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Mitochondrial p32 is upregulated in Myc expressing brain cancers and mediates glutamine addiction.

Fogal V, Babic I, Chao Y, Pastorino S, Mukthavaram R, Jiang P, Cho YJ, Pingle SC, Crawford JR, Piccioni DE, Kesari S - Oncotarget (2015)

Bottom Line: Loss of p32 in glutamine addicted glioma cells induced resistance to glutamine deprivation and imparted sensitivity to glucose withdrawal.Finally, we provide evidence that p32 expression contributes to Myc-induced glutamine addiction of cancer cells.Our findings suggest that Myc promotes the expression of p32, which is required to maintain sufficient respiratory capacity to sustain glutamine metabolism in Myc transformed cells.

View Article: PubMed Central - PubMed

Affiliation: Translational Neuro-Oncology Laboratories, Moores Cancer Center, University of California San Diego, La Jolla, CA.

ABSTRACT
Metabolic reprogramming is a key feature of tumorigenesis that is controlled by oncogenes. Enhanced utilization of glucose and glutamine are the best-established hallmarks of tumor metabolism. The oncogene c-Myc is one of the major players responsible for this metabolic alteration. However, the molecular mechanisms involved in Myc-induced metabolic reprogramming are not well defined. Here we identify p32, a mitochondrial protein known to play a role in the expression of mitochondrial respiratory chain complexes, as a critical player in Myc-induced glutamine addiction. We show that p32 is a direct transcriptional target of Myc and that high level of Myc in malignant brain cancers correlates with high expression of p32. Attenuation of p32 expression reduced growth rate of glioma cells expressing Myc and impaired tumor formation in vivo. Loss of p32 in glutamine addicted glioma cells induced resistance to glutamine deprivation and imparted sensitivity to glucose withdrawal. Finally, we provide evidence that p32 expression contributes to Myc-induced glutamine addiction of cancer cells. Our findings suggest that Myc promotes the expression of p32, which is required to maintain sufficient respiratory capacity to sustain glutamine metabolism in Myc transformed cells.

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Correlation between p32 and Myc expression in human gliomas and glioma cell lines(A) Correlation between Myc and p32 expression in medulloblastoma human tumors. Left panel. Heat map of p32, c-Myc and N-Myc for each medulloblastoma subgroup (c1 through c6) and an additional ATRT subgroup (atypical teratoid/rhabdoid tumors) [23], C5/c1 Medulloblastoma subgroup, characterized by a Myc activation signature [23], exhibits high expression of p32. WNT, Wingless signaling pathway, SHH Sonic Hedgehog signaling pathway. nl cbl, Normal cerebellum samples. (A)-right panel and (B): correlation between p32 and Myc expression in a medulloblastoma array ((A)-right panel) and a mixed glioma (B) as indicated by the % of p32 and Myc positive staining for each core of the arrays. Sequential slides of each array were stained separately with polyclonal anti p32 and c-Myc antibodies. The % of p32 and Myc positive staining for each core was quantified using Aperio software. The Pearson correlation coefficient (r) of linear regression was calculated using data sets deprived of samples expressing low Myc (<15% of staining) but moderate-high p32 (>15% of staining). The immunohistochemistry images (200x magnification) show representative glioma cores (red arrows in graph) exhibiting correlation between Myc and p32 expression. (C) qPCR analysis of p32 and Myc expression in established glioma cell lines (red) and patient derived glioma stem cells (blue) as compared to normal astrocytes (black). The bars shown are normalized to an internal β-actin control and represent the mean ± SEM of at least three independent experiments.
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Figure 2: Correlation between p32 and Myc expression in human gliomas and glioma cell lines(A) Correlation between Myc and p32 expression in medulloblastoma human tumors. Left panel. Heat map of p32, c-Myc and N-Myc for each medulloblastoma subgroup (c1 through c6) and an additional ATRT subgroup (atypical teratoid/rhabdoid tumors) [23], C5/c1 Medulloblastoma subgroup, characterized by a Myc activation signature [23], exhibits high expression of p32. WNT, Wingless signaling pathway, SHH Sonic Hedgehog signaling pathway. nl cbl, Normal cerebellum samples. (A)-right panel and (B): correlation between p32 and Myc expression in a medulloblastoma array ((A)-right panel) and a mixed glioma (B) as indicated by the % of p32 and Myc positive staining for each core of the arrays. Sequential slides of each array were stained separately with polyclonal anti p32 and c-Myc antibodies. The % of p32 and Myc positive staining for each core was quantified using Aperio software. The Pearson correlation coefficient (r) of linear regression was calculated using data sets deprived of samples expressing low Myc (<15% of staining) but moderate-high p32 (>15% of staining). The immunohistochemistry images (200x magnification) show representative glioma cores (red arrows in graph) exhibiting correlation between Myc and p32 expression. (C) qPCR analysis of p32 and Myc expression in established glioma cell lines (red) and patient derived glioma stem cells (blue) as compared to normal astrocytes (black). The bars shown are normalized to an internal β-actin control and represent the mean ± SEM of at least three independent experiments.

Mentions: Myc is central to the genesis of most human cancers, and deregulated Myc is closely correlated with the grade of brain tumor malignancy [21–23, 47]. Microarray analysis of Myc-responsive genes identified p32 as a potential transcriptional target of Myc [43, 44, 46]; as such, we investigated a possible correlation between Myc and p32 expression in malignant brain tumors. We first focused on medulloblastoma. These highly heterogeneous malignant brain tumors, usually found only in children, have been classified into six molecular subgroups, each with a unique combination of chromosomal aberrations [23]. One molecular subgroup, with a particularly aggressive course, is characterized genetically by MYC copy number gains and transcriptionally by enrichment of photoreceptor pathways. Unsupervised clustering of mRNA expression data from 194 medulloblastoma revealed concomitant high expression of p32 and Myc in medulloblastoma with poor clinical outcome (Fig. 2A left panel-c5/c1 subgroup). Correlation of p32 and Myc expression in medulloblastoma tissues was also evident following immunostaining of a medulloblastoma tissue array (Fig. 2A right panel). Similar immunohistochemical analysis was also performed in an array containing glioma subtypes (Fig. 2B). In this case the correlation had a lower Pearson coefficient (r = 0.49) because some tissue cores express low or undetectable levels of Myc, but moderate to high levels of p32 (Supplementary Fig. S1, samples in red box). This is not surprising, since p32 expression is also likely to be regulated by Myc-independent mechanisms. A linear regression analysis excluding these tissues revealed a strong correlation between Myc and p32 expression (r = 0.76) (Fig. 2B). In addition, quantitative RT-PCR analysis showed an upregulation of p32 in glioma cell lines (Fig. 2C red bars) as well as patient-derived glioma stem cells (Fig. 2C, blue bars) compared to normal astrocytes. In agreement with results from the tissue arrays, there was a strong correlation between up-regulation of Myc and p32 in over half of the cell lines tested.


Mitochondrial p32 is upregulated in Myc expressing brain cancers and mediates glutamine addiction.

Fogal V, Babic I, Chao Y, Pastorino S, Mukthavaram R, Jiang P, Cho YJ, Pingle SC, Crawford JR, Piccioni DE, Kesari S - Oncotarget (2015)

Correlation between p32 and Myc expression in human gliomas and glioma cell lines(A) Correlation between Myc and p32 expression in medulloblastoma human tumors. Left panel. Heat map of p32, c-Myc and N-Myc for each medulloblastoma subgroup (c1 through c6) and an additional ATRT subgroup (atypical teratoid/rhabdoid tumors) [23], C5/c1 Medulloblastoma subgroup, characterized by a Myc activation signature [23], exhibits high expression of p32. WNT, Wingless signaling pathway, SHH Sonic Hedgehog signaling pathway. nl cbl, Normal cerebellum samples. (A)-right panel and (B): correlation between p32 and Myc expression in a medulloblastoma array ((A)-right panel) and a mixed glioma (B) as indicated by the % of p32 and Myc positive staining for each core of the arrays. Sequential slides of each array were stained separately with polyclonal anti p32 and c-Myc antibodies. The % of p32 and Myc positive staining for each core was quantified using Aperio software. The Pearson correlation coefficient (r) of linear regression was calculated using data sets deprived of samples expressing low Myc (<15% of staining) but moderate-high p32 (>15% of staining). The immunohistochemistry images (200x magnification) show representative glioma cores (red arrows in graph) exhibiting correlation between Myc and p32 expression. (C) qPCR analysis of p32 and Myc expression in established glioma cell lines (red) and patient derived glioma stem cells (blue) as compared to normal astrocytes (black). The bars shown are normalized to an internal β-actin control and represent the mean ± SEM of at least three independent experiments.
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Figure 2: Correlation between p32 and Myc expression in human gliomas and glioma cell lines(A) Correlation between Myc and p32 expression in medulloblastoma human tumors. Left panel. Heat map of p32, c-Myc and N-Myc for each medulloblastoma subgroup (c1 through c6) and an additional ATRT subgroup (atypical teratoid/rhabdoid tumors) [23], C5/c1 Medulloblastoma subgroup, characterized by a Myc activation signature [23], exhibits high expression of p32. WNT, Wingless signaling pathway, SHH Sonic Hedgehog signaling pathway. nl cbl, Normal cerebellum samples. (A)-right panel and (B): correlation between p32 and Myc expression in a medulloblastoma array ((A)-right panel) and a mixed glioma (B) as indicated by the % of p32 and Myc positive staining for each core of the arrays. Sequential slides of each array were stained separately with polyclonal anti p32 and c-Myc antibodies. The % of p32 and Myc positive staining for each core was quantified using Aperio software. The Pearson correlation coefficient (r) of linear regression was calculated using data sets deprived of samples expressing low Myc (<15% of staining) but moderate-high p32 (>15% of staining). The immunohistochemistry images (200x magnification) show representative glioma cores (red arrows in graph) exhibiting correlation between Myc and p32 expression. (C) qPCR analysis of p32 and Myc expression in established glioma cell lines (red) and patient derived glioma stem cells (blue) as compared to normal astrocytes (black). The bars shown are normalized to an internal β-actin control and represent the mean ± SEM of at least three independent experiments.
Mentions: Myc is central to the genesis of most human cancers, and deregulated Myc is closely correlated with the grade of brain tumor malignancy [21–23, 47]. Microarray analysis of Myc-responsive genes identified p32 as a potential transcriptional target of Myc [43, 44, 46]; as such, we investigated a possible correlation between Myc and p32 expression in malignant brain tumors. We first focused on medulloblastoma. These highly heterogeneous malignant brain tumors, usually found only in children, have been classified into six molecular subgroups, each with a unique combination of chromosomal aberrations [23]. One molecular subgroup, with a particularly aggressive course, is characterized genetically by MYC copy number gains and transcriptionally by enrichment of photoreceptor pathways. Unsupervised clustering of mRNA expression data from 194 medulloblastoma revealed concomitant high expression of p32 and Myc in medulloblastoma with poor clinical outcome (Fig. 2A left panel-c5/c1 subgroup). Correlation of p32 and Myc expression in medulloblastoma tissues was also evident following immunostaining of a medulloblastoma tissue array (Fig. 2A right panel). Similar immunohistochemical analysis was also performed in an array containing glioma subtypes (Fig. 2B). In this case the correlation had a lower Pearson coefficient (r = 0.49) because some tissue cores express low or undetectable levels of Myc, but moderate to high levels of p32 (Supplementary Fig. S1, samples in red box). This is not surprising, since p32 expression is also likely to be regulated by Myc-independent mechanisms. A linear regression analysis excluding these tissues revealed a strong correlation between Myc and p32 expression (r = 0.76) (Fig. 2B). In addition, quantitative RT-PCR analysis showed an upregulation of p32 in glioma cell lines (Fig. 2C red bars) as well as patient-derived glioma stem cells (Fig. 2C, blue bars) compared to normal astrocytes. In agreement with results from the tissue arrays, there was a strong correlation between up-regulation of Myc and p32 in over half of the cell lines tested.

Bottom Line: Loss of p32 in glutamine addicted glioma cells induced resistance to glutamine deprivation and imparted sensitivity to glucose withdrawal.Finally, we provide evidence that p32 expression contributes to Myc-induced glutamine addiction of cancer cells.Our findings suggest that Myc promotes the expression of p32, which is required to maintain sufficient respiratory capacity to sustain glutamine metabolism in Myc transformed cells.

View Article: PubMed Central - PubMed

Affiliation: Translational Neuro-Oncology Laboratories, Moores Cancer Center, University of California San Diego, La Jolla, CA.

ABSTRACT
Metabolic reprogramming is a key feature of tumorigenesis that is controlled by oncogenes. Enhanced utilization of glucose and glutamine are the best-established hallmarks of tumor metabolism. The oncogene c-Myc is one of the major players responsible for this metabolic alteration. However, the molecular mechanisms involved in Myc-induced metabolic reprogramming are not well defined. Here we identify p32, a mitochondrial protein known to play a role in the expression of mitochondrial respiratory chain complexes, as a critical player in Myc-induced glutamine addiction. We show that p32 is a direct transcriptional target of Myc and that high level of Myc in malignant brain cancers correlates with high expression of p32. Attenuation of p32 expression reduced growth rate of glioma cells expressing Myc and impaired tumor formation in vivo. Loss of p32 in glutamine addicted glioma cells induced resistance to glutamine deprivation and imparted sensitivity to glucose withdrawal. Finally, we provide evidence that p32 expression contributes to Myc-induced glutamine addiction of cancer cells. Our findings suggest that Myc promotes the expression of p32, which is required to maintain sufficient respiratory capacity to sustain glutamine metabolism in Myc transformed cells.

Show MeSH
Related in: MedlinePlus