2019 Fiscal Year Research-status Report
The role of O-GlcNAcylation on Notch1 in the self-renewal of muscle satellite cells
| Project/Area Number |
19K16517
|
|---|
| Research Institution | Nagoya University |
|---|
Principal Investigator |
LO PEIWEN 名古屋大学, 医学系研究科, 研究機関研究員 (20822406)
|
|---|
| Project Period (FY) |
2019-04-01 – 2021-03-31
|
| Keywords | O-GlcNAcylation / Notch1 / satellite cell / regeneration / skeletal muscle |
|---|
| Outline of Annual Research Achievements |
Skeletal muscle has a great regeneration capacity that relies on its resident stem cells, satellite cells, but it declines with aging. Notch, a transmembrane receptor, can promote muscle regeneration via its regulation in quiescent satellite cells. Thus, we speculated extracellular O-GlcNAcylation on Notch, the important post-translational modification we found in the previous study, might regulate the regeneration capacity of skeletal muscle. Extracellular O-GlcNAcylation of Notch1 is catalyzed by EGF Domain Specific O-Linked N-Acetylglucosamine Transferase(Eogt). Thus, we verified whether Eogt-catalyzed O-GlcNAcylation would be related to muscle in the beginning. As a result, we found Eogt knockout affect muscle contractibility in the cap of cage hanging assay. In addition, we accomplished the genotyping of pTRE-FLAG-Notch1 transgenic mice, which are generated by using microinjection. These transgenic mice will be applied for inducible FLAG Notch1 transgenic mice. In our preliminary results, Eogt may be involved with the composition of the myofibres. We expect this study can contribute to the improvement of muscle rejuvenation for senior citizens and the therapy for muscular dystrophy patients with Notch1 defects.
|
| Current Status of Research Progress |
Current Status of Research Progress
3: Progress in research has been slightly delayed.
Reason
1. If Eogt knockout would affect muscle contractile?
Cage-cap hanging assay showed Eogt knockout reduced the time of cage-cap hanging obviously. According to the preliminary results, Eogt KO may affect the composition of muscle, augmenting the amount of collagen and altering the contribution of cross-section area in muscle. Subsequently, I will evaluate the expression of Eogt, Notch1 and Notch1-regulated gene in Eogt KO and wild-type mice.
2. If Notch1 glycosylation is altered between muscle satellite cells of young and aged mice?
Generation of FLAG-mNotch1 transgenic mice
The transgenic mice cross with wild-type have been evaluated their genotyping. Next, I will examine the expression of FLAG-Notch1 in the fibroblast of these transgenic mice to obtain premium FLAG-Notch1 inducible mice.
|
| Strategy for Future Research Activity |
1. To address if Eogt knockout(KO) would affect muscle contractile, I will measure the cross-section area of muscle in Eogt KO and wild-type mice. Also, I will evaluate if Eogt KO would extend the contribution of collagen. Subsequently, I will detect the expression level of Eogt, Notch1 and Notch1-regulated gene.
2. To address if O-GlcNAcylation Would Affect Muscle Regeneration? I will examine if the recovery rate of muscle will be changed in Eogt ko mice by using cardiotoxin injury assay.
3. To address if O-GlcNAcylaiton on Notch1 is altered between muscle satellite cells of young and aged mice, we will generate inducible FLAG-Notch1 transgenic mice. The capability of FLAG-Notch1 expression in pTRE-FLAG-Notch1 transgenic mice will be tested by using in vitro fibroblast examination.
|
