Tetrapeptide 60-63 of human ribosomal protein uS3 is crucial for translation initiation

Elena Babaylova, Alexey Malygin, Alexander Gopanenko, Dmitri Graifer, Galina Karpova

Research output: Contribution to journalArticlepeer-review

4 Citations (Scopus)

Abstract

Conserved ribosomal protein uS3 contains a decapeptide fragment in positions 55-64 (human numbering), which has a very specific ability to cross-link to various RNA derivatives bearing aldehyde groups, likely provided by K62. It has been shown that during translation in the cell-free protein-synthesizing system, uS3 becomes accessible for such cross-linking only after eIF3j leaves the mRNA binding channel of the 40S ribosomal subunit. We studied the functional role of K62 and its nearest neighbors in the ribosomal assembly and translation with the use of HEK293T-derived cell cultures capable of producing FLAG-tagged uS3 (uS3FLAG) or its mutant form with amino acid residues at positions 60-63 replaced with alanines. Analysis of polysome profiles from the respective cells and cytosol lysates showed that the mutation significantly affected the uS3 ability to participate in the assembly of 40S subunits, but it was not essential for their maturation and did not prevent the binding of mRNAs to 40S subunits during translation initiation. The most striking effect of the replacement of amino acid residues in the above uS3 positions was that it almost completely deprived the 40S subunits of their ability to form 80S ribosomes, suggesting that the 48S pre-initiation complexes assembled on these subunits were defective in the binding of 60S subunits. Thus, our results revealed the previously unknown crucial role of the uS3 tetrapeptide 60GEKG63 in translation initiation related to maintaining the proper structure of the 48S complex, most likely via the prevention of premature mRNA loading into the ribosomal channel.

Original languageEnglish
Article number194411
Pages (from-to)194411
Number of pages8
JournalBiochimica et biophysica acta. Gene regulatory mechanisms
Volume1862
Issue number9
DOIs
Publication statusPublished - 1 Sep 2019

Keywords

  • Amino acid replacements
  • KH domain
  • Mammalian ribosomal protein uS3
  • Pre-initiation complexes
  • Small ribosomal subunit maturation
  • Translation initiation
  • S3
  • EIF3
  • STATES
  • MESSENGER-RNA
  • ENTRY CHANNEL

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