Abstract
Ribosomal subunits are assembled on a precursor rRNA that includes four spacers in addition to mature rRNA sequences. The 5' external transcribed spacer (5' ETS) is the most prominent one that recruits U3 snoRNA and a plethora of proteins during the early assembly of 90S small subunit preribosomes. Here, we have conducted a comprehensive mutational analysis of 5' ETS by monitoring the processing and assembly of a plasmid-expressed pre-18S RNA. Remarkably, nearly half of the 5' ETS sequences, when depleted individually, are dispensable for 18S rRNA processing. The dispensable elements largely bind at the surface of the 90S structure. Defective assembly of 5' ETS completely blocks the last stage of 90S formation yet has little effect on the early assembly of 5' and central domains of 18S rRNA. Our study reveals the functional regions of 5' ETS and provides new insight into the assembly hierarchy of 90S preribosomes.