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Name
Email
Telephone
Professor Tsukita, Sachiko., Ph.D. mail +81-6-6879-3320
Assistant Prof. Matsui, Takeshi, Ph.D. mail-2 +81-6-6879-3322
Assistant Prof. Tamura, Atsushi, Ph.D. mail-3 +81-6-6879-3322

FAX +81-6-6878-3329
Postal Mail Address Graduate School of Frontier Biosciences, /Department of Pathology, Graduate School of Medicine, Osaka University, 2-2 Yamadaoka, Suita, Osaka 565-0871 Japan


The major focus of our research is the functional analysis of ‘field’ created by epithelial cell sheet formation in the body and cell.  Individual epithelial cells mechanically adhere to each other strongly to create epithelial cell sheets that cover the body and organ surfaces, and to produce compartments in the body.  For epithelial cell sheets to play the role of functional septa, epithelial cells form a clear polarity, and the cell membrane differentiates into the inner apical, junctional, lateral, and basal membranes.  The formation of the cell-cell junction (intercellular adhesion apparatus) that surrounds epithelial cells like a ‘headband’ creates apparent differences between the cell membranes present inside and outside of the inner body.  We are currently investigating the principle behind the construction of ‘field’ in the body and analyzing its functions,  while performing molecular- and individual-level analyses.

1 Analysis of apical membrane

There are specialized structures, such as microvilli and primary cilia, in the apical membranes of cells, and we have identified ezrin/radixin/moesin (ERM) proteins and Odf2 as essential proteins for the construction of these structures.  ERM proteins are important for the construction of microvilli, and they also integrate functional membrane proteins such as transporters and channels, being involved in various physiological and pathological conditions.  How are ERM proteins related to intracellular signals?  What is the role of the primary cilium possessed by most somatic cells, based on the findings that when cells lose Odf2, primary cilia disappears?  We are currently performing mainly individual-level analyses to answer these questions.

fig-1

Fig.1 Schematic representation of ERM family and activation signaling of ERM proteins (left).  Radixin knockout mice showed progressive degeneration of cochlear stereocilia (right). 


2 Functional Analysis of ZO-1

ZO-1 is a lining protein of intercellular junctions identified in mouse liver bile capillary fractions by Tsukita Laboratory.  We have recently succeeded in the preparation of epithelial cell lines without ZO-1 and ZO-2, and revealed that ZO-1 and ZO-2 form scaffolds for TJ formation.

fig-2

 

 

 

 

 

Fig. 2 It was demonstrated that ZO-1 dimers not only polymerized claudin directly below the cell membrane, but also determined the accurate localization of TJ strands.


3 Functional Analysis of Claudin Family

Claudins with molecular masses of ~23 kD bear four transmembrane domains and are essential for TJ strand formation.  In the body, 24 types of claudin form TJ in various combinations.  How is this variety of claudin involved in the diversity of intracellular barrier function?  A recently identified example is an involvement in epithelial cell proliferation.

fig-3

Fig. 3 Analysis of knockout mice suggested that a certain type of claudin is involved in the control of cell proliferation.


4 Regulation of epithelial cell proliferation

Epithelial cells, attracting the interest of our laboratory, are considered to be the origin of 90% of malignant neoplasms.  Elucidation of the epithelial cell-specific proliferation mechanism is awaited in this regard.  It has recently been revealed that the analyses of ERM and junctional proteins performed by our laboratory are linked to the analysis of epithelial cell proliferation at the molecular level.

fig-4

Fig. 4 Actin-binding protein was found to be localized in the intercellular junction, whose expression varies with cell density.


5 Analysis of cell-cell junction (intercellular adhesion apparatus)

Multicellular organisms cannot be establised without intercellular adhesion.  Our laboratory has performed the separation, identification, and analysis of component proteins of junctions specialized for intercellular adhesion by adding various modifications to classic biochemical methods, and proceeded with studies by employing unique approaches.  We developed a preparation method of fractions composing adherens and tight junctions, and analyzed many major proteins.  We recently incorporated a further modified analytical technique to analyze unknown junctional component proteins, aiming at a comprehensive understanding of the junction.  We also recently became able to approach the principle of construction of tight junctions by claudin using cells with supressed expression of ZO-1/2.

fig-5

Fig. 5 The bile capillary fraction was applied to two-dimensional electrophoresis, and spots concentrated in the junction were subjected to mass spectrometric analysis.  Many novel membrane proteins were identified, and their functions are being analyzed by the RNAi method and through the generation of knockout mice. 

‘Do you enjoy your research?’  This question cannot be asked many times in advanced research laboratories.  Researchers are too busy, attaching greater importance to efficiency.  Our laboratory sets ‘enjoying research’ as a basis.  Individual researchers aim to enjoy research based on individual consciousness and responsibilities.  Enjoying, of course, does not mean playing.  Problems and distress because of difficulties with research are also included in the concept of enjoyment.  Individually doing what you want is not the way.  Organized connections among individual researchers form a ‘field’, and the gathering of several persons, working together and not singly, may lead to much more enjoyable and significant studies.  Don’t you think that this is the real attraction of research?
## Application for graduate students is accepted.


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