2013 Fiscal Year Annual Research Report
多能性幹細胞の分化過程における染色体の機能的構造変化
| Project/Area Number |
12F02079
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| Research Institution | Kyoto University |
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Principal Investigator |
中辻 憲夫 京都大学, 物質―細胞統合システム拠点, 教授
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| Co-Investigator(Kenkyū-buntansha) |
KAFER Georgia 京都大学, 物質―細胞統合システム拠点, 外国人特別研究員
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| Keywords | Stem cells / Histones / Acetylation / Chromatin / Trophoblast / Differentiation / Pluripotency / Immunofluorescence |
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| Research Abstract |
My research project looks at cellular events involved in the differentiation of human embryonic stem (hES) cells into a specialized type of cell called a trophoblast cell. The major part of the project is centered on understanding how chromatin (the architecture of the nucleus) changes when hES cells loose their pluripotency and differentiate into the specialized trophoblast cells. I specifically study proteins called 'histones' which are the basic unit of chromatin. I am particularly interested in a type of histone protein called 'histone 2A variant Z' (H2AZ). I am interested in H2AZ because it can have different effects on the chromatin architecture when by becoming modified. These modifications primarily concern the addition of small molecules called "acetylation" or "methylation". To study the changes in the modification of H2AZ I use a technique called immunofluorencese (to fluorescently label the proteins or RNA molecules I want to study) I then image the cell nucleus with a very sensitive microscope and then perform a specialized type of image processing, and analysis that was writen in my laboratory on the captured image. Using these techniques, I have found that status of H2AZ acetylation in the stem cell changes very quickly (in as little as 12 hours) following treatment to induce differentiation. I have studied the protein complexes that both acetylate and de-acetylate H2AZ under hES differentiation and found that de-acetylation of H2AZ is critical for differentiation to proceed. Current work on this part of the project aims to further investigate the reason for the dynamic changes in the modification status of H2AZ during hES differentiation and determine which histone deacetylase complex (HDAC) is responsible.
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| Current Status of Research Progress |
Current Status of Research Progress
2: Research has progressed on the whole more than it was originally planned.
Reason
I have successfully performed all of the experiments I had hoped to by this point in time except for one very critical experiment called "ChIP-Sequencing". However, I plan to complete this experiment in the very near future. This is going to be an important experiment as it will give me important information that will help me to decide how significant the changes in H2AZ acetylation really are for hES biology and the progression of differentiation.
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| Strategy for Future Research Activity |
A major priority is the completion of the ChIP-Sequencing experiment as outlined in 11. The results of this experiment may give us new ideas regarding the role of H2AZ in stem cell biology, which we can then pursue. Aside from this (and related) experiments, my future experimental plan is to specifically target the protein complex HDAC2 which I hypothesize is primarily responsible for the de-aetylation of H2AZ during early differentiation. This work will be very novel, as the complex that de-aetylates H2AZ is not yet reported. This work will involve using chemical inhibitors, performing genetic modification experiments to knockdown the protein of interest together with immunofluorescence to study the effect of the inhibition/knockdown.
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Research Products
(2 results)
