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Mislocalization of the exitatory amino-acid transporters (EAATs) in human astrocytoma and non-astrocytoma cancer cells: effect of the cell confluence.

Varini K, Benzaria A, Taïeb N, Di Scala C, Azmi A, Graoudi S, Maresca M - J. Biomed. Sci. (2012)

Bottom Line: Although non-astrocytoma cancers express EAATs, the localization of EAATs and the handling of L-glutamate in that case have not been investigated.Taken together, our results demonstrated that the mislocalization of the EAATs and its associated altered handling of glutamate are not restricted to astrocytomas but were also found in human non-astrocytoma cancers.Importantly, we found that a cell contact-dependent signal caused the relocalization of EAATs at the plasma membrane at least in human non-astrocytoma cancer cells, resulting in the correction of the altered transport of glutamate in such cancer cells but not in astrocytoma.

View Article: PubMed Central - HTML - PubMed

Affiliation: InteractionsCellulaires Neuroimmunes et Pathologies du Système Nerveux Central, CRN2M, CNRS UMR 6231, University of Aix-Marseille 2 and Aix-Marseille 3, Faculté de Médecine-Secteur Nord, Université de la Méditerranée, Marseille, France.

ABSTRACT

Background: Astrocytomas are cancers of the brain in which high levels of extracellular glutamate plays a critical role in tumor growth and resistance to conventional treatments. This is due for part to a decrease in the activity of the glutamate transporters, i.e. the Excitatory Amino Acid Transporters or EAATs, in relation to their nuclear mislocalization in astrocytoma cells. Although non-astrocytoma cancers express EAATs, the localization of EAATs and the handling of L-glutamate in that case have not been investigated.

Methods: We looked at the cellular localization and activity of EAATs in human astrocytoma and non-astrocytoma cancer cells by immunofluorescence, cell fractionation and L-glutamate transport studies.

Results: We demonstrated that the nuclear mislocalization of EAATs was not restricted to astrocytoma and happened in all sub-confluent non-astrocytoma cancer cells we tested. In addition, we found that cell-cell contact caused the relocalization of EAATs from the nuclei to the plasma membrane in all human cancer cells tested, except astrocytoma.

Conclusions: Taken together, our results demonstrated that the mislocalization of the EAATs and its associated altered handling of glutamate are not restricted to astrocytomas but were also found in human non-astrocytoma cancers. Importantly, we found that a cell contact-dependent signal caused the relocalization of EAATs at the plasma membrane at least in human non-astrocytoma cancer cells, resulting in the correction of the altered transport of glutamate in such cancer cells but not in astrocytoma.

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Cell fractionation of primary and malignant astrocytes. Sub-confluent primary astrocytes and STTG-1 cells were subjected to cell fractionation and western-blot analysis as explained in Materials and Methods. Nitrocellulose membranes were probed with antibodies directed against EAAT1/GLAST or EAAT2/GLT-1 or against antigens specific of each compartment (i.e. Hsp90, EGF-R and Histone 3 for the cytosolic, membrane and nuclear fractions, respectively). Cy = cytosolic fraction, Mb = membrane fraction, Nu = nuclear fraction.
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Figure 3: Cell fractionation of primary and malignant astrocytes. Sub-confluent primary astrocytes and STTG-1 cells were subjected to cell fractionation and western-blot analysis as explained in Materials and Methods. Nitrocellulose membranes were probed with antibodies directed against EAAT1/GLAST or EAAT2/GLT-1 or against antigens specific of each compartment (i.e. Hsp90, EGF-R and Histone 3 for the cytosolic, membrane and nuclear fractions, respectively). Cy = cytosolic fraction, Mb = membrane fraction, Nu = nuclear fraction.

Mentions: Cell fractionation and western analysis confirmed the IF observations (Figure 3). Results demonstrated that EAATs were mainly associated to the membrane fraction and the nuclear fraction in primary astrocytes and STTG-1 cells, respectively.


Mislocalization of the exitatory amino-acid transporters (EAATs) in human astrocytoma and non-astrocytoma cancer cells: effect of the cell confluence.

Varini K, Benzaria A, Taïeb N, Di Scala C, Azmi A, Graoudi S, Maresca M - J. Biomed. Sci. (2012)

Cell fractionation of primary and malignant astrocytes. Sub-confluent primary astrocytes and STTG-1 cells were subjected to cell fractionation and western-blot analysis as explained in Materials and Methods. Nitrocellulose membranes were probed with antibodies directed against EAAT1/GLAST or EAAT2/GLT-1 or against antigens specific of each compartment (i.e. Hsp90, EGF-R and Histone 3 for the cytosolic, membrane and nuclear fractions, respectively). Cy = cytosolic fraction, Mb = membrane fraction, Nu = nuclear fraction.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3293732&req=5

Figure 3: Cell fractionation of primary and malignant astrocytes. Sub-confluent primary astrocytes and STTG-1 cells were subjected to cell fractionation and western-blot analysis as explained in Materials and Methods. Nitrocellulose membranes were probed with antibodies directed against EAAT1/GLAST or EAAT2/GLT-1 or against antigens specific of each compartment (i.e. Hsp90, EGF-R and Histone 3 for the cytosolic, membrane and nuclear fractions, respectively). Cy = cytosolic fraction, Mb = membrane fraction, Nu = nuclear fraction.
Mentions: Cell fractionation and western analysis confirmed the IF observations (Figure 3). Results demonstrated that EAATs were mainly associated to the membrane fraction and the nuclear fraction in primary astrocytes and STTG-1 cells, respectively.

Bottom Line: Although non-astrocytoma cancers express EAATs, the localization of EAATs and the handling of L-glutamate in that case have not been investigated.Taken together, our results demonstrated that the mislocalization of the EAATs and its associated altered handling of glutamate are not restricted to astrocytomas but were also found in human non-astrocytoma cancers.Importantly, we found that a cell contact-dependent signal caused the relocalization of EAATs at the plasma membrane at least in human non-astrocytoma cancer cells, resulting in the correction of the altered transport of glutamate in such cancer cells but not in astrocytoma.

View Article: PubMed Central - HTML - PubMed

Affiliation: InteractionsCellulaires Neuroimmunes et Pathologies du Système Nerveux Central, CRN2M, CNRS UMR 6231, University of Aix-Marseille 2 and Aix-Marseille 3, Faculté de Médecine-Secteur Nord, Université de la Méditerranée, Marseille, France.

ABSTRACT

Background: Astrocytomas are cancers of the brain in which high levels of extracellular glutamate plays a critical role in tumor growth and resistance to conventional treatments. This is due for part to a decrease in the activity of the glutamate transporters, i.e. the Excitatory Amino Acid Transporters or EAATs, in relation to their nuclear mislocalization in astrocytoma cells. Although non-astrocytoma cancers express EAATs, the localization of EAATs and the handling of L-glutamate in that case have not been investigated.

Methods: We looked at the cellular localization and activity of EAATs in human astrocytoma and non-astrocytoma cancer cells by immunofluorescence, cell fractionation and L-glutamate transport studies.

Results: We demonstrated that the nuclear mislocalization of EAATs was not restricted to astrocytoma and happened in all sub-confluent non-astrocytoma cancer cells we tested. In addition, we found that cell-cell contact caused the relocalization of EAATs from the nuclei to the plasma membrane in all human cancer cells tested, except astrocytoma.

Conclusions: Taken together, our results demonstrated that the mislocalization of the EAATs and its associated altered handling of glutamate are not restricted to astrocytomas but were also found in human non-astrocytoma cancers. Importantly, we found that a cell contact-dependent signal caused the relocalization of EAATs at the plasma membrane at least in human non-astrocytoma cancer cells, resulting in the correction of the altered transport of glutamate in such cancer cells but not in astrocytoma.

Show MeSH
Related in: MedlinePlus