Alternative splicing results in differential expression, activity, and localization of the two forms of arginyl-tRNA-protein transferase, a component of the N-end rule pathway

Mol Cell Biol. 1999 Jan;19(1):182-93. doi: 10.1128/mcb.19.1.182.

Abstract

The N-end rule relates the in vivo half-life of a protein to the identity of its N-terminal residue. The underlying ubiquitin-dependent proteolytic system, called the N-end rule pathway, is organized hierarchically: N-terminal aspartate and glutamate (and also cysteine in metazoans) are secondary destabilizing residues, in that they function through their conjugation, by arginyl-tRNA-protein transferase (R-transferase), to arginine, a primary destabilizing residue. We isolated cDNA encoding the 516-residue mouse R-transferase, ATE1p, and found two species, termed Ate1-1 and Ate1-2. The Ate1 mRNAs are produced through a most unusual alternative splicing that retains one or the other of the two homologous 129-bp exons, which are adjacent in the mouse Ate1 gene. Human ATE1 also contains the alternative 129-bp exons, whereas the plant (Arabidopsis thaliana) and fly (Drosophila melanogaster) Ate1 genes encode a single form of ATE1p. A fusion of ATE1-1p with green fluorescent protein (GFP) is present in both the nucleus and the cytosol, whereas ATE1-2p-GFP is exclusively cytosolic. Mouse ATE1-1p and ATE1-2p were examined by expressing them in ate1Delta Saccharomyces cerevisiae in the presence of test substrates that included Asp-betagal (beta-galactosidase) and Cys-betagal. Both forms of the mouse R-transferase conferred instability on Asp-betagal (but not on Cys-betagal) through the arginylation of its N-terminal Asp, the ATE1-1p enzyme being more active than ATE1-2p. The ratio of Ate1-1 to Ate1-2 mRNA varies greatly among the mouse tissues; it is approximately 0.1 in the skeletal muscle, approximately 0.25 in the spleen, approximately 3.3 in the liver and brain, and approximately 10 in the testis, suggesting that the two R-transferases are functionally distinct.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Alternative Splicing*
  • Amino Acid Sequence
  • Aminoacyltransferases / genetics*
  • Aminoacyltransferases / metabolism*
  • Animals
  • Arabidopsis / genetics
  • Aspartic Acid
  • Base Sequence
  • Cell Line, Transformed
  • Cell Nucleus
  • Cysteine
  • Cytosol
  • DNA, Complementary
  • Drosophila melanogaster / genetics
  • Exons
  • Gene Expression Regulation
  • Glutamic Acid
  • Humans
  • Mice
  • Molecular Sequence Data
  • beta-Galactosidase

Substances

  • DNA, Complementary
  • Aspartic Acid
  • Glutamic Acid
  • Aminoacyltransferases
  • arginyltransferase
  • beta-Galactosidase
  • Cysteine

Associated data

  • GENBANK/AF079096
  • GENBANK/AF079097
  • GENBANK/AF079098
  • GENBANK/AF079099
  • GENBANK/AF079100
  • GENBANK/AF079101