How do gene mutations lead to cancer? Myeloid leukemia is one of the best model systems to address this question, because hematopoiesis and myeloid differentiation are well understood and many mutations of genes implicated in myeloid leukemia evolution have been reported. Among them, the TET oncogene family member 2 (TET2) mis-sense mutations have been identified in several independent cohorts of patients with massive myeloproliferative disorders. However, the mechanism by which TET2 mutations cause these diseases is completely unknown. Importantly, TET1 catalyzes the conversion of 5-methylcytosine in DNA to 5-hydroxymethylcytosine, which hints that epigenetic regulation plays critical roles for leukemia development caused by TET2 mutations. The major goal of this project is to reveal how the TET2 mutations are involved in the pathogenesis of myeloid malignancies. To do this, an interdisciplinary strategy combining genomics, proteomics molecular biology and cell biology will be employed. The genome-wide approaches will be employed to profile the target genes of TET2 in hematopoietic stem cells (HSCs), with subsequent validation by traditional methods. TET2 protein complexes will be purified using proteomics approach to identify the proteins involved in TET function and regulation. The importance of TET2, its regulator proteins and their target genes on HSCs differentiation will be confirmed by cell biological analysis and murine model. This project will shed light on epigenetic regulation of TET2 proteins in hematopoiesis and contribute to our understanding of how myeloid cells acquire the malignancy through TET2 mutations.

Telomeres are critical for genome maintenance, as they prevent the natural chromosome ends from being recognized as damaged DNA. While telomere length decreases during the replication of normal cells, immortal cancer cells acquire the pathways that keep telomere length abnormally. Understanding the dynamics and differences of telomere regulation in normal and tumor cells is essential for comprehension of chromosome inheritance mechanism and the successful combating of cancer. Human telomeres associate with a number of factors that can influence chromosome integrity, such as the telomeric core complex and proteins involved in damage checkpoints, repair and recombination. Since cohesin, a ring-like complex that holds sister chromatids, is involved in checkpoints, repair and recombination pathways, it is tempting to hypothesize that cohesin acts in telomere maintenance in interaction with these pathways. To deepen our knowledge about telomere dynamics in both normal and cancer cells, I propose two specific aims: AIM 1 will focus on the novel role of cohesin in telomere maintenance, and AIM 2 is designated to explore telomere dynamics in cancer cells. I will examine the cell cycle dependent localization of cohesin at telomeres and the telomeric effects of cohesin suppression in AIM 1. Since SMC1, a cohesin subunit, is phosphorylated by damage checkpoint kinase, I will also investigate the importance of the phosphorylation in telomere dynamics. In AIM 2 I will study the dynamic binding of proteins and the replication pattern at telomeres in different types of immortal cells, with the goal to utilize the results to develop targeting strategies for different cancer cell types.

Mechanisms by which organ specific stem cells initiate differentiation (or cell cycle exit) remain largely unknown. In addition it is poorly understood, how cell cycle regulators post-translationally regulate organogenesis. I here propose to explore the link between the cell cycle and retinal development by examining the function and regulation of Id proteins (Inhibitor of differentiation) in retinal stem cells. By utilizing retina of medaka fish (Oryzias latipes), which is amenable to genetic and cell biological analyses, I will first determine the functions of Id in the proliferation of retinal stem cell. In addition, to complement limitations of the medaka retinal system with respect to biochemical analysis, I will be building on my previous expertise and will be using frog (Xenopus laevis) eggs to biochemically address the molecular regulation of Id. For this experiment, I will test two hypotheses concerning the interaction of Id with the APC/C, a ubiquitin ligase specifically degrading cell cycle regulators: (1) The APC/C destroys Id to allow differentiation of retinal stem cell, and (2) Id inhibits the APC/C to maintain stem cells with pluripotency. Finally, on the basis of the results from the Xenopus oocyte system, I will genetically test the models using the medaka fish retina. These results will allow developing the control of cellular differentiation by the cell cycle as new field in developmental biology, which will significantly impact also on other biological processes, such as diseases derived from genetic or developmental abnormalities.

It has been suggested that the large recognition diversity of cell surface proteins is required for complex neuronal wiring. Drosophila Down syndrome cell-adhesion molecule (Dscam) is a cell surface protein that belongs to immunoglobulin superfamily. Dscam can generate up to 38016 isoforms by alternative splicing and shows specific homophilic binding. Based on these features, Dscam is thought to be a very attractive molecule that could explain the mechanism of specific neuronal circuit formation. Dscam homophilic binding induces repulsion signaling and thereby contributes to dendritic self-avoidance and axon branching. However, the molecular mechanisms that translate adherent homophilic binding into repulsion are not understood. Here, I suggest to identify novel binding proteins of Dscam by biochemical method and to investigate their importance in vivo by genetic method. My host supervisor Dr. Schmucker has suggested that each Dscam isoform has non-redundant function during axon targeting of mechanosensory neurons. To examine this possibility, it is important to investigate expression pattern and localization of Dscam isoforms at protein level. To this end, I propose to use extracellular domain of Dscam isoform fused to a reporter protein such as protein phosphatase (AP). Next, I investigate whether intercellular Dscam-Dscam interactions could function as adhesion during axon/dendrite targeting and synapse development. To this end, I examine that whether same sets of isoforms are expressed at pre- and post-synaptic region and examine the role of Dscam in localization of neuronal transmitter receptor and synapse transmission.

This research project will provide fundamental new insights into the mechanisms and functions of neuronal network activity in the basal ganglia, brain areas critical for movement. The main goal is to elucidate how synchronized neuronal oscillations are either advantageous or counterproductive for the operation of these brain circuits during movement. Excessive synchronization of rhythmic neuronal activity in the basal ganglia is a critical functional change accompanying Parkinson’s disease (PD). Studies in unmedicated PD patients show that synchronized oscillations preferentially occur at ‘beta’ frequencies (15-30 Hz), and suggest that, by inappropriately coordinating neuronal activities, exaggerated beta oscillations may play ‘anti-movement’ roles in PD. To answer the key questions of where and how exaggerated beta oscillations emerge, I will exploit a rat model of PD that shows these oscillations. I will conduct large-scale electrophysiological recordings throughout the basal ganglia and partner circuits, thus addressing ‘Where’. By applying pharmacological manipulations, large-scale network recordings and intracellular recordings of single neurons in key circuit components, I will determine the inputs, outputs and intrinsic properties necessary for generating beta oscillations, thus addressing ‘How’. Finally, after defining the synchronized oscillations present during both normal and Parkinsonian movement, I will test whether inducing beta oscillations causes movement problems and whether targeted disruption of beta oscillations restores performance. Thus, I will also lay foundations for developing new therapies that better counteract these pathological rhythms in PD.

Helper and regulatory CD4+ T cells are major determinants and modulators of immune responses.
To date, well-defined helper CD4+ T cell subsets are Th1, Th2, Th17 and immunosuppressive regulatory T cells(Treg). These subsets can be characterized by the high expression of “lineage-specific” transcription factors (TFs), namely, T-bet/Runx3, GATA-3/c-Maf, RORgt, and FoxP3, in Th1, Th2, Th17 and Treg, respectively. However, it remains to be determined what signals control the expression of these lineage-specific TFs and other undefined key genes, and which genes are the targets of these TFs during T cell differentiation. In order to understand the dynamics of gene regulation during T cell development, I will combine mathematical modelling with genome-wide analysis of transcriptional regulation by microarray, and molecular and immunological experiments. With help of mathematical modelling, I will try to identify key target genes of the lineage-specific TFs with statistically ranked data. I will also use mathematical models of the “lineage-specific” TF interactions, most of which are in fact expressed by other T cell lineages, to identify the factors that determine T cell fate and assess the models by immunological experiments. Finally, I will predict signaling pathways that control the development of these T cell subsets by mathematical modelling and assess the prediction by molecular and immunological experiments including retroviral gene transduction and RNA interference. This will establish an iterative approach between mathematical modelling and wet experiments in this field.

The dentate gyrus of the hippocampus is one of the brain regions in which neurons are generated during adulthood. Increasing evidence supports the view that adult newborn granule neurons play an important role in hippocampus-dependent learning and memory. In recent years, it has been reported that immature new neurons approximately one month after their birth may make particularly significant contributions to memory formation. However, at what exact stage the young new neurons become important for memory formation, and the memory processes they mediate, have not been yet determined. Therefore it is necessary to identify the specific functions of young new neurons in memory formation in order to understand the mechanisms of newborn-neuron-based memory processing in the hippocampus. Identifying a specific function of young new neurons in learning and memory requires the establishment of new temporally and spatially controlled ablation techniques. Here we propose to develop a virus-based ablation system capable of specifically ablating young newborn neurons at different times during memory processing. Furthermore, we will evaluate the direct effect on memory formation resulting from virus-based specific elimination of young newborn neurons. Through these analyses, we aim to determine the specific function of young newborn neurons as a component of learning and memory. The achievement of the goals of this project will provide us with new insight into the issue of what kind of memory young newborn neurons are involved in, and give us an idea as to how newborn neurons compute memory information.

A major function of the circadian clock is to precisely coordinate responses of an organism to daily environmental stimuli. The molecular mechanisms underlying the integration of environmental stimuli and the clock are poorly understood in all organisms. My objective is to understand how environmental signals are integrated into the circadian clock and the function of specific genes involved in this integration, using Arabidopsis as a model system. My proposal is designed to provide insight into the function of EARLY FLOWERING3, a gene that modulates the effects of light into the clock. The specific aims of this proposal are to: 1. Characterize interactions between ELF3 and known clock associated proteins in vivo. Biochemical and genetic interactions between specific clock proteins that are co-expressed and interact in vitro will be analyzed. 2. Identify novel proteins that interact with ELF3. Yeast two-hybrid assays and affinity purification techniques coupled with mass spectrometry will be used to identify novel ELF3 interacting proteins. Identified candidates will be analyzed by biochemical and genetic approaches in vivo. 3. Identify suppressors of elf3 mutant phenotypes. A suppressor screen will be performed to identify genes that genetically interact with ELF3. Relevance: The circadian clock allows for the proper coordination of daily human behaviors, such as sleep/wake cycles. Research to elucidate the complex molecular mechanisms underlying the circadian clock in plants will complement similar analyses in other systems, ultimately leading to a better understanding of the circadian clock in humans, allowing for the treatment of circadian disorders.

Mediator is a large multi-protein complex critical for eukaryotic gene regulation. It is critical for gene regulation in all promoters since it functions as an interface between a wide variety of transcription factors bound to upstream regulatory sequences and RNA polymerase II. The mechanism by which Mediator controls gene regulation, however, remains obscure. Structural and functional studies on Mediator are essential to elucidate the Mediator mechanisms. However, the size (over 1 Mega Daltons), and the complexity (21 subunits) have prevented from such studies over the years. I will take a full advantage of the robust multi-protein expression system established in Takagi’s lab to obtain a large quantity of Mediator subcomplex, termed “Head module” (7 subunits, 223 kDa), and solve its structure by X-ray crystallography and EM. Additionally, I will apply genome-wide yeast two-hybrid array, and proteomics, and yeast genetics to identify the factors interacting with Head module. As Head module controls the action of Mediator, combined results will shed a light on how Mediator controls transcription.&#12288 Previously, I revealed the nuclear transport mechanism by solving the structure of human Transportin 1 (single protein) complexed with its substrates by X-ray crystallography. My proposed project will not only extend my own expertise of X-ray crystallography from solving the structure of a single protein to multi-protein complex, but also to learn new techniques such as protein complex expression, biochemical assays, yeast genetics and EM through collaboration to broaden my expertise. Thus, these will be indispensable for my career development.

The vascular system is essential for animal survival as it transports nutrients and oxygen throughout the body. Although several recent breakthroughs have enhanced our understanding of the genetic control of vasculogenesis and angiogenesis, the molecular mechanisms of vascular development still remain largely unknown. In general, forward genetic screen is one of the most powerful tools to reveal new clues of vascular development, and zebrafish is the best suited animal for such genetics screen among the animals with a closed circulatory system. Moreover, the transparency of zebrafish embryos and larvae enables one to observe the entire image of three-dimensionally expanded vascular system and to perform in vivo live imaging of blood vessel development. The accessibility of zebrafish allows various manipulations such as generation of chimeric fishes, pharmacological analysis. Therefore, we use zebrafish to reveal mechanisms of vascular development. Previous forward genetic screening identified the mutant with vascular defect, mts (Jin et al., 2007.) Through the analysis of this mutants, we propose to investigate the mechanisms of vascular development. Moreover, we will observe the individual steps of blood vessel development in detail, paying particular attention to individual cell behavior, cytoskeletal dynamics, and the process of endothelial cells polarization by in vivo time-lapse imaging. With the establishment of conditional knock down technology, we will attempt to reveal the mechanism of these steps. The analysis described above should give us further insights into blood vessel development.

The synapse is a specialized cell-cell contact site between neurons or neurons and muscles, and precision of synapse formation is critical in the formation of a neural circuit. While previous studies have identified the molecules that are sufficient for synaptogenesis, little is known about molecular mechanisms that inhibit synaptogenesis with non-target cells. To understand how neurons inhibit synaptogenesis between axons and non-target cells, I will utilize the DA9 neuron in C. elegans, in which the presynaptic terminal is restricted to the specific domain in the axon by the local activity of Wnt and its receptor Fz. As this signaling does not appear to belong to any known Wnt signaling cascade, I will identify the downstream components through characterizing mutants that have a similar phenotype to wnt mutants in terms of localization of synaptic markers. Concurrently, I will try to elucidate how Wnt signaling inhibits synaptogenesis, based on two hypotheses: 1. Wnt inhibits synapse formation 2. Wnt stimulates synapse elimination. Stage specific expression or inactivation of Wnt will reveal which step Wnt actually regulates. I also plan on examining the relationship between Wnt signaling and SCF complex (E3 ubiquitin ligase) in synapse elimination, as SCF complex is required for synapse elimination in other neurons. As Wnt signaling is essential in normal human development, and its deregulation causes various diseases including neurological disorder, understanding the molecular mechanisms of inhibition of synaptogenesis by the novel Wnt pathway will contribute to therapeutic treatment of these diseases, as well as in understanding the development of the human brain.

Sec4p is a Rab GTPase required for transport of secretory vesicles from the Golgi to the plasma membrane in yeast. Sec4p localizes on the secretory vesicle and controls vesicle transport, vesicle tethering and vesicle fusion with plasma membrane. In each case, activation of Sec4p by its guanine nucleotide exchange factor Sec2p is necessary for the function. Ypt32p, another Rab that regulates the export of cargo from Golgi to the plasma membrane, is required to recruit Sec2p to the secretory vesicle. Thus, Sec2p links two different Rabs and constitute a Rab-GEF cascade. However, the regulation mechanism of these Rabs in the transport pathway is unclear. The host laboratory has shown that Sec2p also bind to Sec15p, a component of the exocyst complex implicated in vesicle tethering at the plasma membrane. Since Ypt32p and Sec15p bind to Sec2p in a competitive manner, the mechanism that triggers the change in Sec2p to switch from binding Ypt32p to binding Sec15p would be important in regulating the Rab-GEF cascade. Moreover, Sec2p localization relies on the production of phosphatidylinositol 4-phosphate (PI4P) in the Golgi by the phosphatidylinositol 4-kinase Pik1p. In this proposal, I will focus on the regulatory mechanism of Sec2p in the exocytic pathway. First, I plan to establish the roles of Ypt32p and Sec15p in Sec2p localization by identification of their binding sites in Sec2p. Second, I will examine the domain in Sec2p that binds PI4P and other phosphoinositides. Finally, I will test the effects of altering the subcellular phosphoinositide distribution on Sec2p localization and functions, and examine the role of Pik1p on Sec2p localization to the secretory vesicle.

The posttranslational modification (methylation, acetylation, phosphorylation, ubiquitination) of histones at specific residues regulates gene expression in eukaryotic genomes. Ultimately, such epigenetic marks, singly or in combination, decide cellular state and differentiated cell types. The significance of the histone code depends not only on the modification site but also the type and extent of modification. There exist several examples of reader proteins that recognize specific histone marks, which in turn recruit other proteins that facilitate changes in chromatin state. However no methyl-arginine (Rme) reader protein has been identified, even though it has been known that several arginine residues can be methylated. My first goal is to identify as many Rme reader proteins as possible by pull-down assays with synthesized Rme-containing peptides mimicking distinct histone-tail contexts. In addition, I will characterize the recognition mechanism and the extent of modification (mono- and dimethylation) of these modified-arginine peptides to identify the best substrates by structural and energetic characterization. My second goal is to understand the mechanism of combinatorial recognition of two or more histone marks (one of which is a Rme mark) in the context of the nucleosome as the template. The proposed studies should elucidate how arginine methylation, either singly or in combination with other marks, controls chromatin structure and function. My long-term goal is to broaden my structural expertise to functional experiments and use the proposed combined approach to contribute to the exciting developments in epigenetic regulation of the genome.

To ensure their proper function throughout adult life, epithelial tissues balance cell growth, differentiation, and apoptosis. Homeostasis aberration leads to malfunction of tissues and severe diseases, including cancer. TGFb signaling has a significant role in the regulation of homeostasis. As an intriguing property, TGFb signaling is negative regulator of epithelial cell proliferation, whereas it can promote invasion and metastasis during the later stages of tumor progression. Surprisingly, epithelia lacking the essential receptor (TbRII) for TGFb signaling display morphologically normal tissue homeostasis by balancing hyperproliferation and elevated integrin-signaling with elevated apoptosis. However, this balance can be easily disrupted with additional homeostasis challenges. These mice display accelerated wound-repair and increased propensity to form squamous cell carcinomas (SQCCs). To elucidate the detailed molecular mechanisms underlying epithelial homeostasis downstream of TbRII, I will identify up/downregulated genes and posttranslational modifications arising from epidermal TGFb signaling and examine how these changes are affected in TbRII null vs WT epithelia, when subjected to wound-repair and SQCC progression. I will employ an ShRNA screening strategy to evaluate how these changes affect proliferation, apoptosis, cell migration and invasion in keratinocytes in vitro, and employ skin engraftment of ShRNA-infected keratinocytes in vivo and mouse genetics to determine the physiological relevance of my findings. These studies will advance our understanding of tissue homeostasis and may provide useful clinical methods for treatment of wound-healing and cancer.

Germinal centers (GCs) are important sites to establish humoral immunity. A cardinal event in GCs is antibody affinity maturation, in which B cells undergo somatic hypermutation of their antibody genes and selection for mutant clones that have acquired increased affinity for antigen. However, the precise mechanism by which selection occurs in GCs remains elusive. An important question is how high- and low-affinity B cells compete for binding antigen and make decisions on their fate. My proposed study is aimed at clarifying this point and eventually the selection mechanism of GC B cells. To accomplish this specific aim, I will study the behaviors of GC B cells in living lymph nodes with a special focus on the competitive aspect of B-cell selection using two-photon laser-scanning microscopy in combination with experimental skills that I have acquired in my previous research. In the first part of this proposal, I will visualize antigen itself utilizing quantum dots, nanometer-size fluorescent particles, to track antigen distribution within GCs and antigen capture by B cells. With this antigen-tracking system, I will analyze the changes in antigen-capturing behaviors of GC B cells and their interactions with follicular dendritic cells and helper T cells during affinity maturation. To directly examine the competition between B cells, I also propose to establish a system in which the fate of high- and low-affinity B cells can be tracked within the same GC. I believe this study would promote comprehensive understanding of the mechanism underlying antibody affinity maturation and shed light on the essence of humoral immunity.

Nuclear reprogramming is the functional conversion of the genome contained within a differentiated somatic cell to a state of developmental pluripotency, through factors that reside in the pluripotent stem cells, such as embryonic stem cells (ESCs). The aim of this research is to decipher the molecular mechanism of reprogramming, through understanding the roles of two factors recently discovered to play essential roles in reprogramming: Polycomb repressive complex 2 (PRC2) and Oct4. Nuclear reprogramming will be studied using cell fusion between mouse ES and human B cells. PRC2 methylates histone H3 and represses many differentiation-specific genes, but also appears to upregulate a set of genes in ESCs. I will modulate expression of target genes from both categories to test their roles in reprogramming. I will also test PRC2’s requirement in reprogramming cell types at different stages of differentiation, in an attempt to decipher the nature of PRC2’s role in reprogramming. Oct4 is a transcription factor that plays a central role in maintaining ESC identity. To identify Oct4 targets that play roles at different stages of reprogramming, I will immunoprecipitate Oct4 at different times after cell fusion. Identified targets will be tested for their essentiality in reprogramming by RNAi. The significance of this proposal is two fold. First, by advancing our understandings in the reprogramming process, we will further our understandings in ESC identity and the nature of each differentiation processes. Second, this work provides essential knowledge for generating patient-specific stem cells used for cell replacement therapy.

My recent research emphasis has shifted to the analysis of transcriptional control of gene expression by nearby regulatory elements and to a recognition of the role of long-range interactions and three-dimensional organization of chromatin within the nucleus. Insulators are now known as a major endogenous mechanism for epigenetic control of gene expression. Although many recent studies have been devoted to the identification and characterization of the CTCF-dependent insulators, there is still little information and mechanistic understanding of insulators distinct from CTCF. Arylsulfatase insulator (ArsI) is distinct from the CTCF family of insulators and is known to work in various animals from humans to plants. Unlike CTCF-dependent insulators, the ArsI sequence does not have CTCF-binding sites and can work independent of nuclear matrix. ArsI insulator is also found well conserved in sea urchins and there were over 100 putative sites for this fragment in the genome, implying the significance of this DNA fragment. Using biochemical and computational approaches, I would like to identify the proteins that interact with ArsI and reveal the mechanisms of ArsI regulation in vivo. I also expect the study on ArsI mechanisms may lead us to find novel mechanisms of gene regulations, chromatin modifications that are conserved in all organisms.

The random generation of diversity is the feature of immune system, which is essential to counteract numerous pathogens. However, it also allows immune system to attack self tissues and produce autoimmune diseases. Therefore self tolerance, the inability to make an immune response to self, is crucial to avoid autoimmune disease. For T lymphocytes, potentially auto-reactive cells are eliminated during their differentiation in thymus through encountering the autoantigens which are ectopically expressed in thumus. The AIRE (autoimmune regulator) is a key molecule to promote transcription of many ectopic genes in thymus and a critical element for self tolerance. Though it has structural features of a transcription factor, it is difficult to be reconciled with classical site-specific transcriptional factors, because it impacts on extensive transcriptions. To reveal the precise mechanism for transcriptional regulation by aire, we propose to identify the aire associating proteins by the yeast two-hybrid screening method with Ras Recruitment System. In addition, to elucidate the DNA-binding specificity of aire, we will perform high throughput sequencing-by-synthesis approach, ‘picotiter plate pyrosequencing’, following immunoprecipitating the Aire-DNA complex with anti-Aire antibody. The binding sites on genomic DNA will be examined by analyzing the obtained sequences through the database of mouse genome sequences. All these findings will contribute to reveal the molecular mechanisms for transcriptional regulation by AIRE and profound understanding of self tolerance, including the organ-specificities of autoimmune diseases.

The study of axonal growth is a central area of research in neurobiology. However, the cell-intrinsic mechanisms that regulate axonal morphogenesis remain largely to be discovered. The laboratory of Dr. Azad Bonni discovered that the ubiquitin ligase Cdh1-anaphase promoting complex (Cdh1-APC) controls axonal growth and patterning in the mammalian brain. In more recent studies, they identified the transcriptional corepressor SnoN and Id2 as key downstream targets of neuronal Cdh1-APC that promotes axonal growth. The focus of the proposal is to elucidate the mechanisms that regulate the Cdh1-APC-SnoN/Id2 cell-intrinsic pathway in postmitotic neurons and to determine how this pathway in turn controls axonal growth. To characterize the mechanisms that regulate Cdh1-APC in neurons, I will investigate the role of phosphorylation of Cdh1 and Cdh1-interacting proteins in axonal growth. To reveal the mechanisms by which SnoN and Id2 promotes axonal growth, I will determine transcriptional target genes regulated by SnoN and/or Id2, and reveal how these genes regulate the axonal growth. In addition, I would like to test the hypothesis that SnoN and/or Id2 might regulate the expression of microRNAs in neurons, because expression of several microRNAs has been recently shown to be regulated by transcriptional regulators and increasing evidence suggests that microRNAs regulate neuronal morphology. The postdoctoral research will be a great opportunity for me to learn knowledge and techniques of neuronal cell biology, signal transduction, and transcription regulation. HFSP fellowship will promote my professional growth through interdisciplinary research.

Our experience of time is central to our lives. Simply shaking hands means two people have to coordinate their hands to be in the same place at the same time operating clutch and accelerator in a car requires fine timing (remember how hard that was to learn?) and even social situations require timing information - to track someone walking across a room, to interject in conversation and to decide when to leave. These simple acts we take for granted require the use of timing from milliseconds to minutes and hours. Yet the scientific investigation of temporal experience is in its infancy and lags behind our understanding of other aspects of our senses. During my HFSP fellowship I will examine the brain mechanisms of time perception using state of the art techniques of human brain stimulation (TMS) and electrophysiological recording (EEG & fMRI). By stimulating brain areas involved in time perception I will explore the temporal dynamics of early and late brain responses to temporal information and will distinguish between conscious and unconscious processes involved in timing. Studies of vision and audition have shown that perceived time and relative timing of multiple events can be distorted (e.g., flash-lag effect, chronostasis etc.). I will apply TMS to low level visual areas as well as higher cognitive areas associated with attention and memory (e.g. inferior parietal cortex) to explore the extent to which these regions of the brain contribute to those illusory time percepts. The work experiment aims to uncover specific mechanisms associated with temporal perception, and will reveal fundamental characteristics of causal interactions between different cortical areas.