rRNA modifications in an intersubunit bridge of the ribosome strongly affect both ribosome biogenesis and activity.

The presence of nucleotide modifications in rRNA has been known for nearly 40 years; however, information about their roles is sparse. Here, we describe the consequences of depleting modifications from an intersubunit bridge (helix 69) of the ribosomal large subunit in yeast. Helix 69 interacts with both A and P site tRNAs and contains five modifications. Blocking one to two modifications has no apparent effect on cell growth, whereas loss of three to five modifications impairs growth and causes the broadest defects observed thus far for modification loss in any ribosome region. Major effects include the following: (1) reduced amino acid incorporation rates in vivo (25%-60%); (2) increased stop codon readthrough activity; (3) increased sensitivity to ribosome-based antibiotics; (4) reduced rRNA levels (20%-50%), due mainly to faster turnover; and (5) altered rRNA structure in the ribosome. Taken together, the results indicate that this subset of rRNA modifications can influence both ribosome synthesis and function and in synergistic ways.