2021 Fiscal Year Research-status Report
The nature and roles of cellular metabolism and energy in the regulation of neuronal chain migration
| Project/Area Number |
20K06865
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| Research Institution | National Institute of Genetics |
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Principal Investigator |
ZHU YAN 国立遺伝学研究所, 遺伝形質研究系, 助教 (50464235)
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| Project Period (FY) |
2020-04-01 – 2023-03-31
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| Keywords | metabolic pathway / neuronal migration / bioenergetics / ATP sensor |
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| Outline of Annual Research Achievements |
This research has two objectives: (1) visualize and quantify the energy status of chain migrating neurons; (2) unravel the metabolic sources that fuel neuronal migration.
For objective (1), while I have established a system to express genetically encoded ATP sensors (ATeam) in vivo and in vitro, progress in FRET-based imaging of intracellular ATP is hindered by technical difficulties. For objective (2), I have generated a mouse knockout line with deficiency in a transcription factor known to promote OXPHOS metabolic pathway during development. Phenotypic analysis of the mutant line has shown defects in neuronal migration and/or their termination of migration.
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| Current Status of Research Progress |
Current Status of Research Progress
4: Progress in research has been delayed.
Reason
The progress of this study is delayed.
The FRET-based ATP sensor imaging is delayed due to a broken down of a laser line of our common confocal imaging system and the time it took to replace the laser.
Another reason that delayed this project is due to COVID-related family issues.
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| Strategy for Future Research Activity |
(1)I will perform further phenotype analysis of the knockout mice generated to understand the defects in detail, including morphological changes, dendritic pattern and axon projection. As the knockout mice die at birth, I will also employ CRISPR/CAS9 mediated gene editing to knockout the gene of interest via local in utero electroporation and observe the effect in postnatal samples.
(2)I will perform manipulation of metabolic pathways using pharmacological reagents and examine the consequences of such manipulations on chain neuronal migration using in vitro whole tissue culture system.
(3)I will introduce the fluorescent ATP sensors into the migrating neurons and image ATP levels in both fixed and living conditions.
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| Causes of Carryover |
Next year budget will be spent on (1) Hiring cost of a technician who will breed and maintain the transgenic mouse lines, and perform phenotypic analysis; (2) purchasing pharmacological reagents; (3) purchasing antibodies; (4) purchasing animals; (5) tissue culture reagents.
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