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Association of CAD, a multifunctional protein involved in pyrimidine synthesis, with mLST8, a component of the mTOR

Akio Nakashima1, Ippei Kawanishi, Sumiko Eguchi

  • 1Biosignal Research Center, Kobe University, Kobe, 657-8501, Japan.

Journal of Biomedical Science
|April 19, 2013
PubMed
Abstract

Insights

The mechanistic target of rapamycin (mTOR) pathway regulates cell growth. This study reveals that mLST8 links mTOR to CAD, a key enzyme in pyrimidine synthesis, controlling its activity.

Area of Science:

  • Molecular Biology
  • Cellular Signaling

Background:

  • The mechanistic target of rapamycin (mTOR) is a crucial kinase regulating cell growth, proliferation, and survival.
  • Carbamoyl-phosphate synthetase 2, aspartate transcarbamylase, and dihydroorotase (CAD) is vital for de novo pyrimidine synthesis and is regulated by various kinases, but its connection to mTOR remained unknown.

Purpose of the Study:

  • To investigate the potential relationship between the mTOR pathway and CAD.
  • To elucidate the role of mLST8 in mediating interactions between mTOR and CAD.

Main Methods:

  • Co-immunoprecipitation and immunoblot analysis to confirm protein-protein interactions between CAD and mLST8.
  • Transfection of HEK293 cells with FLAG-mLST8 and myc-CAD vectors.
  • Analysis of mutant constructs to identify binding regions.
  • Assessment of CAD enzymatic activity under varying nutrient conditions.

Main Results:

  • CAD was identified as a binding protein of mLST8, a component of mTOR complexes.
  • The interaction between CAD and mLST8 was confirmed for both transfected and endogenous proteins.
  • mLST8 recognizes CAD and mTOR through distinct regions.
  • CAD enzymatic activity was reduced under conditions of mTOR suppression (low amino acids and serum).

Conclusions:

  • mLST8 acts as a bridge connecting CAD and mTOR.
  • CAD is regulated within the mTOR pathway via its association with mLST8.
  • This interaction suggests a novel signaling mechanism linking nutrient status to pyrimidine synthesis.

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