The intergenic region internal ribosome entry site (IGR IRES) of the Dicistroviridae viral family can directly assemble 80S ribosomes and initiate translation at a non-AUG codon from the ribosomal A-site. These functions are directed by two independently folded domains of the IGR IRES. One domain, composed of overlapping pseudoknots II and III (PKII/III), mediates ribosome recruitment. The seco...

Ribosome profiling identifies ribosome positions on translated mRNAs. A prominent feature of published datasets is the near complete absence of ribosomes in 3' untranslated regions (3'UTR) although substantial ribosome density can be observed on non-coding RNAs. Here we perform ribosome profiling in cultured Drosophila and human cells and show that different features of translation are revealed...

In Escherichia coli, rRNAs are initially transcribed with precursor sequences, which are subsequently removed through processing reactions. To investigate the role of precursor sequences, we analyzed ribosome assembly in strains containing mutations in the processing RNases. We observed that defects in 23S rRNA processing resulted in an accumulation of ribosomal subunits and caused a significan...

During translation elongation, the ribosome ratchets along its mRNA template, incorporating each new amino acid and translocating from one codon to the next. The elongation cycle requires dramatic structural rearrangements of the ribosome. We show here that deep sequencing of ribosome-protected mRNA fragments reveals not only the position of each ribosome but also, unexpectedly, its particular ...

In current models, ribosome release from the endoplasmic reticulum (ER) is coupled to the termination of protein translation. Thus, coincident with termination, membrane-bound ribosomes dissociate into their component subunits and are released into the cytosol. Here, we review past and current data and propose that the affinity of the ribosome for the ER membrane is decreased during translation...

Investigating translationally active mRNA involves isolating and sequencing RNA directly associated with the ribosome. Traditionally, isolating these complexes involved sucrose gradient ultracentrifugation, which is tedious and technically difficult. ARTseqTM (active RNA translation) Ribosome Profiling Kits provide an alternative method for obtaining ribosome-protected RNA fragments using size-...

of the Thesis ......................................................................................... xiii Chapter 1: Introduction ......................................................................................... 1 The Role of the Ribosome and Its Structure .............................................. 1 Ribosome Structure Overview........................................................

Many antibiotics target the ribosome and interfere with its translation cycle. Since translation is the source of all cellular proteins including ribosomal proteins, protein synthesis and ribosome assembly are interdependent. As a consequence, the activity of translation inhibitors might indirectly cause defective ribosome assembly. Due to the difficulty in distinguishing between direct and ind...

Structural studies have revealed that the core of the ribosome structure is conserved among ribosomes of all kingdoms. Kingdom-specific ribosomal proteins (r-proteins) are located in peripheral parts of the ribosome. In this work, the interactions between rRNA and r-proteins of eukaryote Saccharomyces cerevisiae ribosome were investigated applying LiCl induced splitting and quantitative mass sp...