| 研究課題/領域番号 |
18K15531
|
|---|
| 研究機関 | 国立研究開発法人理化学研究所 |
|---|
研究代表者 |
BAGGE EMILIE 国立研究開発法人理化学研究所, 脳神経科学研究センター, 研究員 (30813795)
|
|---|
| 研究期間 (年度) |
2018-04-01 – 2020-03-31
|
| キーワード | bipolar disorder / mitochondria / mtdna / mutation / sequencing |
|---|
| 研究実績の概要 |
During the first year of this Kakenhi grant, I have mapped the mtDNA mutation spectrum in six discrete brain regions from aged mice using a model of bipolar disorder (the Polg1(D181A) mutant mouse) and controls. To do this I have implemented and optimized a method for enrichment of mtDNA from very limited brain tissue samples. These samples have been sequenced using next-generation sequencing. I find that mutations are accumulated in a heterogeneous way that is largely unrelated to the expression of the transgene. Furthermore, I find that the mutations spectrum is highly diverse and includes both single nucleotide changes, short and long range deletions, a surprising duplication in a specific sequence of the mtDNA and a very restricted inversion. Furthermore, these observations are brain region-specific, underlying that mitochondrial function likely differs across brain regions. We are currently preparing the manuscript for publication of these results.
In addition to this, great progress have been made in the other aspects of this project in terms of optimization of methods for the analysis of very limited tissue samples for analysis of global gene expression changes and chromatin accessibility.
|
| 現在までの達成度 (区分) |
現在までの達成度 (区分)
2: おおむね順調に進展している
理由
The first part of the project, mtDNA mutation mapping, has progressed as planned and a paper is currently being prepared. For RNA sequencing, it has been necessary to revise the method, as we were not able to generate intact neurons from the adult brain without disturbing the surface proteins. Instead, I have implemented laser capture microdissection which is working very well. Soon these samples will be sequenced. From this we aim to map specific gene expression changes in dysfunctional neurons. For ATAC-sequencing, libraries have been prepared using FACS sorted neuronal nuclei, and are ready for sequencing. This is also a change from the initial plan. However, I feel that the changes in methodology were necessary in order to progress this research project in a timely as well as reproducible manner.
|
| 今後の研究の推進方策 |
Slight changes have been made to some of the methodologies as explained above. However, the overall aim has not changed and we still aim to complete sequencing analysis within the time frame of this project. Shortly, RNA sequencing libraries will be completed. At that time, both RNA-sequencing samples and ATAC-sequencing samples will be analysed using HiSeq. We aim to integrate these data sets to gain new knowledge of the gene expression changes and regulation in our bipolar disorder model.
|
