Assay