2014 Fiscal Year Annual Research Report
Small Bioactive Molecules for Orchestrating Embryonic Stem Cell Differentiation
Project/Area Number |
25870365
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Research Institution | Kyoto University |
Principal Investigator |
PERRON AMELIE 京都大学, 化学研究所, 講師 (40525862)
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Project Period (FY) |
2013-04-01 – 2015-03-31
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Keywords | SAR Studies / Oscillations / Gro/TLE / Hes1 / Regenerative Medicine / Embryonic / Stem Cells / Hypoxia |
Outline of Annual Research Achievements |
a) Structure-Affinity Relationship (SAR) Studies: 176 derivatives were synthetized and tested to optimize D8C, a hit compound we have isolated as a potential Hes1 modulator. JI051 showed a 10-fold improvement as compared to D8C. b) Effect of JI051 on Oscillations in Presomitic Mesoderm (PSM): PSM preparations expressing luciferase under the control of Hes7 showed a decrease in oscillation number upon treatment with JI051, suggesting that JI051 influences ultradian oscillations in Notch signaling pathway (collaboration with Prof. Kageyama). c) Target Validation Procedure: A photoaffinity probe was synthetized for labeling JI051 target. Although pull-down assay revealed a band corresponding to Gro/TLE, competitive displacement with 10-fold excess of JI051 was partially successful, suggesting nonspecific binding. Gro/TLE was then solely excluded as a JI051 target using recombinant Gro/TLE protein, indicating that JI051 may bind to another target. d) Comprehensive Kinase Profile: 211 kinases were tested in order to investigate if kinases such as CK2, a serine/threonine kinase involved in Gro/TLE corepressor activity, could be directly modulated by JI051. The results did not show inhibition or stimulation higher than 50%. e) Effect of JI051 on Gene Expression: The effect of JI051 on Hes1 mRNA revealed an increase in Hes1 mRNA levels as compared to cells treated with DMSO. In order to identify potential targets involved in JI051-mediated Hes1 modulation, microarray analysis was carried out. The gene analysis revealed an up-regulation of several genes involved in hypoxia.
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Research Products
(2 results)