| Project/Area Number |
23KF0035
|
|---|
| Research Category |
Grant-in-Aid for JSPS Fellows
|
| Allocation Type | Multi-year Fund |
|---|
| Section | 外国 |
|---|
| Review Section |
Basic Section 54040:Metabolism and endocrinology-related
|
|---|
| Research Institution | Kanazawa University |
|---|
Principal Investigator |
篁 俊成 金沢大学, 医学系, 教授 (00324111)
|
|---|
| Co-Investigator(Kenkyū-buntansha) |
HEIN KO OO 金沢大学, 医学系, 外国人特別研究員
|
|---|
| Project Period (FY) |
2023-04-25 – 2025-03-31
|
| Project Status |
Granted (Fiscal Year 2023)
|
| Budget Amount *help |
¥2,400,000 (Direct Cost: ¥2,400,000)
Fiscal Year 2024: ¥1,200,000 (Direct Cost: ¥1,200,000)
Fiscal Year 2023: ¥1,200,000 (Direct Cost: ¥1,200,000)
|
|---|
| Keywords | Cysteine redoxome / Selenoprotein P / Reductive stress |
|---|
| Outline of Research at the Start |
The target proteins of Selenop mRNA and SeP protein will be comprehensively searched in brown and white adipose tissues. The functional and molecular mechanism of candidate proteins will be further analyzed to discover the novel targets of Selenop mRNA and SeP protein, which may be the potential therapeutic targets for SeP-mediated metabolic effects.
|
| Outline of Annual Research Achievements |
For the selenoprotein P (SeP) mediated reductive stress target, we have identified more than 1500 significant cysteine residues that are oxidized or reduced under acute cold exposure in the brown adipose tissue (BAT) of WT and Selenop-knockout mice. Among them, 30 cysteine residues are reversible reactive cysteines. We are now focusing on the reactive cysteine of mic19, a member of MICOS, and evaluating its importance using in-silico and functional analysis.
For mRNA-binding proteins, we identified 17 proteins that specifically bind to Selenop mRNA by using the RiboTrap assay. One potential target is adck1 which is also involved in the assembly of MICOS. We have created Selenop mRNA and untranslated UUG-Selenop mRNA expression vector for further confirmation and functional analysis.
|
| Current Status of Research Progress |
Current Status of Research Progress
3: Progress in research has been slightly delayed.
Reason
Currently, we only finished the comprehensive search for the target molecules in BAT samples because of the congestion in the LC-MS analysis workflow of our collaborator.
To express the various mutant variants and mRNA expression vectors in our in vitro assays, we are also trying to establish the immortalized mouse brown adipocyte cell line from the stromal vascular fraction cells of the BAT by using the SV40 vector. For in vivo analysis, we have also started the full-Selenop-mRNA-knockout mice, which are being crossbred with WT mice to obtain a higher genetic purity.
|
| Strategy for Future Research Activity |
We will confirm the functional involvement of reactive cysteine of mic19 by expressing cysteine to alanine mutated variant in immortalized mouse brown adipocytes and check the mitochondria cristae formation using electron microscopy. Next, we will evaluate the regulatory mechanism of the target cysteine redox status and their involvement for the mitochondrial dynamics and functions by using in vitro and in vivo models.
We will also confirm the role of Selenop mRNA in mitochondrial functions by overexpressing the untranslated mRNA to the brown adipocytes. We will further confirm the binding site of mRNA and target proteins by using various mutant models of mRNA.
|
