Polyamine regulation of ribosome pausing at the upstream open reading frame of S-adenosylmethionine decarboxylase.
Synthesis of S-adenosylmethionine decarboxylase (AdoMetDC), a key regulated enzyme in the pathway of polyamine biosynthesis, is feedback-controlled at the level of translation by spermidine and spermine. The peptide product of an upstream open reading frame (uORF) in the mRNA is solely responsible for polyamine regulation of AdoMetDC translation. Using a primer extension inhibition assay and in vitro protein synthesis reactions, we found ribosomes paused at or close to the termination codon of the uORF. This pause was greatly diminished with the altered uORFs' sequences that abolish uORF regulation in vivo. The half-life of the ribosome pause was related to the concentration of polyamines present but was unaffected by magnesium concentration. Furthermore, inhibition of translation initiation at a reporter gene placed downstream of the AdoMetDC uORF directly correlated with the stability of the ribosome pause at the uORF. These observations are consistent with a model in which regulation of ribosome pausing at the uORF by polyamines controls ribosome access to the downstream AdoMetDC reading frame.
Law
G L
GL
Department of Biochemistry, University of Washington, Seattle, 98195-7350, USA.
Raney
A
A
Heusner
C
C
Morris
D R
DR
D012437:Adenosylmethionine Decarboxylase / Q000737:chemistry / Q000235:genetics*; D001483:Base Sequence; D015317:Biogenic Polyamines / Q000502:physiology*; D002474:Cell-Free System; D017931:DNA Primers; D016366:Open Reading Frames; D014176:Protein Biosynthesis / Q000502:physiology; D012270:Ribosomes / Q000502:physiology*
D015317:Biogenic Polyamines; D017931:DNA Primers; D012437:Adenosylmethionine Decarboxylase