Inducible raptor and rictor knockout mouse embryonic fibroblasts

Area of Science:

• Cellular Biology
• Molecular Biology
• Biochemistry

Background:

• The mammalian Target of Rapamycin (mTOR) kinase is crucial for cell growth and metabolism, operating in two distinct complexes: mTORC1 and mTORC2.
• Rapamycin, a common drug, partially inhibits mTORC1 but not mTORC2, limiting research into mTOR signaling.
• A complete understanding of mTOR function requires methods beyond pharmacological inhibition, particularly for mTORC2.

Purpose of the Study:

• To generate novel research tools for dissecting the individual functions of mTORC1 and mTORC2.
• To overcome the limitations of current pharmacological inhibitors for studying mTOR signaling in vivo.
• To enable precise genetic inhibition of mTORC1 and mTORC2 components.

Main Methods:

• Utilized the Cre/LoxP system to create inducible knockout mouse embryonic fibroblasts (MEFs).
• Generated MEFs deficient for raptor (mTORC1 component) and rictor (mTORC2 component) via inducible knockout.
• Confirmed knockout efficiency by assessing raptor or rictor expression levels post-induction.

Main Results:

• Successfully generated inducible knockout MEFs for raptor (iRapKO) and rictor (iRicKO).
• Induction of knockout led to the specific elimination of raptor or rictor expression.
• Demonstrated impaired signaling in the corresponding mTOR complex upon knockout induction.

Conclusions:

• The developed iRapKO and iRicKO MEFs are valuable tools for studying mTORC1 and mTORC2 signaling independently.
• These genetic tools allow for a more comprehensive investigation of mTOR complex functions in biological processes.
• Facilitates deeper understanding of mTOR pathway dysregulation in diseases like cancer.