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Rapid Generation of Amyloid from Native Proteins In vitro
05:48

Rapid Generation of Amyloid from Native Proteins In vitro

Published on: December 5, 2013

Growth factor-mediated stabilization of amyloid precursor protein mRNA is mediated by a conserved 29-nucleotide

L E Rajagopalan1, J S Malter

  • 1Department of Pathology and Laboratory Medicine, University of Wisconsin Medical School, Madison 53792, USA.

Journal of Neurochemistry
|January 5, 2000
PubMed

Insights

A 29-nucleotide element in amyloid precursor protein (APP) mRNA acts as a destabilizer. Growth factors in cell culture media inhibit this element, stabilizing APP mRNA translation and turnover.

Area of Science:

  • Molecular Biology
  • Cell Biology
  • Biochemistry

Background:

  • Amyloid precursor protein (APP) mRNA turnover is regulated by specific sequence elements.
  • A previously identified 29-nucleotide instability element in the APP 3'-untranslated region (UTR) influences mRNA translation and stability.

Purpose of the Study:

  • To investigate the regulatory role of the 29-nucleotide APP mRNA instability element in primary human capillary endothelial cells.
  • To determine the effect of nutritional conditions, specifically growth factors, on the function of this mRNA instability element.

Main Methods:

  • Transfection of in vitro transcribed beta-globin-APP chimeric mRNAs (wild-type and mutant 29-nucleotide element) into primary human capillary endothelial cells.
  • Culturing cells in standard endothelial cell growth medium versus medium lacking supplemental growth factors.
  • Measurement of mRNA decay rates (half-life) using quantitative methods.

Main Results:

  • The 29-nucleotide element in the APP 3'-UTR destabilizes mRNA, with wild-type mRNA showing rapid decay.
  • Removal of growth factors from the culture medium significantly accelerated the decay of wild-type APP mRNA.
  • Mutating the 29-nucleotide element partially protected the mRNA from decay, even in the absence of growth factors.

Conclusions:

  • The 29-nucleotide 3'-UTR element functions as an mRNA destabilizer for amyloid precursor protein.
  • The inhibitory effect of this mRNA destabilizer can be significantly reduced by the presence of specific growth factors in the culture medium.

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