Abstract 446: Transcription Factor EB, Not mTOR is a Therapeutic Target for Rag GTPase Protein C S75Y-based Cardiomyopathy

• Published in: Circulation research Vol. 123; no. Suppl_1
• Main Authors: Kim, Maengjo, Qi, Qiu, Ma, Xiao, Xu, Xiaolei
• Format: Journal Article
• Language: English
• Published: 03-08-2018
• ISSN: 0009-7330; 1524-4571
• DOI: 10.1161/res.123.suppl_1.446

Abstract only
Rationale:
We recently reported the association of S75Y, a point mutation in the Rag GTPase protein C (
RRAGC
) gene, with pediatric DCM. Because RRAGC directly binds and regulates mTOR and the S75Y mutation results in abnormally activated mTOR, these in vitro data prompts that the mTOR pathway could be harnessed for therapeutic benefits.
Methods:
We aimed to integrate
in vitro
neonatal rat ventricle cardiomyocytes (NRVCM) and
in vivo
zebrafish system to elucidate underling mechanism of
RRAGC
-S75Y-based cardiomyopathy, and to seek targeted therapeutic strategies.
Results:
Overexpression of RRAGC with S75Y mutation, but not RRAGC WT, significantly increased cell size of NRVCM, suggesting the establishment of a cellular model of
RRAGC
- S75Y-based cardiomyopathy. At the molecular level, overexpression of RRAGC S75Y mutant increased activity of mammalian target of rapamycin (mTOR) and decreased activity of transcription factor EB (tfeb), a direct downstream target of mTOR. Intriguingly, treatment with rapamycin, a mTOR inhibitor, did not affect cellular hypertrophy, whereas adenovirus-mediated overexpression of TFEB was able to rescue cellular hypertrophy induced by RRAGC S75Y mutation. Consistently, treatment with trehalose, an mTOR-independent tfeb activator, rescued cellular hypertrophy induced by
RRAGC
-S75Y. To confirm the
in vitro
results, we generated a knock-in (KI) zebrafish line harboring the S75Y corresponding point mutation in
rragc
via the TALEN (Transcription Activator-Like Effector Nucleases) technology. The cardiac function of
rragc
KI fish was determined by high frequency echocardiography, and the KI fish showed a significant decrease in fractional shortening compared with control littermates.
Conclusion:
Both cellular and
in vivo
models of
RRAGC
-S75Y-based cardiomyopathy have been generated. Studies in the cellular model suggested that tfeb, but not mTOR, is a therapeutic target for this particular type of cardiomyopathy. We will report our progress in validating this conclusion in our
in vivo
model, and will discuss potential mechanisms.