Abstract
In eukaryotes, the binding of poly(A) binding protein (PAB) to the poly(A) tail is central to maintaining mRNA stability. PABP interacts with the translation termination apparatus, and with eIF4G to maintain 3′–5′ mRNA interactions as part of an mRNA closed loop. It is however unclear how ribosome recycling on a closed loop mRNA is influenced by the proximity of the stop codon to the poly(A) tail, and how post-termination ribosome recycling affects mRNA stability. We show that in a yeast disabled for nonsense mediated mRNA decay (NMD), a PGK1 mRNA with an early stop codon at codon 22 of the reading frame is still highly unstable, and that this instability cannot be significantly countered even when 50% stop codon readthrough is triggered. In an NMD-deficient mutant yeast, stable reporter alleles with more 3′ proximal stop codons could not be rendered unstable through Rli1-depletion, inferring defective Rli1 ribosome recycling is insufficient in itself to trigger mRNA instability. Mathematical modelling of a translation system including the effect of ribosome recycling and poly(A) tail shortening supports the hypothesis that impaired ribosome recycling from 5′ proximal stop codons may compromise initiation processes and thus destabilize the mRNA. A model is proposed wherein ribosomes undergo a maturation process during early elongation steps, and acquire competency to re-initiate on the same mRNA as translation elongation progresses beyond the very 5′ proximal regions of the mRNA.
Highlights
During expression of genetic information into protein in a eukaryote cell, the primary mRNA transcript is processed and exported into the cytoplasm for translation by the cellular ribosomes.The amount of translated protein produced through gene expression is affected by multiple forms of control, for example at the level of the translational efficiency of the mRNA, and through regulation of protein stability
The release factor eRF3 and poly(A) binding protein functionally interact on eukaryote mRNAs, linking translation termination to the process of poly(A) tail shortening and mRNA stability
Poly(A) binding protein interacts with eIF4G as part of a 30 –50 closed loop interaction between the two ends of an mRNA, linking the process of translation termination to ribosome recycling
Summary
During expression of genetic information into protein in a eukaryote cell, the primary mRNA transcript is processed and exported into the cytoplasm for translation by the cellular ribosomes.The amount of translated protein produced through gene expression is affected by multiple forms of control, for example at the level of the translational efficiency of the mRNA, and through regulation of protein stability. During expression of genetic information into protein in a eukaryote cell, the primary mRNA transcript is processed and exported into the cytoplasm for translation by the cellular ribosomes. MRNA stability is imparted by both these elements, in combination with the eIF4 cap-binding complex, and poly(A)-binding protein PABP. These mRNP complexes impart stability by protecting the mRNA from exonucleases that act in a 50 –30 direction (Xrn1p in yeast; [3]) or in a 30 –50 direction PABP binding at the 30 end interacts with the eIF4G component of the eIF4F cap binding complex at the 50 end of the mRNA, creating a pseudo-circular mRNA molecule that enhances stability and translational efficiency [5,6]
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