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Glutamate and asparagine cataplerosis underlie glutamine addiction in melanoma.

Ratnikov B, Aza-Blanc P, Ronai ZA, Smith JW, Osterman AL, Scott DA - Oncotarget (2015)

Bottom Line: Glutamine dependence is a prominent feature of cancer metabolism, and here we show that melanoma cells, irrespective of their oncogenic background, depend on glutamine for growth.In the absence of glutamine, TCA cycle metabolites were liable to depletion through aminotransferase-mediated α-ketoglutarate-to-glutamate conversion and glutamate secretion.Melanocytes use more glutamine for protein synthesis rather than secreting it as glutamate and are less prone to loss of glutamate and TCA cycle metabolites when starved of glutamine.

View Article: PubMed Central - PubMed

Affiliation: Sanford-Burnham Medical Research Institute, La Jolla, California 92037, USA.

ABSTRACT
Glutamine dependence is a prominent feature of cancer metabolism, and here we show that melanoma cells, irrespective of their oncogenic background, depend on glutamine for growth. A quantitative audit of how carbon from glutamine is used showed that TCA-cycle-derived glutamate is, in most melanoma cells, the major glutamine-derived cataplerotic output and product of glutaminolysis. In the absence of glutamine, TCA cycle metabolites were liable to depletion through aminotransferase-mediated α-ketoglutarate-to-glutamate conversion and glutamate secretion. Aspartate was an essential cataplerotic output, as melanoma cells demonstrated a limited capacity to salvage external aspartate. Also, the absence of asparagine increased the glutamine requirement, pointing to vulnerability in the aspartate-asparagine biosynthetic pathway within melanoma metabolism. In contrast to melanoma cells, melanocytes could grow in the absence of glutamine. Melanocytes use more glutamine for protein synthesis rather than secreting it as glutamate and are less prone to loss of glutamate and TCA cycle metabolites when starved of glutamine.

No MeSH data available.


Related in: MedlinePlus

Glutamine is required for growth and used to sustain TCA cyclemetabolite levels and for aspartate and asparagine synthesis(A) Effects of titrating glutamine in DMEM on growth ofmelanoma cells lines. Mutated oncogenes in cell lines are designated:(B) BRAF; (N) NRAS; (P53) TP53. (B) Growth of Lu1205 cellsin DMEM with varied glutamine and supplementation with 3 mM DMaK, 0.1 mMNEAA, 0.1 mM asparagine (Asn), or 5 mM DMGlu. “NEAA-Asn”indicates addition of a reconstituted NEAA mixture lacking asparagine.Growth is shown relative to growth in medium containing 2 mM glutamine(Mean ± SEM of N = 3). (C)Changes in metabolite pools in Lu1205 cells or melanocytes after 6 h inglutamine-free medium(Gln-), relative to cells with 2 mM glutamine(Gln+). Lu1205 Gln− cultures were supplemented as shownwith 3 mM DMaK, 0.1 mM Asn, 0.1 mM aspartate (Asp), or(“Both”) DMaK and Asn. Source data for part (C) are shownin SupplementaryDataset 1.
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Figure 2: Glutamine is required for growth and used to sustain TCA cyclemetabolite levels and for aspartate and asparagine synthesis(A) Effects of titrating glutamine in DMEM on growth ofmelanoma cells lines. Mutated oncogenes in cell lines are designated:(B) BRAF; (N) NRAS; (P53) TP53. (B) Growth of Lu1205 cellsin DMEM with varied glutamine and supplementation with 3 mM DMaK, 0.1 mMNEAA, 0.1 mM asparagine (Asn), or 5 mM DMGlu. “NEAA-Asn”indicates addition of a reconstituted NEAA mixture lacking asparagine.Growth is shown relative to growth in medium containing 2 mM glutamine(Mean ± SEM of N = 3). (C)Changes in metabolite pools in Lu1205 cells or melanocytes after 6 h inglutamine-free medium(Gln-), relative to cells with 2 mM glutamine(Gln+). Lu1205 Gln− cultures were supplemented as shownwith 3 mM DMaK, 0.1 mM Asn, 0.1 mM aspartate (Asp), or(“Both”) DMaK and Asn. Source data for part (C) are shownin SupplementaryDataset 1.

Mentions: We tested the glutamine dependence of nine melanoma lines with differentoncogenic drivers (4 mutant BRAF, 4 mutant NRAS, 1 mutant p53). All required atleast 1 mM glutamine for maximal growth (Figure 2A) and there was no proliferation in the absence of glutamine(Figure S1A, S1B),while most cell lines could grow in the absence of glucose (Figure S1C). In contrast,growth of melanocytes was similar with or without glutamine (Figure S1D). Asmelanocytes were grown in a melanocyte-specific medium, we checked growth ofLu1205 melanoma cells in this medium and confirmed that growth was substantiallyinhibited by the absence of glutamine (Figure S1E).


Glutamate and asparagine cataplerosis underlie glutamine addiction in melanoma.

Ratnikov B, Aza-Blanc P, Ronai ZA, Smith JW, Osterman AL, Scott DA - Oncotarget (2015)

Glutamine is required for growth and used to sustain TCA cyclemetabolite levels and for aspartate and asparagine synthesis(A) Effects of titrating glutamine in DMEM on growth ofmelanoma cells lines. Mutated oncogenes in cell lines are designated:(B) BRAF; (N) NRAS; (P53) TP53. (B) Growth of Lu1205 cellsin DMEM with varied glutamine and supplementation with 3 mM DMaK, 0.1 mMNEAA, 0.1 mM asparagine (Asn), or 5 mM DMGlu. “NEAA-Asn”indicates addition of a reconstituted NEAA mixture lacking asparagine.Growth is shown relative to growth in medium containing 2 mM glutamine(Mean ± SEM of N = 3). (C)Changes in metabolite pools in Lu1205 cells or melanocytes after 6 h inglutamine-free medium(Gln-), relative to cells with 2 mM glutamine(Gln+). Lu1205 Gln− cultures were supplemented as shownwith 3 mM DMaK, 0.1 mM Asn, 0.1 mM aspartate (Asp), or(“Both”) DMaK and Asn. Source data for part (C) are shownin SupplementaryDataset 1.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4480687&req=5

Figure 2: Glutamine is required for growth and used to sustain TCA cyclemetabolite levels and for aspartate and asparagine synthesis(A) Effects of titrating glutamine in DMEM on growth ofmelanoma cells lines. Mutated oncogenes in cell lines are designated:(B) BRAF; (N) NRAS; (P53) TP53. (B) Growth of Lu1205 cellsin DMEM with varied glutamine and supplementation with 3 mM DMaK, 0.1 mMNEAA, 0.1 mM asparagine (Asn), or 5 mM DMGlu. “NEAA-Asn”indicates addition of a reconstituted NEAA mixture lacking asparagine.Growth is shown relative to growth in medium containing 2 mM glutamine(Mean ± SEM of N = 3). (C)Changes in metabolite pools in Lu1205 cells or melanocytes after 6 h inglutamine-free medium(Gln-), relative to cells with 2 mM glutamine(Gln+). Lu1205 Gln− cultures were supplemented as shownwith 3 mM DMaK, 0.1 mM Asn, 0.1 mM aspartate (Asp), or(“Both”) DMaK and Asn. Source data for part (C) are shownin SupplementaryDataset 1.
Mentions: We tested the glutamine dependence of nine melanoma lines with differentoncogenic drivers (4 mutant BRAF, 4 mutant NRAS, 1 mutant p53). All required atleast 1 mM glutamine for maximal growth (Figure 2A) and there was no proliferation in the absence of glutamine(Figure S1A, S1B),while most cell lines could grow in the absence of glucose (Figure S1C). In contrast,growth of melanocytes was similar with or without glutamine (Figure S1D). Asmelanocytes were grown in a melanocyte-specific medium, we checked growth ofLu1205 melanoma cells in this medium and confirmed that growth was substantiallyinhibited by the absence of glutamine (Figure S1E).

Bottom Line: Glutamine dependence is a prominent feature of cancer metabolism, and here we show that melanoma cells, irrespective of their oncogenic background, depend on glutamine for growth.In the absence of glutamine, TCA cycle metabolites were liable to depletion through aminotransferase-mediated α-ketoglutarate-to-glutamate conversion and glutamate secretion.Melanocytes use more glutamine for protein synthesis rather than secreting it as glutamate and are less prone to loss of glutamate and TCA cycle metabolites when starved of glutamine.

View Article: PubMed Central - PubMed

Affiliation: Sanford-Burnham Medical Research Institute, La Jolla, California 92037, USA.

ABSTRACT
Glutamine dependence is a prominent feature of cancer metabolism, and here we show that melanoma cells, irrespective of their oncogenic background, depend on glutamine for growth. A quantitative audit of how carbon from glutamine is used showed that TCA-cycle-derived glutamate is, in most melanoma cells, the major glutamine-derived cataplerotic output and product of glutaminolysis. In the absence of glutamine, TCA cycle metabolites were liable to depletion through aminotransferase-mediated α-ketoglutarate-to-glutamate conversion and glutamate secretion. Aspartate was an essential cataplerotic output, as melanoma cells demonstrated a limited capacity to salvage external aspartate. Also, the absence of asparagine increased the glutamine requirement, pointing to vulnerability in the aspartate-asparagine biosynthetic pathway within melanoma metabolism. In contrast to melanoma cells, melanocytes could grow in the absence of glutamine. Melanocytes use more glutamine for protein synthesis rather than secreting it as glutamate and are less prone to loss of glutamate and TCA cycle metabolites when starved of glutamine.

No MeSH data available.


Related in: MedlinePlus