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Subcellular Partitioning of Protein Tyrosine Phosphatase 1B to the Endoplasmic Reticulum and Mitochondria Depends Sensitively on the Composition of Its Tail Anchor.

Fueller J, Egorov MV, Walther KA, Sabet O, Mallah J, Grabenbauer M, Kinkhabwala A - PLoS ONE (2015)

Bottom Line: Here, we show that PTP1B's mitochondrial localization is general (observed across diverse mammalian cell lines) and sensitively dependent on the transmembrane domain length, C-terminal charge and hydropathy of its short (≤35 amino acid) tail anchor.Our electron microscopy of specific DAB precipitation revealed that PTP1B localizes via its tail anchor to the outer mitochondrial membrane (OMM), with fluorescence lifetime imaging microscopy establishing that this OMM pool contributes to the previously reported cytoplasmic interaction of PTP1B with endocytosed epidermal growth factor receptor.Further functional elucidation of the newly recognized mitochondrial pool of PTP1B will likely be important for understanding its complex roles in cellular responses to external stimuli, cell proliferation and diseased states.

View Article: PubMed Central - PubMed

Affiliation: Department of Systemic Cell Biology, Max Planck Institute of Molecular Physiology, Otto-Hahn-Strasse 11, 44227, Dortmund, Germany; Center for Molecular Biology (ZMBH), DKFZ-ZMBH Alliance, Heidelberg University, Im Neuenheimer Feld 282, 69120, Heidelberg, Germany.

ABSTRACT
The canonical protein tyrosine phosphatase PTP1B is an important regulator of diverse cellular signaling networks. PTP1B has long been thought to exert its influence solely from its perch on the endoplasmic reticulum (ER); however, an additional subpopulation of PTP1B has recently been detected in mitochondria extracted from rat brain tissue. Here, we show that PTP1B's mitochondrial localization is general (observed across diverse mammalian cell lines) and sensitively dependent on the transmembrane domain length, C-terminal charge and hydropathy of its short (≤35 amino acid) tail anchor. Our electron microscopy of specific DAB precipitation revealed that PTP1B localizes via its tail anchor to the outer mitochondrial membrane (OMM), with fluorescence lifetime imaging microscopy establishing that this OMM pool contributes to the previously reported cytoplasmic interaction of PTP1B with endocytosed epidermal growth factor receptor. We additionally examined the mechanism of PTP1B's insertion into the ER membrane through heterologous expression of PTP1B's tail anchor in wild-type yeast and yeast mutants of major conserved ER insertion pathways: In none of these yeast strains was ER targeting significantly impeded, providing in vivo support for the hypothesis of spontaneous membrane insertion (as previously demonstrated in vitro). Further functional elucidation of the newly recognized mitochondrial pool of PTP1B will likely be important for understanding its complex roles in cellular responses to external stimuli, cell proliferation and diseased states.

No MeSH data available.


Related in: MedlinePlus

PTP1B localizes to the mitochondria in COS-7 cells.Confocal images of COS-7 cells expressing mCitrine-PTP1B along with the mitochondrial marker Tom20-mTagBFP. The mCitrine-PTP1B chimera localized to the general ER (arrowhead) and at a higher local concentration to the mitochondria (arrow). Scale bar: 20 μm.
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pone.0139429.g002: PTP1B localizes to the mitochondria in COS-7 cells.Confocal images of COS-7 cells expressing mCitrine-PTP1B along with the mitochondrial marker Tom20-mTagBFP. The mCitrine-PTP1B chimera localized to the general ER (arrowhead) and at a higher local concentration to the mitochondria (arrow). Scale bar: 20 μm.

Mentions: Upon its expression in COS-7 cells, a chimera of mCitrine with the second splice variant of PTP1B (ending in FLFNSNT and the central focus of our current study) exhibited the same strikingly high concentration to structures coincident with the mitochondria (Fig 2, arrow indicates an individual mitochondrion) as compared to its general distribution along the ER (Fig 2, arrowhead indicates a mitochondria-free region of the ER). Its higher local concentration to mitochondrial structures was quantitatively verified by plotting the mCitrine-PTP1B brightness distribution (Fig 2 histograms) in a peripheral region free from mitochondria (blue histogram corresponding to the solid box in the images) and in a peripheral region containing mitochondria (red histogram corresponding to the dashed box in the images). The brightness distribution in the mitochondria-free region truncates at roughly 100 counts/pixel, whereas the distribution in the mitochondria-containing region extends to roughly 250 counts/pixel.


Subcellular Partitioning of Protein Tyrosine Phosphatase 1B to the Endoplasmic Reticulum and Mitochondria Depends Sensitively on the Composition of Its Tail Anchor.

Fueller J, Egorov MV, Walther KA, Sabet O, Mallah J, Grabenbauer M, Kinkhabwala A - PLoS ONE (2015)

PTP1B localizes to the mitochondria in COS-7 cells.Confocal images of COS-7 cells expressing mCitrine-PTP1B along with the mitochondrial marker Tom20-mTagBFP. The mCitrine-PTP1B chimera localized to the general ER (arrowhead) and at a higher local concentration to the mitochondria (arrow). Scale bar: 20 μm.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4592070&req=5

pone.0139429.g002: PTP1B localizes to the mitochondria in COS-7 cells.Confocal images of COS-7 cells expressing mCitrine-PTP1B along with the mitochondrial marker Tom20-mTagBFP. The mCitrine-PTP1B chimera localized to the general ER (arrowhead) and at a higher local concentration to the mitochondria (arrow). Scale bar: 20 μm.
Mentions: Upon its expression in COS-7 cells, a chimera of mCitrine with the second splice variant of PTP1B (ending in FLFNSNT and the central focus of our current study) exhibited the same strikingly high concentration to structures coincident with the mitochondria (Fig 2, arrow indicates an individual mitochondrion) as compared to its general distribution along the ER (Fig 2, arrowhead indicates a mitochondria-free region of the ER). Its higher local concentration to mitochondrial structures was quantitatively verified by plotting the mCitrine-PTP1B brightness distribution (Fig 2 histograms) in a peripheral region free from mitochondria (blue histogram corresponding to the solid box in the images) and in a peripheral region containing mitochondria (red histogram corresponding to the dashed box in the images). The brightness distribution in the mitochondria-free region truncates at roughly 100 counts/pixel, whereas the distribution in the mitochondria-containing region extends to roughly 250 counts/pixel.

Bottom Line: Here, we show that PTP1B's mitochondrial localization is general (observed across diverse mammalian cell lines) and sensitively dependent on the transmembrane domain length, C-terminal charge and hydropathy of its short (≤35 amino acid) tail anchor.Our electron microscopy of specific DAB precipitation revealed that PTP1B localizes via its tail anchor to the outer mitochondrial membrane (OMM), with fluorescence lifetime imaging microscopy establishing that this OMM pool contributes to the previously reported cytoplasmic interaction of PTP1B with endocytosed epidermal growth factor receptor.Further functional elucidation of the newly recognized mitochondrial pool of PTP1B will likely be important for understanding its complex roles in cellular responses to external stimuli, cell proliferation and diseased states.

View Article: PubMed Central - PubMed

Affiliation: Department of Systemic Cell Biology, Max Planck Institute of Molecular Physiology, Otto-Hahn-Strasse 11, 44227, Dortmund, Germany; Center for Molecular Biology (ZMBH), DKFZ-ZMBH Alliance, Heidelberg University, Im Neuenheimer Feld 282, 69120, Heidelberg, Germany.

ABSTRACT
The canonical protein tyrosine phosphatase PTP1B is an important regulator of diverse cellular signaling networks. PTP1B has long been thought to exert its influence solely from its perch on the endoplasmic reticulum (ER); however, an additional subpopulation of PTP1B has recently been detected in mitochondria extracted from rat brain tissue. Here, we show that PTP1B's mitochondrial localization is general (observed across diverse mammalian cell lines) and sensitively dependent on the transmembrane domain length, C-terminal charge and hydropathy of its short (≤35 amino acid) tail anchor. Our electron microscopy of specific DAB precipitation revealed that PTP1B localizes via its tail anchor to the outer mitochondrial membrane (OMM), with fluorescence lifetime imaging microscopy establishing that this OMM pool contributes to the previously reported cytoplasmic interaction of PTP1B with endocytosed epidermal growth factor receptor. We additionally examined the mechanism of PTP1B's insertion into the ER membrane through heterologous expression of PTP1B's tail anchor in wild-type yeast and yeast mutants of major conserved ER insertion pathways: In none of these yeast strains was ER targeting significantly impeded, providing in vivo support for the hypothesis of spontaneous membrane insertion (as previously demonstrated in vitro). Further functional elucidation of the newly recognized mitochondrial pool of PTP1B will likely be important for understanding its complex roles in cellular responses to external stimuli, cell proliferation and diseased states.

No MeSH data available.


Related in: MedlinePlus