Department of Toxicology, NC State University, Faculty, Yoshiaki Tsuji, Ph.D.

Mailing address:
Department of Environmental
and Molecular Toxicology
Box 7633, NC State University
Raleigh, NC 27695-7633

Shipping address:
Suite 1104, 850 Main Campus Dr.
Raleigh, NC 27606

Phone 919.515.2274
Fax 919.515.7169

Paul

Kensuke

Yoshiaki Tsuji, PhD
Associate Professor
Department of Environmental and Molecular Toxicology

Phone: 919-513-1106
Fax: 919-515-7169
e-mail: yoshiaki_tsuji@ncsu.edu

Education
Ph.D. in Cell Biology, Hiroshima University School of Medicine, Hiroshima, Japan
Postdoctoral Fellow, Stanford University School of Medicine, Palo Alto, California

Research Interests

Signaling Pathways Leading to Transcriptional Activation of Antioxidant Genes in Oxidative Stress
Chromatin Remodeling and Gene Regulation
Disorder of Iron Homeostasis and Neurodegeneration
Frataxin and Mitochondrial Oxidative Stress

Research Interests and Plans

Our research interests have been to define the molecular mechanisms by which cellular susceptibility to cytotoxic agents and stresses are determined in normal and cancer cells. We are particularly interested in the roles and regulation of genes encoding key proteins involved in iron metabolism . Within this area, we are currently studying the oxidative stress -mediated regulation of ferritin and frataxin. While iron plays an essential role in many cellular functions, excess iron is potentially harmful because it can catalyze the formation of reactive oxygen species (ROS) via the Fenton chemistry. Ferritin, a major iron storage protein that plays a prominent role in maintaining intracellular iron homeostasis, serves as a suppressor of the ROS generation by sequestration of intracellular excess free-iron. Frataxin is a mitochondrial protein that appears to regulate iron homeostasis and oxidative stress in mitochondria. Friedreich ataxia, the most frequent inherited ataxia, is associated with an intronic GAA repeat expansion of the frataxin gene, resulting in reduced transcription of the frataxin gene and neurodegeneration by unidentified molecular mechanisms.

Paul Ray (Graduate Student)

Ferritin and Neurodegeneration

My current research interests involve the regulation of iron homeostasis and its involvement in disease processes. Of primary focus is the iron binding protein ferritin, which serves as a storage depot for intracellular iron. In this manner iron is readily available for use, while at the same time prevented from over-accumulating in the cell, where it could take part in the Fenton reaction and produce reactive oxidative species.

a) Resveratrol and ferritin regulation

We have recently discovered that the polyphenolic compound resveratrol, found in red grape skins, affects transcriptional regulation of the ferritin H gene. In this capacity, resveratrol serves as an anti-oxidant by preventing excess iron from producing oxidative stress via the Fenton reaction. My project is focused on elucidating the pathway of resveratrol mediated ferritin H gene regulation, from cellular signaling molecules to transcription factors, to the specific areas of the ferritin H gene promoter.

b) Organophosphates and iron homeostasis in neurodegenerative diseases

Oxidative stress and perturbed intracellular iron levels have been implicated in certain neurodegenerative diseases. Organophosphates, such as the nerve agent soman and the pesticide chlorpyrifos, have been shown to induce oxidative stress in neuronal cells. My research aim is to investigate ferritin regulation in neuronal organophosphate exposure as part of an inquiry into the role of organophosphorous compounds in neurodegenerative diseases.

Bo-Wen Huang (Graduate Student)

Characterization of novel regulators of the ferritin H ARE

Transcription factors dominate the physiology and play a pivotal role in gene regulation. We found that several b-zip transcription factors including ATF1 and JunD are involved in the regulation of the ferritin H gene via the ARE. To investigate molecular mechanisms by which these ARE binding proteins regulate the transcription of the ferritin H gene, we performed yeast-two hybrid screening and have been characterizing coregulators that interact with and regulate the transcription factors bound to the ARE. My projects focus on ATF1-upstream signaling pathway. We try to find out the upstream pathways which regulate ATF1 activity and how exogenous stimulators or stresses regulate ATF1 activity.

Kensuke Sakamoto (Visiting Graduate Student)

CREB, CREB-associated proteins and Neurodegeneration

Excess iron induces oxidative stress, which is implicated to cause several neurodegenerative disorders, such as Parkinson's disease (PD) and Alzheimer's disease (AD). However, iron has essential roles in various cellular events, therefore, fine-tuning intracellular iron level is important for neurons. We have recently identified CREB/ATF1 as transcriptional repressors of the ferritin H gene. To understand molecular mechanism by which CREB transcription factors repress the ARE enhancer activity, we performed yeast two-hybrid assay and have identified several CREB binding proteins harboring corepressor activities. Since CREB is an important transcription factor for viability and generation of neuronal cells, I will further characterize regulatory mechanism of CREB through interaction with binding proteins, as well as CREB-mediated regulation of iron metabolism through transcriptional regulation of the ferritin H gene in neuronal cells.

Other Research Projects

1. PI3K-AKT signaling pathway and chromatin remodeling

2. Frataxin and Mitochondrial Iron Metabolism

We are also interested in studying mitochondrial oxidative stress and iron homeostasis through understanding functions of frataxin. Friedreich Ataxia (FRDA) is an autosomal recessive disease (with an estimated prevalence of 1 in 50,000) characterized by progressive gait, limb ataxia, loss of position sense and cardiomyopathy. FRDA is most commonly caused by the hyperexpansion of a GAA repeat in the first intron of the frataxin gene, which results in a marked transcriptional repression of the frataxin gene. Frataxin is a mitochondrial protein conserved from yeast to man, but with no homology with proteins of known function.

The frataxin gene exhibits tissue-specific expression, whose mRNA is most abundant in the heart, followed by the liver and skeletal muscle. The disease phenotype is prominent in the heart tissue and nervous system, suggesting that the required amount of frataxin in each tissue may be variable. The disruption of the yeast frataxin homolog ( YFH1 ) in yeast strain (deltayfh1 ) causes an accumulation of iron in mitochondria and induced their sensitivity to oxidative stress (hydrogen peroxide and iron overload). Mitochondria isolated from cultured FRDA fibroblasts have been shown to contain more iron than mitochondria from control normal counterparts. These results suggest that frataxin is involved in regulation of mitochondrial iron metabolism, such as iron efflux. Thus, FRDA, caused by deficiency of frataxin, is a mitochondrial disorder mediated by oxidative stress, reminiscent of the clinically similar ataxia of autosomal recessive vitamin E deficiency. However, the molecular mechanisms that regulate transcription of frataxin and its tissue-specific expression, biological function and mitochondrial deficit associated with FRDA remain unclear.

We currently plan to: 1) identify the binding partners of frataxin by yeast two-hybrid assays, and then in vitro transfection experiments to identify the functional frataxin protein complex, 2) isolate and characterize the 5' enhancer/promoter region of the frataxin gene to define the tissue-specific and basal enhancer elements of the frataxin gene, and 3) assess roles of frataxin and interacting molecules in cellular susceptibility to cytotoxic and stress-inducing agents. These studies will lead us not only to understand the biological function of frataxin, but also to elucidate the strategies and agents to stimulate the transcription of the frataxin gene that could derepress frataxin transcription caused by the GAA repeat expansion in FRDA patients.

Former Lab members:

Graduate Students

Selected Publications

K. Sakamoto, K. Iwasaki, H. Sugiyama and Y.Tsuji. “Role of the Tumor Suppressor PTEN in Antioxidant Responsive Element-mediated Transcription and Associated Histone Modifications” Mol. Biol. Cell, 20: 1606-1617 (2009)

E.L. MacKenzie and Y. Tsuji . "Elevated Intracellular Calcium Increases Ferritin H Expression through an NFAT-independent Posttranscriptional Mechanism Involving mRNA Stabilization." Biochem. J. , 411: 107-113 (2008)

F. Zhang, W. Wang, Y. Tsuji, S.V. Torti and F.M. Torti. “Post-transcriptional Modulation of Iron Homeostasis during p53-dependent Growth Arrest" J. Biol. Chem., 283: 33911-33918 (2008)

E.L. MacKenzie, P. D. Ray and Y. Tsuji. “Role and Regulation of Ferritin H in Rotenone-mediated Mitochodrial Oxidative Stress” Free Radic. Biol. Med., 44: 1762-1771 (2008)

E.L. MacKenzie, K. Iwasaki and Y. Tsuji. "Intracellular Iron Transport and Storage: From Molecular Mechanisms to Health Implications." Antioxid. Redox Signaling, 10: 997-1030 (2008) Comprehensive Invited Review

K. Iwasaki, K. Hailemariam and Y. Tsuji. "PIAS3 Interacts with ATF1 and Regulates the Human Ferritin H Gene through an Antioxidant Responsive Element." J. Biol. Chem., 282: 22335-22343. (2007)

K. Iwasaki , E. L. MacKenzie , K. Hailemariam , K. Sakamoto and Y. Tsuji. "Hemin-mediated Regulation of an Antioxidant Responsive Element of the Human Ferritin H Gene and Role of Ref-1 during Erythroid Differentiation of K562 Cells." Mol. Cell. Biol., 26: 2845-2856. (2006)

E.Jennings-Gee, Y. Tsuji , E. C. Pietsch, E. Moran, J.S. Mymryk, F. M. Torti and S. V. Torti. "Coordinate Inhibition of Cytokine-mediated Induction of Ferritin H, Manganese Superoxide Dismutase, and Interleukin-6 by the Adenovirus E1A Oncogene." J. Biol. Chem., 281: 16428-16435. (2006)

Y. Tsuji. "JunD Activates Transcription of the Human Ferritin H Gene through an Antioxidant Response Element during Oxidative Stress." Oncogene, 24: 7567-7578. (2005)