UC San Diego

Sestrin is a family of proteins known to regulate cell growth by TOR signaling modulation. While overexpression of dSesn in the eye only caused a slight reduction in size, coexpression of dSesn with TOR activators caused a reduction in size compared to single expression of TOR activators in the eye. Immunoblots indicate that TOR signaling is reduced with the expression of dSesn, indicated by reduced phosphorylation of TOR targets, 4E-BP and S6K. The dSesn-/- organisms showed increased lipid accumulation compared to wildtype, which was suppressed by AICAR and rapamycin feeding or expression of wildtype dSesn. Immunoblots of larvae fat bodies indicate that dSesn-/- organisms have decreased AMPK phosphorylation and increased S6K and 4E-BP phosphorylation. Cardiac function of dSesn-/- organism shows reduced heart rate, increased arrhythmia and increased diameter, which were rescued by rapamycin and AICAR feeding. Administration of vitamin E, an antioxidant, only rescued arrhythmia. Hand-GAL4 driver was used to express TOR downstream components specifically at the heart. Overexpression of an active form of 4E-BP suppressed translation to decrease heart diameter and rescue dSesn-/- heart rate, while overexpression of Atg1 resulted in aggravating arrhythmicity. NOVA screening of dSesn and mSesn1 may provide insights into understanding the structural basis of Sestrins. NOVA screening on 50,000 (dSesn) and 35,000 (mSesn1) flies in eyes and wings was performed, however, failed to isolate novel mutations that affect Sestrin function. Alternative genetic strategies, such as suppression of synthetic lethality by strong expression of Sestrins, may be used to improve sensitivity of the NOVA screening