Rrp5p, Noc1p and Noc2p form a protein module which is part of early large ribosomal subunit precursors in S. cerevisiae

Abstract

Eukaryotic ribosome biogenesis requires more than 150 auxiliary proteins, which transiently interact with pre-ribosomal particles. Previous studies suggest that several of these biogenesis factors function together as modules. Using a heterologous expression system, we show that the large ribosomal subunit (LSU) biogenesis factor Noc1p of Saccharomyces cerevisiae can simultaneously interact with the LSU biogenesis factor Noc2p and Rrp5p, a factor required for biogenesis of the large and the small ribosomal subunit. Proteome analysis of RNA polymerase-I-associated chromatin and chromatin immunopurification experiments indicated that all members of this protein module and a specific set of LSU biogenesis factors are co-transcriptionally recruited to nascent ribosomal RNA (rRNA) precursors in yeast cells. Further ex vivo analyses showed that all module members predominantly interact with early pre-LSU particles after the initial pre-rRNA processing events have occurred. In yeast strains depleted of Noc1p, Noc2p or Rrp5p, levels of the major LSU pre-rRNAs decreased and the respective other module members were associated with accumulating aberrant rRNA fragments. Therefore, we conclude that the module exhibits several binding interfaces with pre-ribosomes. Taken together, our results suggest a co- and post-transcriptional role of the yeast Rrp5p–Noc1p–Noc2p module in the structural organization of early LSU precursors protecting them from non-productive RNase activity.

Disruption of ribosome assembly in yeast blocks cotranscriptional pre-rRNA processing and affects the global hierarchy of ribosome biogenesis

“…We found that many pre-60S subunit assembly factors were enriched in Nog2-containing particles when Drs1 was depleted, especially early assembly factors such as Ssf1, Npa1, and Rrp5, all of which have been reported to dissociate from pre-ribosomes before 27SB pre-rRNA is generated and before Nog2 binds pre-ribosomes in wild-type cells (Fig. 3A,D,E ;Fatica et al 2002;Nissan et al 2002;Dez et al 2004;Hierlmeier et al 2012).…”

“…However, recent studies, including this one (Fig. 1), have shown that 23S pre-rRNA is likely an alternative, on-pathway intermediate to mature 18S rRNA Adams et al 2002;Miles et al 2005;Hierlmeier et al 2012;Jakovljevic et al 2012;Talkish et al 2014). Depletion of Drs1, like many pre-60S subunit assembly factors that function during processing of 27S pre-rRNA, results in a dramatic decrease in 20S pre-rRNA with a corresponding increase in 35S and 23S pre-rRNA levels (Fig.…”

“…Recently, a number of ribosome assembly factors were shown to copurify with RNA polymerase I (Pol I), suggesting that they are recruited to pre-ribosomes cotranscriptionally (Hierlmeier et al 2012). Many of these factors, including Drs1 and Nop7, are proteins that associate with pre-ribosomes early in ribosome biogenesis.…”

“…In wild-type cells, Nop7 copurifies with Pol I, indicating it normally associates with pre-ribosomes cotranscriptionally (Hierlmeier et al 2012). Thus, even upon a shift from coto more post-transcriptional pre-rRNA processing, Nop7 still associates with pre-rRNAs cotranscriptionally.…”

“…However, the 23S intermediate that accumulates in many mutants when early steps of 60S subunit assembly are blocked (including in the absence of Drs1, Fig. 1), can be processed to 18S rRNA and serves as an alternative route for maturation of the small subunit when large subunit assembly is blocked Adams et al 2002;Miles et al 2005;Hierlmeier et al 2012;Jakovljevic et al 2012;Talkish et al 2014).…”