Mol Cell Biochem (2011) 350:127134

DOI 10.1007/s11010-010-0690-4

Characterizing the novel protein p33MONOX

Manisha Mishra Noriko Inoue Klaus Heese

Received: 4 June 2010 / Accepted: 18 September 2010 / Published online: 14 December 2010
Springer Science+Business Media, LLC. 2010

Abstract The novel protein p33MONOX (p33Monooxygenase) was over-expressed in neuroblastoma cells demonstrating its inhibitory effect on the phosphorylation of
the App (amyloid precursor protein) and Bcl2 (B-cell
lymphoma 2) proteins but mediating higher activation of
Mapk1/3 (mitogen-activated protein kinase 1/3). We
employed a variety of cell biology techniques to show the
localization of p33MONOX to the cytoplasm of pyramidal
neurons in the mouse brain hippocampus. We also carried
out a yeast-two-hybrid screening plus co-immunoprecipitation and bio-informatics to determine COBRA1 (cofactor
of BRCA1 (breast cancer type 1)), NOL12 (nucleolar
protein 12), and PRNP (prion protein) as p33MONOXinteracting proteins. Bio-computational analyses revealed a
flavine-containing monooxygenase (FMO)-1 motif, thus
linking p33MONOX to a group of previously characterized proteins, the MICALs (molecule interacting with
CasL). Concluding, p33MONOX might regulate pre- and

Electronic supplementary material The online version of this

article (doi:10.1007/s11010-010-0690-4) contains supplementary
material, which is available to authorized users.
M. Mishra
K. Heese (&)
Department of Molecular and Cell Biology,
School of Biological Sciences, College of Science,
Nanyang Technological University, 60 Nanyang Drive,
637551 Singapore, Singapore
e-mail: kheese@ntu.edu.sg
N. Inoue
Medical Center for Translational Research,
Osaka University Hospital, Suita, Osaka, Japan

post-transcriptional control of dynamic processes related to

growth cone guidance.
Keywords Alzheimers disease
Apoptosis

Neurodegeneration

Introduction
Alzheimers Disease (AD) remains the most common cause
of dementia in all age groups, characterized by progressive
neurodegeneration and profound cognitive deficits [14].
Much of what is known about AD revolves around the
amyloid precursor protein (APP) and presenilin-1 (PSEN1)
and PSEN2 [5]. However, there remains a myriad of other
proteins that could possibly play an equally crucial role in the
development of AD. Although the etiology of sporadic AD is
poorly understood, there is evidence that aberrant iron
deposition, oxidative stress and mitochondria insufficiency
play a role in the pathogenesis of sporadic AD and other
aging-related neurodegenerative disorders. The excessive
generation of free radicals may promote neurofibrillary
tangle (NFT) formation as well as amyloid deposition in AD
brains. Conversely, the neurotoxic effects of certain amyloid
fragments may be mediated by free radical intermediates
[68]. P33MONOX was discovered in our recent study on
brain site-specific gene-expression analysis, when we compared the gene-expression pattern in the temporal and
occipital lobe of early stage AD subjects with control
patients [9]. In that study, p33MONOX, a novel gene with
unknown functions as yet, was identified to be down-regulated in the occipital lobe of an early stage AD patient. In the
current study, we characterized the potential biological significance of p33MONOX using molecular and cell biological as well as bio-computational analyses.

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Materials and methods

Reagents
All reagents used for experiments were purchased from
Sigma-Aldrich (Milwaukee, WI, USA) unless otherwise
stated.
Cell culture and transfection
Rat B104 and human SHSY5Y neuroblastoma cells as well
as rat PC12 cells (all from American Type Culture Collection (ATCC, Manassas, VA, USA)) were maintained in
Dulbeccos Modified Eagle Medium (D-MEM/F12(1:1))
plus 10% fetal bovine serum (FBS; Invitrogen, (Gibco),
Carlsbad, CA, USA) at 37C in humidified 5% CO2/95%
air. A p33Monox expression construct was generated by
inserting rat p33Monox cDNA in-frame with the red fluorescent protein (DsRed) (pDsRed-Express-N1; BD Biosciences Clontech, Palo Alto, CA, USA) at the C-terminus
of p33Monox (p33-CT-DsRed). B104 cells were transiently transfected with the p33-CT-DsRed expression
vector using the Lipofectamine 2000 (Invitrogen) transfection reagent (according to the manufacturers protocol)
and maintained in D-MEM medium containing 10% FBS
at 37C. The transfected cells were then visualized by
fluorescence microscopy (Nikon eclipse TE2000U, Nikon,
Singapore). SHSY5Y, B104, and PC12 cells were stably
transfected using a lentivirus expression system (p33 in
EF.CMV.Gfp-Lenti-vector (elongation factor 1 alpha,
cytomegalovirus promotors, green fluorescent protein;
JHU-55, ATCC); co-expression of p33Monox and green
Gfp) (control, mock-transfection) according to the manufacturers protocol (Invitrogen) as briefly described in
Supplementary materials and methods [10, 11].
Western blot analyses
Total protein cell lysates were subjected to western blot
analyses as described previously (Supplementary materials
and methods) [9, 10].
Animal material, immunohistochemistry (IHC)
and immunocytochemistry (ICC)
Experimental methods, including the killing of animals,
were performed in accordance with the International
Guiding Principles for Animal Research (WHO) and
approved by the local Institutional Animal Care & Use
Committee (NTU-IACUC). Mouse tissues were isolated
(C57BL/6J mice from the Animal Facility Centre at the
National University (NUS) of Singapore) after humane
killing of the animals using approved anaesthetic methods.

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Mol Cell Biochem (2011) 350:127134

Mouse brain perfusion, IHC, and ICC were performed

as described previously (Supplementary materials and
methods) [11].
ProQuestTM two-hybrid-system with GatewayTM
technology
The two-hybrid-system is an in vivo yeast-based system
that identifies the interaction between two proteins (here
X = p33MONOX and Y = human brain cDNA library or
COBRA1) by reconstituting an active transcription factor.
The analysis was performed according to the manufacturers protocol (Invitrogens brain ProQuestTM twohybrid-system, Singapore) using p33MONOX as bait. In
the ProQuestTM two-hybrid-system, in comparison to
standard two-hybrid-systems, false positives are reduced
because three independent transcription events (from distinct promoters) must occur at independent chromosomal
loci. Positive clones were confirmed by retransformation
assays and protein co-immunoprecipitation (Co-IP, Supplementary materials and methods) [10].

Results
p33MONOX protein sequence analysis
Bio-computational analyses of the p33MONOX protein
sequence among the species of human, mouse, and rat
showed high sequence similarity (Fig. 1) pointing to the
possibility that p33MONOX plays a crucial role that is
evolutionarily conserved.
The most encouraging information about p33MONOX
is the presence of a flavine-containing monooxygenase
(FMO)-1 motif. The proteins comprising the FMO motif
belong to a family of microsomal NADPH (nicotinamide
adenine dinucleotide phosphate)- and oxygen-dependent
flavoenzymes (with flavin adenine nucleotide (FAD) as a
co-factor) that are distributed ubiquitously in mammalian
species, and catalyze the oxidation of soft nucleophilic
heteroatom centers in drugs, pesticides, and xenobiotics,
using nucleotides as electron donors. FMO-1 catalyzes the
N-oxygenation of secondary and tertiary amines. In some
contexts, while performing the oxidation, they can generate
reactive oxygen species (ROS) [1215]. Thus,
p33MONOX protein is more likely to be a NADPHdependent oxidoreductase.
Sub-cellular localization and neuronal expression
of p33Monox in the mouse brain
We investigated the sub-cellular localization of p33Monox
to obtain more information about its physiological role,

Mol Cell Biochem (2011) 350:127134

129

Fig. 1 Characteristic features

of the p33MONOX protein
sequence. Aligned protein
sequences of p33MONOX in
the human (H, black), mouse
(M, blue), and rat (R, green)
species, with red alphabets
denoting the varying amino
acid. There is evidently a high
degree of conservation in the
protein sequences amongst the
species, suggesting a pivotal
functional significance of
p33MONOX. Bio-informatical
analyses of p33MONOXs
protein sequence revealed a
potential flavine-containing
monooxygenase (FMO)-1 motif
and several Ser-/Thrphosphorylation sites

distribution, and site of activity in the cell. For this purpose, a p33Monox-DsRed fluorescent fusion protein was
transiently expressed in B104 neuroblastoma cells. Using
fluorescence microscopy, we show that p33Monox
expression was confined to the neuronal cytoplasm thus
confirming the bio-computational analysis data (Fig. 2a).
Besides, ICC of nerve growth factor (Ngf)-differentiated
PC12 cells also verified the localization of p33Monox in
the cytoplasm with a substantial expression in the axonal
growth cone (Fig. 2b).
Furthermore, we conducted an IHC analysis of the
mouse brain for the p33Monox protein expression and
localization. This analysis revealed that p33Monox was

physiologically expressed in neuronal pyramidal cells of

the hippocampus and also in the neurons of the cortex
(Fig. 3).
p33Monox-mediated neuronal signaling
To further corroborate the physiological significance of
p33Monox, we analyzed its effect on the activation of
pivotal proteins involved in neuronal survival and differentiation. Interestingly, upon over-expression in neuronal
cells, p33Monox inhibited the phosphorylation of App
(reduced Ab formation [16]) and Bcl2 (the functional significance of the dynamic phosphorylation status (regulated

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Mol Cell Biochem (2011) 350:127134

Fig. 2 Sub-cellular localization

of p33Monox. a B104 cells
were transfected with a
p33-CT-DsRed expression
vector as described in
Supplementary materials and
methods. Microscopic picture of
a representative B104 cell under
bright-field, red fluorescence
(revealing cytoplasmic
localization of p33Monox), and
UV light (indicating nuclear
DAPI staining), respectively;
scale bar = 20 lm. b Co-ICC
of Ngf-differentiated PC12 cells
with p33Monox (green) and
Tuba1a (tubulin, red) or Syp
(synaptophysin, red) as
indicated. DAPI staining was
used to indicate the nucleus.
Representative pictures are
shown. Pictures show the strong
co-localization (merged, yellow)
of p33Monox with tubulin in the
cytoplasm and with Syp in the
axonal growth cones (arrows).
Scale bar = 50 lm

by various kinases and protein phosphatase PP2A) has been

discussed conflictive [1722]) while Erk1/2 (Mapk1/3) was
activated (sustained phosphorylation is required for neuronal differentiation [23]) (Fig. 4a).
p33MONOX interacts with COBRA1, NOL12,
and PRNP
Additional clarification about p33Monoxs potential cellular function was obtained by the yeast-two-hybrid screen
combined with bio-informatic analyses (NCBI (National
Center for Biotechnology information), EMBL-EBI
(European Bioinformatics Institute), and SIB (Swiss Institute of Bioinformatics, (Swiss-Prot & Tremble, ExPASy,
and Proteomics tools)) databases were used). The analyses
revealed that p33MONOX interacts with several proteins

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involved in the control of gene transcription, such

as COBRA1 (the co-factor of BRCA1 is a newly characterized member of the negative elongation factor (NELF)
complex; confirmed by Co-IP (Fig. 4b)) [2428], NOL12
(nucleolar protein 12, also known as ribosomal RNA processing protein 17) [29, 30], and the prion protein (PRNP
or PrP with isoform 2 as a potential growth suppressor
that arrests the cell cycle at the G0/G1 phase) [3133]
(Supplementary Tables 1 and 2).
NOL12 itself interacts with several other proteins
known to be pivotal regulators of gene transcription and
cell cycle progression such as: SAP18 (or Sin3A-associated
protein), CDK4 (cyclin-dependent kinase 4), SF3B3 (subunit 3 of the splicing factor 3b protein complex), and
SLC25A38 (solute carrier family 25, member 38), a
mitochondrial carrier protein that is widely expressed

Mol Cell Biochem (2011) 350:127134

131

Fig. 3 Co-IHC analysis of p33Monox (green) and Mtap2 (red) in

the mouse brain revealed that p33Monox is expressed in the
cytoplasm of pyramidal neurons in the hippocampus. Top left entire
hippocampus formation including CA1, CA2, CA3 regions, and the
dentate gyrus. Scale bar = 100 lm. Right enlarged CA1 regions.

Scale bar = 20 lm. Bottom/left further magnified picture of the CA1

region, clearly shows the cytoplasmic localization of p33Monox.
Scale bar = 5 lm. Bottom/right representative picture taken form the
cortex area thus indicating that p33Monox is generally expressed in
neurons in the mouse brain. Scale bar = 20 lm

in the central nervous system [34]. Interestingly, by direct

interaction NOL12 links p33Monox to SOD2 (superoxide
dismutase 2). This protein is a member of the iron/
manganese superoxide dismutase family that binds to the
superoxide byproducts of oxidative phosphorylation and
converts them to hydrogen peroxide and diatomic oxygen.
Mutations in this gene have been associated with premature
aging and sporadic motor neuron disease [35, 36] (further
details in Supplementary Table 2).

that p33Monox showed an interesting inhibition of the

phosphorylation of App and Bcl2 as well as an enhanced
activation of Mapk1/3 [16, 20, 23]. We also found that
p33Monox forms a complex with Cobra1, Nol12, and Prnp.
Cobra1, also known as Nelf-b that is involved in controlling axonal growth [25, 26], associates with the product of
the breast cancer susceptibility gene Brca1, thus hinting at
a plausible role of p33Monox in sequestering Cobra1 and
thereby leading to profound effects on gene transcription
signals during dynamic neurite outgrowth processes.
Consequently, p33Monox can be likened to a group of
cytosolic proteins, the MICALs (molecule interacting with
CasL), which are also oxidoreductases that utilize FAD as a
co-factor and communicate via ROS [3739]. During the
past decade, we have begun to recognize that controlled
production of ROS and regulated redox modifications of

Discussion
In the present study, we revealed the specific sites of
p33Monox expression and its intensive localization in
neural axonal growth cones. Apart from this, we observed

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Mol Cell Biochem (2011) 350:127134

Fig. 5 Schematic illustration of the potential action of p33MONOX

in mediating neuronal survival, differentiation, and axonal outgrowth.
In light of experimental and bio-informatical evidences it can be
speculated that p33MONOX may act as platform to recruit downstream effectors (e.g., COBRA1, NOL12, PRNP, SOD2, NF-jB,
ROS, kinases, or phosphatases) to their site of action. The activity of
these effectors could then be selectively modulated by redox
modifications of key amino acid residues that could either be the
direct effect of p33MONOXs monooxygenase activity or be the
indirect consequence of a local increase in ROS. P33MONOXmediated de-phosphorylation of App and Bcl2 as well as sustained
phosphorylation of Mapk1/3 are required for the control of neuronal
survival, differentiation, and growth-cone extension
Fig. 4 a p33Monox inhibits the phosphorylation of pivotal signaling
molecules as indicated. Neuronal B104 cells were transfected with
p33Monox as described in Supplementary materials and methods.
Thereafter, the phosphorylation status was checked by western
blotting. C control, non-transfected, GFP mock-transfection with
GFP, p33Monox p33Monox-transfected. b p33MONOX Co-IP. Upon
transfection of neuronal SHSY5Y cells, Co-IP was performed as
described in Supplementary materials and methods to confirm the
interaction between p33MONOX and COBRA1. Control Co-IP with
unspecific serum

transcription factors or enzymes (such as kinases and

phosphatases) are an essential part of signal transduction
pathways [4042]. Similar to p33Monox, the MICALs are
expressed in neuronal axons that associate with several
cytoskeletal/-associated proteins, and are required for
semaphorin-mediated repulsive axon guidance that is crucial for neuronal development. P33Monox may be another
candidate for directly mediating the cytoskeletal alterations
characteristic of semaphorin signaling and could be a novel
target for the attenuation of axonal repulsion (Fig. 5).
Given the presence of high amounts of ROS and other
oxidants in the spinal cord after injury [43], and that aging
and AD have previously been closely linked to the accumulation of oxidative stress [4446], regulation of redox
signaling using antioxidants and specific enzyme inhibitors
may be a powerful approach for encouraging neuronal
regeneration [37]. The plausible link between MICAL,
p33Monox, and SOD2 points to a role of p33Monox

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in controlling ROS that have been recently suggested to be a

key factor in the cellular changes of an AD brain as several
reports have suggested that mitochondrial abnormalities and
oxidative stress play a role in sporadic AD [4749]. For
instance, the heme-oxygenase-1 (HO-1), a member of the
stress protein superfamily that operates with the NADPH
cytochrome P450 reductase to oxidize heme, is widely
accepted as a sensitive and fairly ubiquitous up-regulated
marker of oxidative stress. It has also been shown to be
consistently co-localized to NFTs and senile plaques in AD
brains [5052].
In conclusion, our data has shed new insights on the
possible involvement of the novel protein p33Monox in the
regulation of neuronal survival, differentiation, and axonal
outgrowth and the connection among p33Monox, oxidative
stress and AD will be a gripping topic for future investigations, especially in view of potential antioxidant therapies.
Acknowledgments This study was supported by an A*STAR grant
(BMRC/04/1/22/19/360) to K.H. We thank Ms H.J. Tang and
S. Yusof (both from the School of Biological Sciences, Nanyang
Technical University) for technical assistance. We are particularly
grateful to Prof. Dr. R. Li (Department of Molecular Medicine,
Institute of Biotechnology, The University of Texas Health Science
Center, 15355 Lambda Drive, San Antonio, TX, 78245-3207, USA)
for providing us the anti-COBRA1 antibody.

Mol Cell Biochem (2011) 350:127134

References
1. Welsh KA, Butters N, Hughes JP, Mohs RC, Heyman A (1992)
Detection and staging of dementia in Alzheimers disease. Use of
the neuropsychological measures developed for the Consortium
to Establish a Registry for Alzheimers Disease. Arch Neurol
49:448452
2. Terry RD, Masliah E, Salmon DP, Butters N, DeTeresa R, Hill R,
Hansen LA, Katzman R (1991) Physical basis of cognitive
alterations in Alzheimers disease: synapse loss is the major
correlate of cognitive impairment. Ann Neurol 30:572580
3. Heese K, Akatsu H (2006) Alzheimers diseasean interactive
perspective. Curr Alzheimer Res 3:109121
4. Mattson MP (2004) Pathways towards and away from Alzheimers disease. Nature 430:631639
5. Selkoe DJ, Schenk D (2003) Alzheimers disease: molecular
understanding predicts amyloid-based therapeutics. Annu Rev
Pharmacol Toxicol 43:545584
6. Butterfield DA (2002) Amyloid beta-peptide (1-42)-induced
oxidative stress and neurotoxicity: implications for neurodegeneration in Alzheimers disease brain. A review. Free Radic Res
36:13071313
7. Butterfield DA, Castegna A, Lauderback CM, Drake J (2002)
Evidence that amyloid beta-peptide-induced lipid peroxidation
and its sequelae in Alzheimers disease brain contribute to neuronal death. Neurobiol Aging 23:655664
8. Pappolla MA, Chyan YJ, Omar RA, Hsiao K, Perry G, Smith
MA, Bozner P (1998) Evidence of oxidative stress and in vivo
neurotoxicity of beta-amyloid in a transgenic mouse model of
Alzheimers disease: a chronic oxidative paradigm for testing
antioxidant therapies in vivo. Am J Pathol 152:871877
9. Yokota T, Mishra M, Akatsu H, Tani Y, Miyauchi T, Yamamoto
T, Kosaka K, Nagai Y, Sawada T, Heese K (2006) Brain sitespecific gene expression analysis in Alzheimers disease patients.
Eur J Clin Invest 36:820830
10. Heese K, Yamada T, Akatsu H, Yamamoto T, Kosaka K, Nagai
Y, Sawada T (2004) Characterizing the new transcription regulator protein p60TRP. J Cell Biochem 91:10301042
11. Nehar S, Mishra M, Heese K (2009) Identification and characterisation of the novel amyloid-beta peptide-induced protein p17.
FEBS Lett 583:32473253
12. Massey V (1995) Introduction: flavoprotein structure and mechanism. FASEB J 9:473475
13. Yeung CK, Lang DH, Thummel KE, Rettie AE (2000) Immunoquantitation of FMO1 in human liver, kidney, and intestine.
Drug Metab Dispos 28:11071111
14. Ziegler DM (1990) Flavin-containing monooxygenases: enzymes
adapted for multisubstrate specificity. Trends Pharmacol Sci
11:321324
15. Massey V (1994) Activation of molecular oxygen by flavins and
flavoproteins. J Biol Chem 269:2245922462
16. Lee MS, Kao SC, Lemere CA, Xia W, Tseng HC, Zhou Y, Neve
R, Ahlijanian MK, Tsai LH (2003) APP processing is regulated
by cytoplasmic phosphorylation. J Cell Biol 163:8395
17. Ito T, Deng X, Carr B, May WS (1997) Bcl-2 phosphorylation
required for anti-apoptosis function. J Biol Chem 272:1167111673
18. Deng X, Ito T, Carr B, Mumby M, May WS Jr (1998) Reversible
phosphorylation of Bcl2 following interleukin 3 or bryostatin 1 is
mediated by direct interaction with protein phosphatase 2A.
J Biol Chem 273:3415734163
19. Deng X, Gao F, Flagg T, May WS Jr (2004) Mono- and multisite
phosphorylation enhances Bcl2s antiapoptotic function and
inhibition of cell cycle entry functions. Proc Natl Acad Sci USA
101:153158

133
20. Yamamoto K, Ichijo H, Korsmeyer SJ (1999) BCL-2 is phosphorylated and inactivated by an ASK1/Jun N-terminal protein
kinase pathway normally activated at G(2)/M. Mol Cell Biol
19:84698478
21. Chang BS, Minn AJ, Muchmore SW, Fesik SW, Thompson CB
(1997) Identification of a novel regulatory domain in Bcl-X(L)
and Bcl-2. EMBO J 16:968977
22. Liu XA, Liao K, Liu R, Wang HH, Zhang Y, Zhang Q, Wang Q,
Li HL, Tian Q, Wang JZ (2010) Tau dephosphorylation potentiates apoptosis by mechanisms involving a failed dephosphorylation/activation of Bcl-2. J Alzheimers Dis 19:953962
23. Kholodenko BN (2007) Untangling the signalling wires. Nat Cell
Biol 9:247249
24. McChesney PA, Aiyar SE, Lee OJ, Zaika A, Moskaluk C, Li R,
El-Rifai W (2006) Cofactor of BRCA1: a novel transcription
factor regulator in upper gastrointestinal adenocarcinomas. Cancer Res 66:13461353
25. Kramer PR, Wray S (2001) Nasal embryonic LHRH factor
(NELF) expression within the CNS and PNS of the rodent. Brain
Res Gene Expr Patterns 1:2326
26. Kramer PR, Wray S (2000) Novel gene expressed in nasal region
influences outgrowth of olfactory axons and migration of
luteinizing hormone-releasing hormone (LHRH) neurons. Genes
Dev 14:18241834
27. Sun J, Blair AL, Aiyar SE, Li R (2007) Cofactor of BRCA1
modulates androgen-dependent transcription and alternative
splicing. J Steroid Biochem Mol Biol 107:131139
28. Narita T, Yamaguchi Y, Yano K, Sugimoto S, Chanarat S, Wada
T, Kim DK, Hasegawa J, Omori M, Inukai N, Endoh M, Yamada
T, Handa H (2003) Human transcription elongation factor NELF:
identification of novel subunits and reconstitution of the functionally active complex. Mol Cell Biol 23:18631873
29. Suzuki S, Kanno M, Fujiwara T, Sugiyama H, Yokoyama A,
Takahashi H, Tanaka J (2006) Molecular cloning and characterization of Nop25, a novel nucleolar RNA binding protein, highly
conserved in vertebrate species. Exp Cell Res 312:10311041
30. Stelzl U, Worm U, Lalowski M, Haenig C, Brembeck FH,
Goehler H, Stroedicke M, Zenkner M, Schoenherr A, Koeppen S,
Timm J, Mintzlaff S, Abraham C, Bock N, Kietzmann S, Goedde
A, Toksoz E, Droege A, Krobitsch S, Korn B, Birchmeier W,
Lehrach H, Wanker EE (2005) A human proteinprotein interaction network: a resource for annotating the proteome. Cell
122:957968
31. Juanes ME, Elvira G, Garcia-Grande A, Calero M, Gasset M (2009)
Biosynthesis of prion protein nucleocytoplasmic isoforms by
alternative initiation of translation. J Biol Chem 284:27872794
32. Satoh J, Obayashi S, Misawa T, Sumiyoshi K, Oosumi K, Tabunoki H (2009) Protein microarray analysis identifies human
cellular prion protein interactors. Neuropathol Appl Neurobiol
35:1635
33. Haigh CL, Lewis VA, Vella LJ, Masters CL, Hill AF, Lawson
VA, Collins SJ (2009) PrPC-related signal transduction is influenced by copper, membrane integrity and the alpha cleavage site.
Cell Res 19:10621078
34. Haitina T, Lindblom J, Renstrom T, Fredriksson R (2006)
Fourteen novel human members of mitochondrial solute carrier
family 25 (SLC25) widely expressed in the central nervous system. Genomics 88:779790
35. Miao L, St Clair DK (2009) Regulation of superoxide dismutase
genes: implications in disease. Free Radic Biol Med 47:344356
36. Lynn S, Huang EJ, Elchuri S, Naeemuddin M, Nishinaka Y,
Yodoi J, Ferriero DM, Epstein CJ, Huang TT (2005) Selective
neuronal vulnerability and inadequate stress response in superoxide dismutase mutant mice. Free Radic Biol Med 38:817828

123

134
37. Terman JR, Mao T, Pasterkamp RJ, Yu HH, Kolodkin AL (2002)
MICALs, a family of conserved flavoprotein oxidoreductases,
function in plexin-mediated axonal repulsion. Cell 109:887900
38. Massey V, Palmer G (1966) On the existence of spectrally distinct classes of flavoprotein semiquinones. A new method for the
quantitative production of flavoprotein semiquinones. Biochemistry 5:31813189
39. Ventura A, Pelicci PG (2002) Semaphorins: green light for redox
signaling? Sci STKE 2002: pe44
40. Kamata H, Hirata H (1999) Redox regulation of cellular signalling. Cell Signal 11:114
41. Finkel T (1998) Oxygen radicals and signaling. Curr Opin Cell
Biol 10:248253
42. Meng TC, Fukada T, Tonks NK (2002) Reversible oxidation and
inactivation of protein tyrosine phosphatases in vivo. Mol Cell
9:387399
43. Juurlink BH, Paterson PG (1998) Review of oxidative stress in
brain and spinal cord injury: suggestions for pharmacological and
nutritional management strategies. J Spinal Cord Med 21:309334
44. Lovell MA, Ehmann WD, Butler SM, Markesbery WR (1995)
Elevated thiobarbituric acid-reactive substances and antioxidant
enzyme activity in the brain in Alzheimers disease. Neurology
45:15941601
45. Schuessel K, Leutner S, Cairns NJ, Muller WE, Eckert A (2004)
Impact of gender on upregulation of antioxidant defence mechanisms in Alzheimers disease brain. J Neural Transm 111:11671182

123

Mol Cell Biochem (2011) 350:127134

46. Nunomura A, Chiba S, Lippa CF, Cras P, Kalaria RN, Takeda A,
Honda K, Smith MA, Perry G (2004) Neuronal RNA oxidation is
a prominent feature of familial Alzheimers disease. Neurobiol
Dis 17:108113
47. Hirai K, Aliev G, Nunomura A, Fujioka H, Russell RL, Atwood
CS, Johnson AB, Kress Y, Vinters HV, Tabaton M, Shimohama
S, Cash AD, Siedlak SL, Harris PL, Jones PK, Petersen RB, Perry
G, Smith MA (2001) Mitochondrial abnormalities in Alzheimers
disease. J Neurosci 21:30173023
48. Behl C (2005) Oxidative stress in Alzheimers disease: implications for prevention and therapy. Subcell Biochem 38:6578
49. Reddy PH (2006) Amyloid precursor protein-mediated free radicals and oxidative damage: implications for the development and
progression of Alzheimers disease. J Neurochem 96:113
50. Premkumar DR, Smith MA, Richey PL, Petersen RB, Castellani
R, Kutty RK, Wiggert B, Perry G, Kalaria RN (1995) Induction of
heme oxygenase-1 mRNA and protein in neocortex and cerebral
vessels in Alzheimers disease. J Neurochem 65:13991402
51. Schipper HM, Cisse S, Stopa EG (1995) Expression of heme
oxygenase-1 in the senescent and Alzheimer-diseased brain. Ann
Neurol 37:758768
52. Smith MA, Kutty RK, Richey PL, Yan SD, Stern D, Chader GJ,
Wiggert B, Petersen RB, Perry G (1994) Heme oxygenase-1 is
associated with the neurofibrillary pathology of Alzheimers
disease. Am J Pathol 145:4247