Mechanisms of mTORC1 activation by RHEB and inhibition by PRAS40

Area of Science:

• Biochemistry
• Structural Biology
• Cell Biology

Background:

• The mechanistic target of rapamycin complex 1 (mTORC1) is a crucial regulator of cell growth and metabolism.
• mTORC1 activity is modulated by nutrients, energy status, and growth factors.
• Key components include the mTOR kinase, RAPTOR, RHEB (activator), and PRAS40 (inhibitor).

Purpose of the Study:

• To elucidate the structural mechanisms underlying mTORC1 activation and regulation.
• To investigate the role of RHEB in allosterically activating mTORC1 kinase activity.
• To define the structural basis for substrate recognition and inhibition by PRAS40.

Main Methods:

• Cryo-electron microscopy (cryo-EM) to determine the structure of mTORC1 and activated RHEB-mTORC1.
• X-ray crystallography to resolve structures of RAPTOR-TOS motif complexes, mTOR FRB-substrate complexes, and mTOR-PRAS40 complexes.
• Biochemical assays to assess kinase activity and the impact of mutations.

Main Results:

• The cryo-EM structure of RHEB-mTORC1 reveals RHEB binding induces global conformational changes that allosterically activate the mTOR kinase active site.
• Cancer-associated mutations were mapped to regions maintaining the inactive state, suggesting they mimic RHEB-mediated activation.
• Crystal structures defined TOS motif recognition by RAPTOR and identified a second substrate recruitment mechanism via the mTOR FRB domain.
• PRAS40 was shown to inhibit both substrate recruitment sites.

Conclusions:

• Structural and biochemical data explain how mTORC1 selects substrates and how its kinase activity is regulated.
• The findings provide mechanistic insights into mTORC1 activation by RHEB and cancer-associated mutations.
• These discoveries offer a foundation for understanding mTORC1 dysregulation in diseases like cancer.