Recent Advances on the Structure and Function of RNA Acetyltransferase Kre33/NAT10

Abstract

Ribosome biogenesis is one of the most energy demanding processes in the cell. In eukaryotes, the main steps of this process occur in the nucleolus and include pre-ribosomal RNA (pre-rRNA) processing, post-transcriptional modifications, and assembly of many non-ribosomal factors and ribosomal proteins in order to form mature and functional ribosomes. In yeast and humans, the nucleolar RNA acetyltransferase Kre33/NAT10 participates in different maturation events, such as acetylation and processing of 18S rRNA, and assembly of the 40S ribosomal subunit. Here, we review the structural and functional features of Kre33/NAT10 RNA acetyltransferase, and we underscore the importance of this enzyme in ribosome biogenesis, as well as in acetylation of non-ribosomal targets. We also report on the role of human NAT10 in Hutchinson-Gilford progeria syndrome.

Keywords: Hutchinson–Gilford progeria syndrome; Kre33; NAT10; SSU processome; nuclear localization signal; nuclear pore complex; nucleolar localization signal; post-transcriptional modifications; ribosome biogenesis.

Figure 1

Processing of pre-ribosomal RNA (pre-rRNA) and small subunit (SSU) processome formation in the yeast Saccharomyces cerevisiae. (A) Scheme of 35S pre-rRNA processing steps. The 35S precursor is cleaved and trimmed at different sites to form the mature 18S (blue), 5.8S_L, 5.8S_S (red) and 25S (yellow) rRNAs. Processing takes place in different cellular compartments (nucleolus, nucleoplasm and cytoplasm), represented by rectangles. Cleavage sites are indicated on the transcripts and are detailed in the text. The final maturation steps occur in the cytoplasm once the late precursors have been exported via the nuclear pore complex (NPC, colored in green). (B) Representation of SSU processome formation. The ribosomal DNA (rDNA) is transcribed by RNA polymerase I (Pol I) (green circles). When visualized by electron microscopy, actively transcribed rDNA units appear as Christmas trees, wherein chromatin forms the trunk, nascent rRNA transcripts are the branches, and the terminal knob seen at the 5′ end of transcripts is the forming SSU processome. The small black circles represent the newly formed SSU knobs and the larger black circles are the mature SSU knobs, where the 5′ETS and the 18S rRNA are packed. The large SSU knob is then cleaved at site A2 (represented by the black scissors). The large subunit precursor (pre-LSU) knobs are shown in grey circles. This schematic representation is adapted from Osheim et al. [70].

Figure 2

Conserved domains and motifs of Kre33. (A) Representation of yeast Kre33 highlighting the conserved domains/motifs. From left to right, the N-terminal extension (NTE), containing putative nuclear localization signal (NLS) and nucleolar localization signal (NoLS) motifs, colored in dark blue and magenta, respectively. Overlapping NLS and NoLS regions are shown in purple. The NTE is followed by a domain of unknown function called DUF1726 (light blue), a helicase domain (blue), three TmcA-specific motifs (TS1, TS2 and TS3) colored respectively in light green, dark green and yellow, a N-acetyltransferase domain (orange) and a possible tRNA-binding domain (paprika). These highly conserved domains precede a C-terminal extension (CTE) harboring a putative coiled-coil motif (red) followed by the NLS (blue) and NoLS (magenta), with overlapping regions in purple. Point mutations generated by Sharma et al. [22] are indicated. (B) Tridimensional representation of bacterial TmcA and yeast Kre33. Structures were modeled with I-TASSER [101]. Conserved motifs were colored using PyMOL with the color code used in (A).

Figure 3

Alignment of Kre33/NAT10 from different species. Sequences from yeast, human, mouse, D. melanogaster (DROME), S. pombe (SCHPO), A. thaliana (ARATH) and C. elegans (CAEEL) were used to generate an alignment with Clustal Omega [102]. (A) The N-terminal extension precedes the DUF1726 (black bold letters) and harbors eukaryote-specific motifs, such as the NLS (blue) and the NoLS (magenta); overlapping NLS/NoLS sequences are in purple. (B) The C-terminal extension follows the tRNA-binding domain (black bold letters): it contains a coiled-coil motif (red) and finishes with the NLS (blue)/NoLS (magenta); the NLS/NoLS overlap is in purple. The putative NLS, NoLS and coiled-coil motif were identified with cNLS mapper [103], NoD [104] and COILS [105].

Figure 4

Hypothetical model for Kre33 acetylation of RNAs. In the nucleolus, the Kre33 homodimer and Enp2 protein may interact together via their C-terminal coiled-coil motifs (in red) before joining the SSU processome. In this particle, Kre33 catalyzes the acetylation of two 18S rRNA cytosines: each modification is guided by a different box C/D small nucleolar RNA (snoRNA), snR4 (C1280 in helix 34) and snR45 (C1773 in helix 45). Acetylation of tRNAs and mRNAs could occur in the nucleoplasm, where Kre33 needs to interacts with the adaptor protein Tan1 in order to acetylate tRNAs. It is not known if acetylation of mRNAs requires adaptor protein(s). Nevertheless, it remains possible that acetylation of tRNAs and mRNAs could take place in the nucleolus since tRNAs and mRNAs have previously been detected in this compartment [51,54,91,92].