| Project/Area Number |
22K15112
|
|---|
| Research Category |
Grant-in-Aid for Early-Career Scientists
|
| Allocation Type | Multi-year Fund |
|---|
| Review Section |
Basic Section 44010:Cell biology-related
|
|---|
| Research Institution | Institute of Physical and Chemical Research |
|---|
Principal Investigator |
Li Xue 国立研究開発法人理化学研究所, 生命機能科学研究センター, 研究員 (50936507)
|
|---|
| Project Period (FY) |
2022-04-01 – 2024-03-31
|
| Project Status |
Completed (Fiscal Year 2023)
|
| Budget Amount *help |
¥4,550,000 (Direct Cost: ¥3,500,000、Indirect Cost: ¥1,050,000)
Fiscal Year 2023: ¥2,210,000 (Direct Cost: ¥1,700,000、Indirect Cost: ¥510,000)
Fiscal Year 2022: ¥2,340,000 (Direct Cost: ¥1,800,000、Indirect Cost: ¥540,000)
|
|---|
| Keywords | KIF5C / Kinesin 1 / Microtubule binding / Cortical neuron / Cortical Neuron / TIRF / STED Microscopy / kinesin KIF5C / axon microtubule / TurboID / STED |
|---|
| Outline of Research at the Start |
1) optimizations of culture cortical neuron cells from mice Embryonic mice at E16 will be performed.
2) Several KIF5C-TurboID dimer constructs will be purified, and KIF5C binding activity to MT will be measured which is critical in our study.
3) Identify KIF5C partners using TurboID. The purified KIF5C-TurboID dimer will be delivered into semi-intact neurons, where KIF5C binds to MT and TurboID labels proteins in proximity of KIF5C with biotin. Then the biotinylated proteins will be identified by mass spectrometry, and we are going to verify the hits and uncover the binding mechanism behind.
|
| Outline of Final Research Achievements |
In this study, we focus on the identification of molecular machinery in cells regulating or adjusting the binding affinity of KIF5C (Kinesin 1) to microtubules, of which the importance has been highlighted after the analysis of KIF5C mutants found in patients with severe malformations of cortical development and microcephaly. We made several KIF5C binding constructs to mimic selective binding and have established KIF5C-TurboID and KIF5C-APEX2 to label key protein players in semi-intact cortical neurons. We are now proceeding to hits identification using mass spectrometry analyses. We expect to reveal key regulators of KIF5C binding that could be potentially helpful for early diagnosis and treatment of the related brain diseases.
|
| Academic Significance and Societal Importance of the Research Achievements |
KIF5C変異体は、大脳皮質発達の重篤な奇形や小頭症の患者で発見され、その重要性が明らかになっている。本研究では、KIF5C（キネシン1）の微小管への結合親和性を制御する細胞内分子機構の同定を目指した。KIF5C結合の主要な制御因子が明らかになれば、関連する脳疾患の早期診断や治療に役立つ可能性がある。
|
