Molecular Architecture of a 40S Ribosomal Subunit Maturation Intermediate.

The biogenesis of eukaryotic ribosomes is a highly regulated process and requires approximately 200 assembly factors. The process begins with transcription of ribosomal RNA (rRNA) in the nucleolus followed by maturation events in both the nucleolus and the cytoplasm. Although many assembly factors have been identified, detailed knowledge of their specific functions and binding locations on ribosomal precursor subunits is lacking. To understand the maturation of the 40S subunit (40S) in more detail, we characterized the 3D structure of a late cytoplasmic 40S subunit precursor using cryo-EM. To understand the function and binding sites of the seven assembly factors associated at this stage, we calculated the 3D structures of premature 40S ribosomal complexes lacking individual assembly factors. The localization of assembly factors reveals how their binding inhibits multiple steps in the formation of the translation competent 80S complex. Collectively, the assembly factors obstruct the binding sites of initiation factors, prevent the opening of the messenger RNA (mRNA) channel, block 60S subunit (60S) joining, and disrupt the rRNA nucleotides comprising the decoding site. In addition to their functions in formation of mature subunits, this work reveals how assembly factors act as external cues in the maturation status of 40S, with their dissociation being a prerequisite for proper 80S complex formation. This regulation is important as 80S complexes containing premature 40S subunits are rapidly degraded and aberrations in ribosome biogenesis are related to numerous human pathologiesPh.D.Biological ChemistryUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/86323/1/cloucks_1.pd