Spatial and functional relationship between poly(ADP-ribose) polymerase-1 and poly(ADP-ribose) glycohydrolase in the brain

M. F. Poitras; D. W. Koh; S. W. Yu; S. A. Andrabi; A. S. Mandir; G. G. Poirier; V. L. Dawson; T. M. Dawson

doi:10.1016/j.neuroscience.2007.04.062

Spatial and functional relationship between poly(ADP-ribose) polymerase-1 and poly(ADP-ribose) glycohydrolase in the brain

M. F. Poitras, D. W. Koh, S. W. Yu, S. A. Andrabi, A. S. Mandir, G. G. Poirier, V. L. Dawson, T. M. Dawson

School of Medicine

Research output: Contribution to journal › Article › peer-review

34 Scopus citations

Abstract

Poly(ADP-ribose) polymerases (PARPs) are members of a family of enzymes that utilize nicotinamide adenine dinucleotide (NAD⁺) as substrate to form large ADP-ribose polymers (PAR) in the nucleus. PAR has a very short half-life due to its rapid degradation by poly(ADP-ribose) glycohydrolase (PARG). PARP-1 mediates acute neuronal cell death induced by a variety of insults including cerebral ischemia, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced Parkinsonism, and CNS trauma. While PARP-1 is localized to the nucleus, PARG resides in both the nucleus and cytoplasm. Surprisingly, there appears to be only one gene encoding PARG activity, which has been characterized in vitro to generate different splice variants, in contrast to the growing family of PARPs. Little is known regarding the spatial and functional relationships of PARG and PARP-1. Here we evaluate PARG expression in the brain and its cellular and subcellular distribution in relation to PARP-1. Anti-PARG (α-PARG) antibodies raised in rabbits using a purified 30 kDa C-terminal fragment of murine PARG recognize a single band at 111 kDa in the brain. Western blot analysis also shows that PARG and PARP-1 are evenly distributed throughout the brain. Immunohistochemical studies using α-PARG antibodies reveal punctate cytosolic staining, whereas anti-PARP-1 (α-PARP-1) antibodies demonstrate nuclear staining. PARG is enriched in the mitochondrial fraction together with manganese superoxide dismutase (MnSOD) and cytochrome C (Cyt C) following whole brain subcellular fractionation and Western blot analysis. Confocal microscopy confirms the co-localization of PARG and Cyt C. Finally, PARG translocation to the nucleus is triggered by NMDA-induced PARP-1 activation. Therefore, the subcellular segregation of PARG in the mitochondria and PARP-1 in the nucleus suggests that PARG translocation is necessary for their functional interaction. This translocation is PARP-1 dependent, further demonstrating a functional interaction of PARP-1 and PARG in the brain.

Original language	English (US)
Pages (from-to)	198-211
Number of pages	14
Journal	Neuroscience
Volume	148
Issue number	1
DOIs	https://doi.org/10.1016/j.neuroscience.2007.04.062
State	Published - Aug 10 2007

Keywords

NMDA
immunostaining
mitochondria
nucleus
subcellular fractionation

ASJC Scopus subject areas

General Neuroscience

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10.1016/j.neuroscience.2007.04.062

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@article{2fe6c6773952434e8f20ddcda97b36e9,

title = "Spatial and functional relationship between poly(ADP-ribose) polymerase-1 and poly(ADP-ribose) glycohydrolase in the brain",

abstract = "Poly(ADP-ribose) polymerases (PARPs) are members of a family of enzymes that utilize nicotinamide adenine dinucleotide (NAD+) as substrate to form large ADP-ribose polymers (PAR) in the nucleus. PAR has a very short half-life due to its rapid degradation by poly(ADP-ribose) glycohydrolase (PARG). PARP-1 mediates acute neuronal cell death induced by a variety of insults including cerebral ischemia, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced Parkinsonism, and CNS trauma. While PARP-1 is localized to the nucleus, PARG resides in both the nucleus and cytoplasm. Surprisingly, there appears to be only one gene encoding PARG activity, which has been characterized in vitro to generate different splice variants, in contrast to the growing family of PARPs. Little is known regarding the spatial and functional relationships of PARG and PARP-1. Here we evaluate PARG expression in the brain and its cellular and subcellular distribution in relation to PARP-1. Anti-PARG (α-PARG) antibodies raised in rabbits using a purified 30 kDa C-terminal fragment of murine PARG recognize a single band at 111 kDa in the brain. Western blot analysis also shows that PARG and PARP-1 are evenly distributed throughout the brain. Immunohistochemical studies using α-PARG antibodies reveal punctate cytosolic staining, whereas anti-PARP-1 (α-PARP-1) antibodies demonstrate nuclear staining. PARG is enriched in the mitochondrial fraction together with manganese superoxide dismutase (MnSOD) and cytochrome C (Cyt C) following whole brain subcellular fractionation and Western blot analysis. Confocal microscopy confirms the co-localization of PARG and Cyt C. Finally, PARG translocation to the nucleus is triggered by NMDA-induced PARP-1 activation. Therefore, the subcellular segregation of PARG in the mitochondria and PARP-1 in the nucleus suggests that PARG translocation is necessary for their functional interaction. This translocation is PARP-1 dependent, further demonstrating a functional interaction of PARP-1 and PARG in the brain.",

keywords = "NMDA, immunostaining, mitochondria, nucleus, subcellular fractionation",

author = "Poitras, {M. F.} and Koh, {D. W.} and Yu, {S. W.} and Andrabi, {S. A.} and Mandir, {A. S.} and Poirier, {G. G.} and Dawson, {V. L.} and Dawson, {T. M.}",

note = "Funding Information: We thank Joke Wortel for technical support and Weza Cotman for secretarial support. Supported by grants from the NIH (NS39148), the American Heart Association, and the Mary Lou McIlhaney Scholar Award. M.F.P was supported by NSERC of Canada. G.G.P. is supported by the Medical Research Council of Canada. Under an agreement between The Johns Hopkins University and MGI, T.M.D. and V.L.D. are entitled to a share of sales royalty received by the university from MGI. The terms of this arrangement are being managed by the university in accordance with its conflict-of-interest policies. T.M.D. is the Leonard and Madlyn Abramson Professor of Neurodegenerative Diseases.",

year = "2007",

month = aug,

day = "10",

doi = "10.1016/j.neuroscience.2007.04.062",

language = "English (US)",

volume = "148",

pages = "198--211",

journal = "Neuroscience",

issn = "0306-4522",

publisher = "Elsevier Limited",

number = "1",

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TY - JOUR

T1 - Spatial and functional relationship between poly(ADP-ribose) polymerase-1 and poly(ADP-ribose) glycohydrolase in the brain

AU - Poitras, M. F.

AU - Koh, D. W.

AU - Yu, S. W.

AU - Andrabi, S. A.

AU - Mandir, A. S.

AU - Poirier, G. G.

AU - Dawson, V. L.

AU - Dawson, T. M.

N1 - Funding Information: We thank Joke Wortel for technical support and Weza Cotman for secretarial support. Supported by grants from the NIH (NS39148), the American Heart Association, and the Mary Lou McIlhaney Scholar Award. M.F.P was supported by NSERC of Canada. G.G.P. is supported by the Medical Research Council of Canada. Under an agreement between The Johns Hopkins University and MGI, T.M.D. and V.L.D. are entitled to a share of sales royalty received by the university from MGI. The terms of this arrangement are being managed by the university in accordance with its conflict-of-interest policies. T.M.D. is the Leonard and Madlyn Abramson Professor of Neurodegenerative Diseases.

PY - 2007/8/10

Y1 - 2007/8/10

N2 - Poly(ADP-ribose) polymerases (PARPs) are members of a family of enzymes that utilize nicotinamide adenine dinucleotide (NAD+) as substrate to form large ADP-ribose polymers (PAR) in the nucleus. PAR has a very short half-life due to its rapid degradation by poly(ADP-ribose) glycohydrolase (PARG). PARP-1 mediates acute neuronal cell death induced by a variety of insults including cerebral ischemia, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced Parkinsonism, and CNS trauma. While PARP-1 is localized to the nucleus, PARG resides in both the nucleus and cytoplasm. Surprisingly, there appears to be only one gene encoding PARG activity, which has been characterized in vitro to generate different splice variants, in contrast to the growing family of PARPs. Little is known regarding the spatial and functional relationships of PARG and PARP-1. Here we evaluate PARG expression in the brain and its cellular and subcellular distribution in relation to PARP-1. Anti-PARG (α-PARG) antibodies raised in rabbits using a purified 30 kDa C-terminal fragment of murine PARG recognize a single band at 111 kDa in the brain. Western blot analysis also shows that PARG and PARP-1 are evenly distributed throughout the brain. Immunohistochemical studies using α-PARG antibodies reveal punctate cytosolic staining, whereas anti-PARP-1 (α-PARP-1) antibodies demonstrate nuclear staining. PARG is enriched in the mitochondrial fraction together with manganese superoxide dismutase (MnSOD) and cytochrome C (Cyt C) following whole brain subcellular fractionation and Western blot analysis. Confocal microscopy confirms the co-localization of PARG and Cyt C. Finally, PARG translocation to the nucleus is triggered by NMDA-induced PARP-1 activation. Therefore, the subcellular segregation of PARG in the mitochondria and PARP-1 in the nucleus suggests that PARG translocation is necessary for their functional interaction. This translocation is PARP-1 dependent, further demonstrating a functional interaction of PARP-1 and PARG in the brain.

AB - Poly(ADP-ribose) polymerases (PARPs) are members of a family of enzymes that utilize nicotinamide adenine dinucleotide (NAD+) as substrate to form large ADP-ribose polymers (PAR) in the nucleus. PAR has a very short half-life due to its rapid degradation by poly(ADP-ribose) glycohydrolase (PARG). PARP-1 mediates acute neuronal cell death induced by a variety of insults including cerebral ischemia, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced Parkinsonism, and CNS trauma. While PARP-1 is localized to the nucleus, PARG resides in both the nucleus and cytoplasm. Surprisingly, there appears to be only one gene encoding PARG activity, which has been characterized in vitro to generate different splice variants, in contrast to the growing family of PARPs. Little is known regarding the spatial and functional relationships of PARG and PARP-1. Here we evaluate PARG expression in the brain and its cellular and subcellular distribution in relation to PARP-1. Anti-PARG (α-PARG) antibodies raised in rabbits using a purified 30 kDa C-terminal fragment of murine PARG recognize a single band at 111 kDa in the brain. Western blot analysis also shows that PARG and PARP-1 are evenly distributed throughout the brain. Immunohistochemical studies using α-PARG antibodies reveal punctate cytosolic staining, whereas anti-PARP-1 (α-PARP-1) antibodies demonstrate nuclear staining. PARG is enriched in the mitochondrial fraction together with manganese superoxide dismutase (MnSOD) and cytochrome C (Cyt C) following whole brain subcellular fractionation and Western blot analysis. Confocal microscopy confirms the co-localization of PARG and Cyt C. Finally, PARG translocation to the nucleus is triggered by NMDA-induced PARP-1 activation. Therefore, the subcellular segregation of PARG in the mitochondria and PARP-1 in the nucleus suggests that PARG translocation is necessary for their functional interaction. This translocation is PARP-1 dependent, further demonstrating a functional interaction of PARP-1 and PARG in the brain.

KW - NMDA

KW - immunostaining

KW - mitochondria

KW - nucleus

KW - subcellular fractionation

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AN - SCOPUS:34547783237

SN - 0306-4522

VL - 148

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JO - Neuroscience

JF - Neuroscience

IS - 1

ER -

Spatial and functional relationship between poly(ADP-ribose) polymerase-1 and poly(ADP-ribose) glycohydrolase in the brain

Abstract

Keywords

ASJC Scopus subject areas

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