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Review
. 2024 Aug 9;14(8):975.
doi: 10.3390/biom14080975.

Putting It All Together: The Roles of Ribosomal Proteins in Nucleolar Stages of 60S Ribosomal Assembly in the Yeast Saccharomyces cerevisiae

Taylor N Ayers  1 , John L Woolford  1
Affiliations
Review

Putting It All Together: The Roles of Ribosomal Proteins in Nucleolar Stages of 60S Ribosomal Assembly in the Yeast Saccharomyces cerevisiae

Taylor N Ayers et al. Biomolecules. .

Abstract

Here we review the functions of ribosomal proteins (RPs) in the nucleolar stages of large ribosomal subunit assembly in the yeast Saccharomyces cerevisiae. We summarize the effects of depleting RPs on pre-rRNA processing and turnover, on the assembly of other RPs, and on the entry and exit of assembly factors (AFs). These results are interpreted in light of recent near-atomic-resolution cryo-EM structures of multiple assembly intermediates. Results are discussed with respect to each neighborhood of RPs and rRNA. We identify several key mechanisms related to RP behavior. Neighborhoods of RPs can assemble in one or more than one step. Entry of RPs can be triggered by molecular switches, in which an AF is replaced by an RP binding to the same site. To drive assembly forward, rRNA structure can be stabilized by RPs, including clamping rRNA structures or forming bridges between rRNA domains.

Keywords: assembly factors; cryo-electron microscopy; large ribosomal subunit; pre-rRNA processing; protein–rRNA interactions; rRNA folding; ribosomal proteins; ribosome assembly; ribosome assembly intermediates.

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Conflict of interest statement

The authors declare no conflicts of interest.

Figures

Figure 1
Figure 1
Each neighborhood of rRNA and RPs in the crystal structure of the yeast mature large ribosomal subunit [47], in the order with which they assemble: (A) The entire large subunit. Ribosomal proteins are color coded according to the neighborhood in which they are located. Ribosomal proteins colored in white assemble after nucleolar stages and thus are not discussed in this review; (B) domain I (red); (C) domain II (orange); (D) proximal stem, PS (pink); (E) domain VI (purple); (F) domain III (yellow); (G) domain IV (green); (H) domain V (blue); (I) central protuberance, CP (navy blue). Structures are shown in the subunit interface view on the left and the solvent-exposed surface on the right, except for domain II. In this case, orientations are shown to optimize the visualization of each RP. Note, though ribosomal protein L1 is present, it is not resolved in the mature LSU crystal structure (PDB: 4V88).
Figure 2
Figure 2
Domain I (RPs L8, L13, L15, L36): (A) Eight assembly factors (AFs) (cyan) bind to the exterior of the domain I RNP in the Noc1-Noc2 particle during early stages of assembly (PDB: 8E5T) [26]; (B) the assembly hierarchy implied by the effects of depleting RPs L8, L15, or L36; (C) binding of L8 and L15 to domain I rRNA and to each other in Noc1-Noc2 pre-ribosomes (PDB: 8E5T). Flexible rRNA is depicted as transparent in the secondary structure. Regions where L8 and L15 contact rRNA are indicated; (D) the globular domain of L8 (red) binds to rRNA domain I, while the 70-amino acid long N-terminal domain (grey) binds to rRNA domain V (PDB: 4V88).
Figure 3
Figure 3
Domain II: (A) Structure of node 1 containing RPs L4, L7, L18, and L32 (left), and of node 2 containing RPs L6, L14, L16, L20, and L33 (right) (PDB: 4V88). Domain VI (purple) is shown docked onto domain II rRNA. The bottom panel depicts the contact points of each protein with the rRNA; (B) a molecular switch between the co-transcriptional Noc1-Noc2 particle and the State 2 intermediate (PDB: 8E5T, 6C0F) [20,26]. State A (lower resolution) precedes State 2 and is not shown; (C) docking of domain VI rRNA onto domain II rRNA during the transition from the co-transcriptional Noc1-Noc2 particle to the State 2 intermediate. Panels below show extensions of L14 and L16 binding to domain VI after compaction. Truncations of L14 and L16 C-termini (grey) result in lethality (PDB: 8E5T, 6C0F); (D) binding of AF Rrp1 to node 1 domain II RPs (PDB: 8E5T).
Figure 3
Figure 3
Domain II: (A) Structure of node 1 containing RPs L4, L7, L18, and L32 (left), and of node 2 containing RPs L6, L14, L16, L20, and L33 (right) (PDB: 4V88). Domain VI (purple) is shown docked onto domain II rRNA. The bottom panel depicts the contact points of each protein with the rRNA; (B) a molecular switch between the co-transcriptional Noc1-Noc2 particle and the State 2 intermediate (PDB: 8E5T, 6C0F) [20,26]. State A (lower resolution) precedes State 2 and is not shown; (C) docking of domain VI rRNA onto domain II rRNA during the transition from the co-transcriptional Noc1-Noc2 particle to the State 2 intermediate. Panels below show extensions of L14 and L16 binding to domain VI after compaction. Truncations of L14 and L16 C-termini (grey) result in lethality (PDB: 8E5T, 6C0F); (D) binding of AF Rrp1 to node 1 domain II RPs (PDB: 8E5T).
Figure 4
Figure 4
Proximal stem (RPs L17, L26, L35, and L37): (A) The structure of the mature proximal stem RNP (PDB: 4V88); (B) processing of the ITS2 spacer rRNA to create the mature proximal stem (PDB: 6C0F, 4V88); (C) Maturation of the proximal stem during the transition from the Noc1-Noc2 particle to the State A intermediate (PDB: 8E5T, 6EM3) [18,26]; (D) a molecular switch resulting in the replacement of AF Noc1 with RP L17 and AF Noc2 with L26, Nsa1, and Rpf1 during the transition from Noc1-Noc2 to State 2 (PDB: 8E5T, 6C0F).
Figure 4
Figure 4
Proximal stem (RPs L17, L26, L35, and L37): (A) The structure of the mature proximal stem RNP (PDB: 4V88); (B) processing of the ITS2 spacer rRNA to create the mature proximal stem (PDB: 6C0F, 4V88); (C) Maturation of the proximal stem during the transition from the Noc1-Noc2 particle to the State A intermediate (PDB: 8E5T, 6EM3) [18,26]; (D) a molecular switch resulting in the replacement of AF Noc1 with RP L17 and AF Noc2 with L26, Nsa1, and Rpf1 during the transition from Noc1-Noc2 to State 2 (PDB: 8E5T, 6C0F).
Figure 5
Figure 5
Domain VI (RPs L3, L9, and L23): (A) The sequential assembly of domain VI from State A to State C intermediates (PDB: 6EM3, 6C0F, 6EM1). Bottom panel: secondary structure diagrams indicating the successive stabilization of domain VI rRNA, including contacts with the domain VI RPs; (B) L3 contacts all six root helices, and is proximal to the 5′ end of 5.8S rRNA and the 3′ end of 25S rRNA in the mature LSU (PDB: 4V88); (C) the assembly of AFs Nog1, Rlp24, and Tif6 proximal to L23 is most affected by depletion of L23 (PDB: 6C0F); (D) the assembly of AFs Nug1, Nsa2, and Dbp10 adjacent to L9 is most affected by its depletion (PDB: 6EM1). Note that Dbp10 is invisible in these particles; (E) L23 stabilizes both conformations of H92 (H92-alt, then H92 in its mature form) (PDB: 8V83, 8V87) [23].
Figure 6
Figure 6
Domain III (RPs L19, L22, L25, L27, L30, L31, L34, and L38): (A) The locations of domain III RPs in mature LSU based on structural features. L19, L31, and L34 are shown embedded on the domain III root helix. L22, L27, L30, L38 are bound to the exterior surface of the domain III rRNA. The L25 globular domain (yellow) binds domain III rRNA, and the N-terminal extension of L25 containing amino acids 1–61 (grey) contacts neighboring rRNA domains (PDB: 4V88); (B) the completion of exit platform construction by assembly of RPs L19, L25, and L31 (PDB: 6C0F, 7NAC) [20,21].
Figure 7
Figure 7
Domain IV (RPs L2 and L43): (A) L2 and L43 are positioned between rRNA domains II, III, IV, and V in the mature LSU (PDB: 4V88); (B) the entry of L2 and L43 enables compaction of rRNA domain IV and stabilizes the rotation of the L1 stalk (PDB: 7NAC, 7U0H, 6YLY) [21,28]. The bottom panel depicts changes in the stabilization of domain IV rRNA during this interval; (C) significant particle remodeling occurs during the entry of L2 and L43 just prior to the exit of pre-60S particles from the nucleolus to the nucleoplasm. AFs that contain intrinsically disordered domains and that exit at this interval are shown in bold. Note that Nip7, Nop2, and the L1 stalk are invisible in State NE1. (PDB: 7NAC, 7U0H).
Figure 8
Figure 8
Domain V (RPs L1, L21, and L28): (A) The location of L21 and L28 in domain V beneath the central protuberance (CP) in the mature LSU. Note that L1 is not resolved in this structure (PDB: 4V88); (B) the C-terminal domain of L21 is stabilized on domain II RPs in State E1 (PDB: 7R7A); (C) a molecular switch: the exit of Brx1 enables the entry of the C-terminal globular domain of L28 (7NAC, 7U0H); (D) the assembly of L10, L24, L29, and L40 during nucleoplasmic and cytoplasmic stages of LSU construction is most affected by the absence of L21 or L28 (PDB: 4V88); (E) the rotation of the L1 stalk during the transition of the state E1 intermediate to state NE2. Note that L1 and H77-78 are not resolved following L1 stalk rotation in the nucleus. L1 and H77-78 are overlayed onto the NE2 structure (PDB: 7NAC, 6YLY).
Figure 9
Figure 9
The polypeptide exit tunnel (RP L39): (A) The polypeptide exit tunnel is comprised of portions of rRNA domains I, II, III, IV, and V. Segments of L4 and L17 are inserted into the tunnel and L39 is embedded near the tunnel exit (PDB: 4V88); (B) the presence of L39 reduces the diameter of the tunnel exit. The space outlined in green highlights the size of the tunnel exit in the absence of L39. In contrast, the space outlined in blue indicates the reduced dimensions of the tunnel exit when L39 is present (PDB: 4V88); (C) particles before and after L39 entry, including RPs surrounding the tunnel exit. The C-terminal tail of Nog1 enters the tunnel coincident with L39 in the Nog2 assembly intermediate (PDB: 6YLY, 3JCT) [28,70].
Figure 10
Figure 10
The central protuberance (RPs L5, L11): RPs L5 and L11 bind to the separately transcribed 5S rRNA and dock onto domain V on top of the LSU to form the central protuberance. The 5S RNP assembles with pre-ribosomes early, but is not visible by cryo-EM until the transition from state NE2 (top) to the Nog2 particle (middle). The 5S RNP undergoes ~180° rotation to form the mature structure (bottom) (PDB: 6YLY, 3JCT, 4V88) [28,47,70]. Abbreviation: peptidyltransferase center, PTC. Adapted from [28].
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References

    1. Petrov A.S., Bernier C.R., Hershkovits E., Xue Y., Waterbury C.C., Hsiao C., Stepanov V.G., Gaucher E.A., Grover M.A., Harvey S.C., et al. Secondary Structure and Domain Architecture of the 23S and 5S rRNAs. Nucleic Acids Res. 2013;41:7522–7535. doi: 10.1093/nar/gkt513. - DOI - PMC - PubMed
    1. Petrov A.S., Bernier C.R., Gulen B., Waterbury C.C., Hershkovits E., Hsiao C., Harvey S.C., Hud N.V., Fox G.E., Wartell R.M., et al. Secondary Structures of rRNAs from All Three Domains of Life. PLoS ONE. 2014;9:e88222. doi: 10.1371/journal.pone.0088222. - DOI - PMC - PubMed
    1. Baßler J., Hurt E. Eukaryotic Ribosome Assembly. Annu. Rev. Biochem. 2019;88:281–306. doi: 10.1146/annurev-biochem-013118-110817. - DOI - PubMed
    1. Fernández-Pevida A., Kressler D., De La Cruz J. Processing of Preribosomal RNA in Saccharomyces Cerevisiae: Yeast Pre-rRNA Processing. WIREs RNA. 2015;6:191–209. doi: 10.1002/wrna.1267. - DOI - PubMed
    1. Vanden Broeck A., Klinge S. Eukaryotic Ribosome Assembly. Annu. Rev. Biochem. 2024;93:189–210. doi: 10.1146/annurev-biochem-030222-113611. - DOI - PubMed

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