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The mitochondrial carrier SLC25A10 regulates cancer cell growth.

Zhou X, Paredes JA, Krishnan S, Curbo S, Karlsson A - Oncotarget (2015)

Bottom Line: The metabolic alteration was linked to an energy metabolic shift from glycolysis to mitochondrial oxidative phosphorylation illustrated by increased expression of glutamate dehydrogenase, decreased expression of lactate dehydrogenase due to down-regulation of hypoxia inducible factor 1α.We identified effects on NADPH production linked to the growth changes observed in SLC25A10 knockdown cells, demonstrated by decreased NADPH production in cells deprived of glutamine.The contribution of SLC25A10 to reprogram cell metabolism and to regulate cell growth suggests SLC25A10 as a novel target for anti-cancer strategies.

View Article: PubMed Central - PubMed

Affiliation: Division of Clinical Microbiology, Department of Laboratory Medicine, Karolinska Institute, Karolinska University Hospital Huddinge, 141 86 Stockholm, Sweden.

ABSTRACT
Dysregulation of cell metabolism is critical for the growth properties of cancer cells. The purpose of this study was to understand the role of substrate transport across the mitochondrial membrane to sustain the metabolic shift and redox defense in cancer cells. Mitochondrial carrier SLC25A10 is up-regulated in a variety of tumors and is involved in regulating intracellular levels of reactive oxygen species. We show that knockdown of SLC25A10 in A549 cells changed the growth properties to a less malignant phenotype and casued increased glutamine dependency and sensitivity to oxidative stress. The metabolic alteration was linked to an energy metabolic shift from glycolysis to mitochondrial oxidative phosphorylation illustrated by increased expression of glutamate dehydrogenase, decreased expression of lactate dehydrogenase due to down-regulation of hypoxia inducible factor 1α. We identified effects on NADPH production linked to the growth changes observed in SLC25A10 knockdown cells, demonstrated by decreased NADPH production in cells deprived of glutamine. The contribution of SLC25A10 to reprogram cell metabolism and to regulate cell growth suggests SLC25A10 as a novel target for anti-cancer strategies.

No MeSH data available.


Related in: MedlinePlus

Growth behavior and morphology changes of SLC25A10 knockdown cellsEstablishment of stable SLC25A10 knockdown cell lines with specific siRNA expression in A549 cell line. (A) SLC25A10 gene expression was analysed by real-time qPCR in different clones (siRNA-SLC-2, siRNA-SLC-4 and siRNA-SLC-5). (B) SLC25A10 protein expression in A549 cells; 1, untransfected cells; 2, siRNA-CON cells; 3, siRNA-SLC-2; 4, siRNA-SLC-5. (C) Growth curve of siRNA-SLC-2 cell line, data is represented as mean ± SD. (D) Cell morphology change (magnification 20X) and mitochondrial staining. Cells with altered mitochondria are indicated with arrows and scale bar represents 20 μm (E) Sizes of colonies. (F) Numbers of colonies. This experiment was repeated 3 times independently, data is presented as mean ± SD, *** represents p < 0, 001. (G) Mitochondrial DRP1 and MFN1 gene expression levels.
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Figure 1: Growth behavior and morphology changes of SLC25A10 knockdown cellsEstablishment of stable SLC25A10 knockdown cell lines with specific siRNA expression in A549 cell line. (A) SLC25A10 gene expression was analysed by real-time qPCR in different clones (siRNA-SLC-2, siRNA-SLC-4 and siRNA-SLC-5). (B) SLC25A10 protein expression in A549 cells; 1, untransfected cells; 2, siRNA-CON cells; 3, siRNA-SLC-2; 4, siRNA-SLC-5. (C) Growth curve of siRNA-SLC-2 cell line, data is represented as mean ± SD. (D) Cell morphology change (magnification 20X) and mitochondrial staining. Cells with altered mitochondria are indicated with arrows and scale bar represents 20 μm (E) Sizes of colonies. (F) Numbers of colonies. This experiment was repeated 3 times independently, data is presented as mean ± SD, *** represents p < 0, 001. (G) Mitochondrial DRP1 and MFN1 gene expression levels.

Mentions: Stable knockdown A549 NSCLC cell lines (siRNA-SLC −2, −4 and −5) with more than 75% reduction of SLC25A10 mRNA were established (Figure 1A). The SLC25A10 protein levels decreased by 73%, 80% and 37% in siRNA-SLC −2, −4 and −5 compared to the siRNA-CON and untransfected cells (Figure 1B). The down-regulation of SLC25A10 did not affect the doubling time of both cell types, however, after reaching confluency the siRNA-CON cells had a higher proliferation rate than the siRNA-SLC cells (Figure 1C). The siRNA-SLC cells grew in a monolayer manner and displayed decreased ability to form cell islands compared to untransfected A549 or siRNA-CON cells (Figure 1D). Moreover, the size of the siRNA-SLC cells was smaller than the size of the siRNA-CON cells (Figure 1D). Since siRNA-SLC cells grew in an even monolayer, soft agar experiments were performed to compare the ability of anchorage-independent growth of SLC25A10 knockdown cells with untransfected and mock control cells. The sizes of colonies from siRNA-SLC were small compared to the colonies formed by untransfected or siRNA-CON cells (Figure 1E) and in addition the number of colonies formed by the siRNA-SLC cells were significantly lower than in untransfected or siRNA-CON cells (Figure 1F).


The mitochondrial carrier SLC25A10 regulates cancer cell growth.

Zhou X, Paredes JA, Krishnan S, Curbo S, Karlsson A - Oncotarget (2015)

Growth behavior and morphology changes of SLC25A10 knockdown cellsEstablishment of stable SLC25A10 knockdown cell lines with specific siRNA expression in A549 cell line. (A) SLC25A10 gene expression was analysed by real-time qPCR in different clones (siRNA-SLC-2, siRNA-SLC-4 and siRNA-SLC-5). (B) SLC25A10 protein expression in A549 cells; 1, untransfected cells; 2, siRNA-CON cells; 3, siRNA-SLC-2; 4, siRNA-SLC-5. (C) Growth curve of siRNA-SLC-2 cell line, data is represented as mean ± SD. (D) Cell morphology change (magnification 20X) and mitochondrial staining. Cells with altered mitochondria are indicated with arrows and scale bar represents 20 μm (E) Sizes of colonies. (F) Numbers of colonies. This experiment was repeated 3 times independently, data is presented as mean ± SD, *** represents p < 0, 001. (G) Mitochondrial DRP1 and MFN1 gene expression levels.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4496216&req=5

Figure 1: Growth behavior and morphology changes of SLC25A10 knockdown cellsEstablishment of stable SLC25A10 knockdown cell lines with specific siRNA expression in A549 cell line. (A) SLC25A10 gene expression was analysed by real-time qPCR in different clones (siRNA-SLC-2, siRNA-SLC-4 and siRNA-SLC-5). (B) SLC25A10 protein expression in A549 cells; 1, untransfected cells; 2, siRNA-CON cells; 3, siRNA-SLC-2; 4, siRNA-SLC-5. (C) Growth curve of siRNA-SLC-2 cell line, data is represented as mean ± SD. (D) Cell morphology change (magnification 20X) and mitochondrial staining. Cells with altered mitochondria are indicated with arrows and scale bar represents 20 μm (E) Sizes of colonies. (F) Numbers of colonies. This experiment was repeated 3 times independently, data is presented as mean ± SD, *** represents p < 0, 001. (G) Mitochondrial DRP1 and MFN1 gene expression levels.
Mentions: Stable knockdown A549 NSCLC cell lines (siRNA-SLC −2, −4 and −5) with more than 75% reduction of SLC25A10 mRNA were established (Figure 1A). The SLC25A10 protein levels decreased by 73%, 80% and 37% in siRNA-SLC −2, −4 and −5 compared to the siRNA-CON and untransfected cells (Figure 1B). The down-regulation of SLC25A10 did not affect the doubling time of both cell types, however, after reaching confluency the siRNA-CON cells had a higher proliferation rate than the siRNA-SLC cells (Figure 1C). The siRNA-SLC cells grew in a monolayer manner and displayed decreased ability to form cell islands compared to untransfected A549 or siRNA-CON cells (Figure 1D). Moreover, the size of the siRNA-SLC cells was smaller than the size of the siRNA-CON cells (Figure 1D). Since siRNA-SLC cells grew in an even monolayer, soft agar experiments were performed to compare the ability of anchorage-independent growth of SLC25A10 knockdown cells with untransfected and mock control cells. The sizes of colonies from siRNA-SLC were small compared to the colonies formed by untransfected or siRNA-CON cells (Figure 1E) and in addition the number of colonies formed by the siRNA-SLC cells were significantly lower than in untransfected or siRNA-CON cells (Figure 1F).

Bottom Line: The metabolic alteration was linked to an energy metabolic shift from glycolysis to mitochondrial oxidative phosphorylation illustrated by increased expression of glutamate dehydrogenase, decreased expression of lactate dehydrogenase due to down-regulation of hypoxia inducible factor 1α.We identified effects on NADPH production linked to the growth changes observed in SLC25A10 knockdown cells, demonstrated by decreased NADPH production in cells deprived of glutamine.The contribution of SLC25A10 to reprogram cell metabolism and to regulate cell growth suggests SLC25A10 as a novel target for anti-cancer strategies.

View Article: PubMed Central - PubMed

Affiliation: Division of Clinical Microbiology, Department of Laboratory Medicine, Karolinska Institute, Karolinska University Hospital Huddinge, 141 86 Stockholm, Sweden.

ABSTRACT
Dysregulation of cell metabolism is critical for the growth properties of cancer cells. The purpose of this study was to understand the role of substrate transport across the mitochondrial membrane to sustain the metabolic shift and redox defense in cancer cells. Mitochondrial carrier SLC25A10 is up-regulated in a variety of tumors and is involved in regulating intracellular levels of reactive oxygen species. We show that knockdown of SLC25A10 in A549 cells changed the growth properties to a less malignant phenotype and casued increased glutamine dependency and sensitivity to oxidative stress. The metabolic alteration was linked to an energy metabolic shift from glycolysis to mitochondrial oxidative phosphorylation illustrated by increased expression of glutamate dehydrogenase, decreased expression of lactate dehydrogenase due to down-regulation of hypoxia inducible factor 1α. We identified effects on NADPH production linked to the growth changes observed in SLC25A10 knockdown cells, demonstrated by decreased NADPH production in cells deprived of glutamine. The contribution of SLC25A10 to reprogram cell metabolism and to regulate cell growth suggests SLC25A10 as a novel target for anti-cancer strategies.

No MeSH data available.


Related in: MedlinePlus