
Visiting Professor Hisao MASAI
Tokyo Metroporitan Institute of Medical Science
TEL: +81-3-5316-3231
E-mail: masai-hs{at}igakuken.or.jp
Lab HP
【Key Words】DNA replication, genome stability, cell cycle, chromatin architecture, DNA replication stress checkpoint, embryonic stem cells, G-quadruplex, RNA-DNA hybrids, cancer cells
Precise duplication of genetic materials is central to the stable maintenance of genomes through generations. Defects in the genome copying processes would generate genomic instability which could ultimately result in various diseases including cancer. The goal of our studies is to understand the molecular basis of how the huge genomes are accurately replicated and the precise copies of the genetic materials are inherited to the next generation. Three billion base pairs of the human genome (2 meter long) are replicated with almost no errors during the 6-8 hr time span of the cell cycle. This requires an extreme level of coordination of temporal and spatial arrangements of chromatin organization and signaling events for initiation of DNA replication (13).
We recently discovered novel and crucial roles of non-standard DNA structures in regulation of DNA replication and transcription. Notably, we found that G-quadruplex structures (Fig. 1), which are widely present on genomes (estimated to be at more than 370,000 locations on the human genome), regulate organization of chromatin architecture and initiation of DNA replication (Fig. 2; 6). Recent reports indicate crucial roles of these non-canonical DNA/RNA structures in diverse biological reactions as well as in pathogenesis of diseases. One of our major goals is to establish a novel principle of the genome by elucidating the fundamental and universal functions of G-quadruplex and other non-B type DNA structures in regulation of various genome functions. Through these efforts, we will also explore the possibility that mutations found in various diseases including cancer are related to alteration of these non-B DNA structures, which are likely to be essential components of genomes but somehow have been disregarded in the past.
Our other major projects include 1) Maintenance of genome integrity and its failure as a cause of diseases: Molecular dissection of cellular responses to replication stress, a major trigger for oncogenesis, and elucidation of mechanisms by which stalled forks are processed and the genome is protected from various insults, and of how the failure of this process leads to diseases and senescence (4,5,8). 2) Chromosome dynamics that determines cell fate and regulates cell proliferation: Elucidation of mechanisms regulating temporal and spatial regulation of genome duplication as well as coordination of replication, repair, recombination and transcription (1,3,7,9,10). 3) Unraveling the universal mechanisms of origin firing and its regulation (genetic and enzymological studies using E.coli as a model). 4) DNA replication and development: Understanding the roles of replication factors or replication timing regulation during development/ differentiation processes or during the functioning of various tissues and organs. We have recently found potential novel and critical roles of Cdc7 kinase in development of brain. 5) DNA replication as target of anti-caner drugs: we have developed specific inhibitors of a replication factor as novel anti-cancer drug, and try to find a highly efficient and side-effect-free therapy for cancer patients by novel combination of cell cycle modulation.
To achieve these goals, we are using E.coli, fission yeast, various mammalian cell lines, embryonic stem cells and model animals. We would like ultimately to apply the basic knowledge on the mechanisms of stable genome maintenance to the diagnosis and therapy of the relevant diseases including cancer.
We are recruiting highly motivated and interested individuals who are communicative and can share excitement with us in the laboratory. We have had students from many foreign countries including Korea, Malaysia, Taiwan, China, Canada, Italy, France, USA and Germany and have been excited to have many different cultures in our laboratory. Please feel free to contact us at any time through e-mail or by telephone.
Selected publications
1 Moriyama et al. (2018) J. Biol. Chem. In press
2 You, Z. and Masai, H. (2017) Nucleic Acids Res. 45, 6495-6506.
3 Toteva et al. (2017) Proc. Natl. Acad. Sci. USA. 114, 1093-1098.
4 Matsumoto et al. (2017) Mol Cell. Biol. 37, pii: e00355-16.
5 Yang et al. (2016) Nature Communications 7:12135
6 Kanoh, Y. et al. (2015) Nature Struct. Mol. Biol. 22, 889-897.
7 Yamazaki, S. et al. (2013) Trends in Genetics. 29, 449-460.
8 Yamada, M. et al. (2013) Genes and Development 27:2459-72.
9 Yamazaki, S. et al. (2012) EMBO J. 31, 3167-3177.
10 Hayano, M. et al. (2012) Genes and Development, 26,137-150.
11 Hayano, M. et al. (2011) Mol. Cell. Biol. 31, 2380-2389.
12 Matsumoto, S. et al. (2011) J. Cell Biol. 195, 387-401.
13 Masai, H. et al. (2010) Ann. Rev. Biochem. 79, 89-130.
Visiting Professor Masanori ITOKAWA
Tokyo Metroporitan Institute of Medical Science
TEL: +81-3-5316-3228
E-mail: Itokawa-ms{at}igakuken.or.jp
Lab HP
【Key Words】mental illness, mind, brain, molecular biology, genome
Why is homo sapience suffered from mental illnesses? Numerous numbers of people in field of religion or philosophy had ever investigated the maze far past. Only three hundred years have passed since medical sciences involved in this theme. We are challenging to resolve the twister interwoven with brain and mind by using methods and tools of biology.
Functional psychiatric disease is the brain disorder causing emotional and thinking difficulty without any abnormal sings in electric encephalography or brain imaging. Schizophrenia is the major one of those as well as mood disorder
We perform genomic and metabolome analysis using blood samples from patients with schizophrenia in order to reveal pathophysiology of the disease. We create animal
and culture cell-based model utilizing genetic polymorphisms and aberrant metabolism seen in the patients.
Human iPS cells induced from a schizophrenic patient carrying the rare genetic variation were differentiated to neural cells to be analyzed for investigation of pathophysiology of the disease.
Schizophrenia is a common disease that the prevalence is around 1% of population at any region of the world. Why has schizophrenia survived natural selection during human evolution? We are also seeking answer of the question by using our models of animals and culture cells.
Ego-function such as self-identity is also disturbed in patients with schizophrenia. We challenge to reveal ego and self-consciousness, the fields that had ever been investigated by religion or philosophy far past,by using tools and methods of molecular biology.
Oxidative stress is a central mediator of advanced glycation end product (AGE) formation, and pyridoxamine[vitamin (vit)B6]] (biosynthesized from pyridoxal in vivo) is known to detoxify reactive carbonyl compounds (RCOs) via carbonyl-amine chemistry.Cellular removal of AGEs hinges largely upon the activity of the zinc metalloenzyme glyoxalase I (GLO1). We detected idiopathic carbonyl stress in a subpopulation of schizophrenia. We first found an interesting case carrying genetic defect of glyoxalase 1 (GLO1)that increased AGEs and decreased vitamin B6 since GLO1 detoxifies AGEs and vitamin B6 is carbonyl scavenger. We obtained 20% of patients showing carbonyl stress by the manner expanding concept of the case over the general schizophrenic patients. This manner can resolve the problem of research on schizophrenia derived from the heterogeneity of the disease. Genetic defect of GLO1 contributes to the stress by 5 time’s higher risk compared to that of intact gene. AGEs level was significantly correlated with negative symptoms of the patients. Pyridoxamine, active vitamin B6, could be the first medicine for negative symptoms of schizophrenia as most of the antipsychotic medicines are not effective for negative symptoms. We here present unique report of resolution of research difficulty due to heterogeneity of schizophrenia and possible discovery of the drug for negative symptoms of the disease.
Figure 1 . Plasma pentos idine accumulation and serum pyridoxal(vitamin B6) depletion.Levels of plasma pentos idine (A) and serum pyridoxal (B ) were analyzed us ing hig h-performance liquid chromatog raphy techniques .Values were compared using the Mann-Whitney U tes t (2 -tailed). Error bars indicate s tandard devi
Visiting Professor Yasushi SAEKI
Tokyo Metroporitan Institute of Medical Science
TEL: +81-3-6834-2329
E-mail: saeki-ys{at}igakuken.or.jp
Lab HP
【Key Words】Ubiquitin-proteasome system, Protein degradation, Proteostasis, Liquid-liquid phase separation, Mass spectrometry
The ubiquitin-proteasome system (UPS) is a crucial protein degradation system that regulates almost all cellular functions in eukaryotic cells. Since the maintenance of protein homeostasis is essential for human health, malfunction of the UPS causes various diseases including cancer, inflammation, and neurodegeneration. Therefore, the UPS regulators are attractive targets for drug discovery, and in fact development of proteasome inhibitors and proteolysis targeting chimeras (PROTACs) has been an important world-wide topic. However, a large part of molecular mechanism underlying the UPS remains elusive due to its complexity, and tools for accurate estimation of UPS function are limited. Our goal is to elucidate the fundamental mechanisms of ubiquitin signaling and proteasomal degradation to understand pathophysiology and to develop therapeutic strategies for UPS-related diseases.
1) Fundamental mechanisms of proteasomal degradation: Previously, it was simply thought that ubiquitylated proteins are directly recognized and degraded by the proteasome. However, we found that ubiquitin-selective chaperone p97 complexed with UFD1-NPL4 and the UBL-UBA proteins play a pivotal role in selection of proteasome substrate and delivery to the proteasome (Mol Cell 2017, Nat Commun 2018, 2019). We also found that branched ubiquitin chains enhance proteasomal degradation of several substrate proteins (PNAS 2018, Mol Cell 2021). Given that multiple proteasome-interacting proteins regulate proteasome activity, proteasomal degradation would be finely regulated at various levels. We are investigating high-order architecture of ubiquitin chains, substrate selectivity of the ubiquitin-binding proteins, and proteasome substrates using newly developed proteomics and chemical tools.
2) Biological significance of proteasome phase separation: Proteasomes are diffusely localized in the cytoplasm and nucleoplasm, but, recently, we have found that hyperosmotic stress induces the ubiquitin-dependent liquid-liquid phase separation (LLPS) of the proteasome, which forms proteolytic droplets in the nucleus (Nature 2020). As this LLPS could be associated with the ubiquitin-positive inclusion bodies observed in various neurodegenerative diseases, we are further investigating whether various stresses induce proteasome droplet.
3) Analysis of proteasome mutant mice: We have generated systemic proteasome mutant mice based on recently identified gene mutations in patients. By analyzing the phenotype of these mice, we aim to understand the pathophysiological mechanism of proteasome-related diseases, ‘proteasomopathy’.
1. Kaiho-Soma et al. Mol Cell 2021. TRIP12 promotes small-molecule-induced degradation through K29/K48-branched ubiquitin chains.
2. Yasuda, Tsuchiya, Kaiho, et al. Nature 2020. Stress- and ubiquitylation-dependent phase separation of the proteasome.
3. Sato, Tsuchiya, Nat Commun 2019. Structural insights into ubiquitin recognition and Ufd1 interaction of Npl4.
4. Tsuchiya, Burana, et al. Nat Commun 2018. Ub-ProT reveals global length and composition of protein ubiquitylation in cells.
5. Ohtake et al. PNAS 2018. K63 ubiquitylation triggers proteasomal degradation by seeding branched ubiquitin chains.
6. Tsuchiya et al. Mol Cell 2017. In vivo ubiquitin linkage-type analysis reveals that the Cdc48-Rad23/Dsk2 axis contributes to K48-linked chain specificity of the proteasome.