Protein kinase N controls a lysosomal lipid switch to facilitate nutrient signalling via mTORC1

Area of Science:

• Cellular signaling
• Molecular biology
• Biochemistry

Background:

• Mechanistic target of rapamycin (mTOR) kinase, functioning in mTORC1 and mTORC2 complexes, is crucial for cell growth and nutrient sensing.
• Dysregulation of mTORC1 is linked to various diseases, and its activity is modulated by phosphoinositide lipids.
• The regulation of PI3KC2-β localization and activity by mitogens, and its role in mTORC1 signaling, remains largely unknown.

Purpose of the Study:

• To elucidate the mechanism by which protein kinase N (PKN) influences mTORC1 signaling.
• To investigate the role of PKN in regulating PI3KC2-β activity and its impact on nutrient signaling downstream of mTORC2.

Main Methods:

• Investigated the interaction between PKN and PI3KC2-β.
• Utilized phosphoinositide lipid analysis to assess PI3KC2-β activity.
• Employed genetic manipulation (loss of PKN2) and site-directed mutagenesis to study the phosphorylation of PI3KC2-β.

Main Results:

• Demonstrated that PKN facilitates mTORC1 signaling by repressing PI3KC2-β-mediated synthesis of phosphatidylinositol-3,4-bisphosphate.
• Showed that active PKN2 phosphorylates PI3KC2-β, promoting its association with inhibitory 14-3-3 proteins.
• Confirmed that loss of PKN2 or disruption of PI3KC2-β phosphorylation impairs nutrient signaling via mTORC1.

Conclusions:

• Uncovered a novel mechanism where mTORC2-activated PKN2 phosphorylates PI3KC2-β, thereby repressing its lipid kinase activity.
• This regulation couples mTORC2 signaling to the control of mTORC1-mediated nutrient responses through local lipid signaling.
• Provides a new understanding of how cellular nutrient signaling pathways are interconnected and regulated.