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Effects of alanine-serine-cysteine transporter 2 on proliferation and invasion of hepatocellular carcinoma

  • 1.

    Department of Pathophysiology, School of Basic Medical Science, Anhui Medical University, Hefei 230032, China

  • 2.

    Department of Hematology, The First Affiliated Hospital of USTC(Anhui Provincial Hospital), Hefei 230001, China

  • 3.

    School of Basic Medicine, Division of Life Sciences and Medicine, University of Science and Technology of China, Hefei 230027, China

Cite this:
https://doi.org/10.52396/JUST-2021-0033
  • Received Date: 28 January 2021
  • Rev Recd Date: 20 February 2021
  • Publish Date: 28 February 2021
    Abstract

  • Abstract

    Metabolic reprogramming is a major feature of tumors, and tumor cells adapt to their glutamine needs by up-regulating the alanine-serine-cysteine transporter 2(ASCT2). It was found that the ASCT2 expression in hepatocellular carcinoma (HCC) tissues was significantly higher than that in normal liver tissues. In addition, the higher expression of ASCT2 in HCC patients was closely associated with poor survival.The knockdown of ASCT2 inhibited the proliferation, clone formation, migration and invasion of HCC cells in vitro.Cell cycle analysis suggested that knockdown of ASCT2 inhibited the proportion of HCC cells in the S phase. In vivo tumorigenic assay confirmed that the knockdown of ASCT2 in HCC cells could significantly inhibit tumor growth.Further studies showed that the knockdown of ASCT2 significantly reduced mitochondrial oxidative phosphorylation(OXPHOS),ATP production,and the phosphorylation level of AKT/S6 in HCC cells.Overall, our results showed that knockdown of ASCT2 could inhibit the malignancy of HCC cells. In addition, the mitochondrial metabolism and phosphorylation level of the AKT/S6 signaling pathway of HCC cells were also inhibited following the ASCT2 inhibition, suggesting that the dysregulated mitochondrial metabolism and abnormal activation of AKT/S6 signaling pathway were closely associated with the HCC progression.

    Abstract

    Metabolic reprogramming is a major feature of tumors, and tumor cells adapt to their glutamine needs by up-regulating the alanine-serine-cysteine transporter 2(ASCT2). It was found that the ASCT2 expression in hepatocellular carcinoma (HCC) tissues was significantly higher than that in normal liver tissues. In addition, the higher expression of ASCT2 in HCC patients was closely associated with poor survival.The knockdown of ASCT2 inhibited the proliferation, clone formation, migration and invasion of HCC cells in vitro.Cell cycle analysis suggested that knockdown of ASCT2 inhibited the proportion of HCC cells in the S phase. In vivo tumorigenic assay confirmed that the knockdown of ASCT2 in HCC cells could significantly inhibit tumor growth.Further studies showed that the knockdown of ASCT2 significantly reduced mitochondrial oxidative phosphorylation(OXPHOS),ATP production,and the phosphorylation level of AKT/S6 in HCC cells.Overall, our results showed that knockdown of ASCT2 could inhibit the malignancy of HCC cells. In addition, the mitochondrial metabolism and phosphorylation level of the AKT/S6 signaling pathway of HCC cells were also inhibited following the ASCT2 inhibition, suggesting that the dysregulated mitochondrial metabolism and abnormal activation of AKT/S6 signaling pathway were closely associated with the HCC progression.
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    • References(44)
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    Sim H W, Knox J. Hepatocellular carcinoma in the era of immunotherapy. Curr. Probl. Cancer, 2018, 42(1): 40-48.
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    Brown K K, Spinelli J B, Asara J M, et al. Adaptive reprogramming of de novo pyrimidine synthesis is a metabolic vulnerability in triple-negative breast cancer. Cancer Discov., 2017, 7(4): 391-399.
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    Alberghina L, Gaglio D. Redox control of glutamine utilization in cancer. Cell Death Dis., 2014, 5: e1561.
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    Geck R C, Toker A. Nonessential amino acid metabolism in breast cancer. Adv. Biol. Regul., 2016, 62: 11-17.
    [31]
    Yoo H C, Park S J, Nam M, et al. A variant of SLC1A5 is a mitochondrial glutamine transporter for metabolic reprogramming in cancer cells. Cell Metab., 2020, 31(2): 267-283.
    [32]
    Broer A, Gauthier-Coles G, Rahimi F, et al. Ablation of the ASCT2 (SLC1A5) gene encoding a neutral amino acid transporter reveals transporter plasticity and redundancy in cancer cells. J. Biol. Chem., 2019, 294(11): 4012-4026.
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    Bothwell P J, Kron C D, Wittke E F, et al. Targeted suppression and knockout of ASCT2 or LAT1 in epithelial and mesenchymal human liver cancer cells fail to inhibit growth. Int. J. Mol. Sci., 2018, 19(7): 2093.
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    Lee P, Malik D, Perkons N, et al. Targeting glutamine metabolism slows soft tissue sarcoma growth. Nat. Commun., 2020, 11(1): 498.
    [35]
    Wang V M, Ferreira R, Almagro J, et al. CD9 identifies pancreatic cancer stem cells and modulates glutamine metabolism to fuel tumour growth. Nat. Cell Biol., 2019, 21(11): 1425-1435.
    [36]
    Wang Y, Bai C, Ruan Y, et al. Coordinative metabolism of glutamine carbon and nitrogen in proliferating cancer cells under hypoxia. Nat. Commun., 2019, 10(1): 201.
    [37]
    Biancur D E, Paulo J A, Malachowska B, et al. Compensatory metabolic networks in pancreatic cancers upon perturbation of glutamine metabolism. Nat. Commun., 2017, 8: 15965.
    [38]
    Pochini L, Scalise M, Galluccio M, et al. Membrane transporters for the special amino acid glutamine: Structure/function relationships and relevance to human health. Front. Chem., 2014, 2: 61.
    [39]
    Hoxhaj G, Manning B D. The PI3K-AKT network at the interface of oncogenic signalling and cancer metabolism. Nat. Rev. Cancer, 2020, 20(2): 74-88.
    [40]
    Pereira O, Teixeira A, Sampaio-Marques B, et al. Signalling mechanisms that regulate metabolic profile and autophagy of acute myeloid leukaemia cells. J. Cell Mol. Med., 2018, 22(10): 4807-4817.
    [41]
    Stiles B L. PI-3-K and AKT: Onto the mitochondria. Adv. Drug Deliv. Rev., 2009, 61(14): 1276-1282.
    [42]
    Robey R B, Hay N. Mitochondrial hexokinases, novel mediators of the antiapoptotic effects of growth factors and Akt. Oncogene, 2006, 25(34): 4683-4696.
    [43]
    Pelicano H, Xu R H, Du M, et al. Mitochondrial respiration defects in cancer cells cause activation of Akt survival pathway through a redox-mediated mechanism. J. Cell Biol., 2006, 175(6): 913-923.
    [44]
    Li T, Han J, Jia L, et al. PKM2 coordinates glycolysis with mitochondrial fusion and oxidative phosphorylation. Protein Cell, 2019, 10(8): 583-594.
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    [1]
    Bray F, Ferlay J, Soerjomataram I, et al. Global cancer statistics 2018: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA Cancer J. Clin., 2018, 68(6): 394-424.
    [2]
    Kulik L, El-Serag H B. Epidemiology and management of hepatocellular carcinoma. Gastroenterology, 2019, 156(2): 477-491.
    [3]
    Yarchoan M, Agarwal P, Villanueva A, et al. Recent developments and therapeutic strategies against hepatocellular carcinoma. Cancer Res., 2019, 79(17): 4326-4330.
    [4]
    Dai W, Xu L, Yu X, et al. OGDHL silencing promotes hepatocellular carcinoma by reprogramming glutamine metabolism. J. Hepatol., 2020, 72(5): 909-923.
    [5]
    Yu L, Kim J, Jiang L, et al. MTR4 drives liver tumorigenesis by promoting cancer metabolic switch through alternative splicing. Nat. Commun., 2020, 11(1): 708.
    [6]
    Gu L, Zhu Y, Lin X, et al. The IKKβ-USP30-ACLY axis controls lipogenesis and tumorigenesis. Hepatology, 2021,73(1):160-174.
    [7]
    Deberardinis R J, Lum J J, Hatzivassiliou G, et al. The biology of cancer: Metabolic reprogramming fuels cell growth and proliferation. Cell Metab., 2008, 7(1): 11-20.
    [8]
    Geck R C, Toker A. Nonessential amino acid metabolism in breast cancer. Adv. Biol. Regul., 2016, 62: 11-17.
    [9]
    Yoo H C, Park S J, Nam M, et al. A variant of SLC1A5 is a mitochondrial glutamine transporter for metabolic reprogramming in cancer cells. Cell Metab., 2020, 31(2): 267-283.
    [10]
    Wise D R, Thompson C B. Glutamine addiction: A new therapeutic target in cancer. Trends Biochem. Sci., 2010, 35(8): 427-433.
    [11]
    Cassago A, Ferreira A P, Ferreira I M, et al. Mitochondrial localization and structure-based phosphate activation mechanism of Glutaminase C with implications for cancer metabolism. Proc. Natl. Acad. Sci. U. S. A., 2012, 109(4): 1092-1097.
    [12]
    Udagawa M, Horie Y, Hirayama C. Aberrant porphyrin metabolism in hepatocellular carcinoma. Biochem. Med., 1984, 31(2): 131-139.
    [13]
    Gao P, Tchernyshyov I, Chang T C, et al. c-Myc suppression of miR-23a/b enhances mitochondrial glutaminase expression and glutamine metabolism. Nature, 2009, 458(7239): 762-765.
    [14]
    Son J, Lyssiotis C A, Ying H, et al. Glutamine supports pancreatic cancer growth through a KRAS-regulated metabolic pathway. Nature, 2013, 496(7443): 101-105.
    [15]
    Wise D R, Deberardinis R J, Mancuso A, et al. Myc regulates a transcriptional program that stimulates mitochondrial glutaminolysis and leads to glutamine addiction. Proc. Natl. Acad. Sci. U. S. A., 2008, 105(48): 18782-18787.
    [16]
    Arriza J L, Kavanaugh M P, Fairman W A, et al. Cloning and expression of a human neutral amino acid transporter with structural similarity to the glutamate transporter gene family. J. Biol. Chem., 1993, 268(21): 15329-15332.
    [17]
    Nakaya M, Xiao Y, Zhou X, et al. Inflammatory T cell responses rely on amino acid transporter ASCT2 facilitation of glutamine uptake and mTORC1 kinase activation. Immunity, 2014, 40(5): 692-705.
    [18]
    Scalise M, Pochini L, Galluccio M, et al. Glutamine transport and mitochondrial metabolism in cancer cell growth. Front. Oncol., 2017, 7: 306.
    [19]
    Scalise M, Pochini L, Panni S, et al. Transport mechanism and regulatory properties of the human amino acid transporter ASCT2 (SLC1A5). Amino Acids, 2014, 46(11): 2463-2475.
    [20]
    Ren P, Yue M, Xiao D, et al. ATF4 and N-Myc coordinate glutamine metabolism in MYCN-amplified neuroblastoma cells through ASCT2 activation. J. Pathol., 2015, 235(1): 90-100.
    [21]
    van Geldermalsen M, Wang Q, Nagarajah R, et al. ASCT2/SLC1A5 controls glutamine uptake and tumour growth in triple-negative basal-like breast cancer. Oncogene, 2016, 35(24): 3201-3208.
    [22]
    Ye J, Huang Q, Xu J, et al. Targeting of glutamine transporter ASCT2 and glutamine synthetase suppresses gastric cancer cell growth. J. Cancer Res. Clin. Oncol., 2018, 144(5): 821-833.
    [23]
    Sun H W, Yu X J, Wu W C, et al. GLUT1 and ASCT2 as predictors for prognosis of hepatocellular carcinoma. PLoS One, 2016, 11(12): e168907.
    [24]
    Hu W, Feng Z. The role of p53 in reproduction, an unexpected function for a tumor suppressor. J. Mol. Cell Biol., 2019, 11(7): 624-627.
    [25]
    Icard P, Fournel L, Wu Z, et al. Interconnection between metabolism and cell cycle in cancer. Trends Biochem. Sci., 2019, 44(6): 490-501.
    [26]
    Han T S, Ban H S, Hur K, et al. The epigenetic regulation of HCC metastasis. Int. J. Mol. Sci., 2018, 19(12): 3978.
    [27]
    Sim H W, Knox J. Hepatocellular carcinoma in the era of immunotherapy. Curr. Probl. Cancer, 2018, 42(1): 40-48.
    [28]
    Brown K K, Spinelli J B, Asara J M, et al. Adaptive reprogramming of de novo pyrimidine synthesis is a metabolic vulnerability in triple-negative breast cancer. Cancer Discov., 2017, 7(4): 391-399.
    [29]
    Alberghina L, Gaglio D. Redox control of glutamine utilization in cancer. Cell Death Dis., 2014, 5: e1561.
    [30]
    Geck R C, Toker A. Nonessential amino acid metabolism in breast cancer. Adv. Biol. Regul., 2016, 62: 11-17.
    [31]
    Yoo H C, Park S J, Nam M, et al. A variant of SLC1A5 is a mitochondrial glutamine transporter for metabolic reprogramming in cancer cells. Cell Metab., 2020, 31(2): 267-283.
    [32]
    Broer A, Gauthier-Coles G, Rahimi F, et al. Ablation of the ASCT2 (SLC1A5) gene encoding a neutral amino acid transporter reveals transporter plasticity and redundancy in cancer cells. J. Biol. Chem., 2019, 294(11): 4012-4026.
    [33]
    Bothwell P J, Kron C D, Wittke E F, et al. Targeted suppression and knockout of ASCT2 or LAT1 in epithelial and mesenchymal human liver cancer cells fail to inhibit growth. Int. J. Mol. Sci., 2018, 19(7): 2093.
    [34]
    Lee P, Malik D, Perkons N, et al. Targeting glutamine metabolism slows soft tissue sarcoma growth. Nat. Commun., 2020, 11(1): 498.
    [35]
    Wang V M, Ferreira R, Almagro J, et al. CD9 identifies pancreatic cancer stem cells and modulates glutamine metabolism to fuel tumour growth. Nat. Cell Biol., 2019, 21(11): 1425-1435.
    [36]
    Wang Y, Bai C, Ruan Y, et al. Coordinative metabolism of glutamine carbon and nitrogen in proliferating cancer cells under hypoxia. Nat. Commun., 2019, 10(1): 201.
    [37]
    Biancur D E, Paulo J A, Malachowska B, et al. Compensatory metabolic networks in pancreatic cancers upon perturbation of glutamine metabolism. Nat. Commun., 2017, 8: 15965.
    [38]
    Pochini L, Scalise M, Galluccio M, et al. Membrane transporters for the special amino acid glutamine: Structure/function relationships and relevance to human health. Front. Chem., 2014, 2: 61.
    [39]
    Hoxhaj G, Manning B D. The PI3K-AKT network at the interface of oncogenic signalling and cancer metabolism. Nat. Rev. Cancer, 2020, 20(2): 74-88.
    [40]
    Pereira O, Teixeira A, Sampaio-Marques B, et al. Signalling mechanisms that regulate metabolic profile and autophagy of acute myeloid leukaemia cells. J. Cell Mol. Med., 2018, 22(10): 4807-4817.
    [41]
    Stiles B L. PI-3-K and AKT: Onto the mitochondria. Adv. Drug Deliv. Rev., 2009, 61(14): 1276-1282.
    [42]
    Robey R B, Hay N. Mitochondrial hexokinases, novel mediators of the antiapoptotic effects of growth factors and Akt. Oncogene, 2006, 25(34): 4683-4696.
    [43]
    Pelicano H, Xu R H, Du M, et al. Mitochondrial respiration defects in cancer cells cause activation of Akt survival pathway through a redox-mediated mechanism. J. Cell Biol., 2006, 175(6): 913-923.
    [44]
    Li T, Han J, Jia L, et al. PKM2 coordinates glycolysis with mitochondrial fusion and oxidative phosphorylation. Protein Cell, 2019, 10(8): 583-594.
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