Mammalian HCA66 protein is required for both ribosome synthesis and centriole duplication.

Ribosome production, one of the most energy-consuming biosynthetic activities in living cells, is adjusted to growth conditions and coordinated with the cell cycle. Connections between ribosome synthesis and cell cycle progression have been described, but the underlying mechanisms remain only partially understood. The human HCA66 protein was recently characterized as a component of the centrosome, the major microtubule-organizing center (MTOC) in mammalian cells, and was shown to be required for centriole duplication and assembly of the mitotic spindle.

BC1-FMRP interaction is modulated by 2'-O-methylation: RNA-binding activity of the tudor domain and translational regulation at synapses.

The brain cytoplasmic RNA, BC1, is a small non-coding RNA that is found in different RNP particles, some of which are involved in translational control. One component of BC1-containing RNP complexes is the fragile X mental retardation protein (FMRP) that is implicated in translational repression. Peptide mapping and computational simulations show that the tudor domain of FMRP makes specific contacts to BC1 RNA.

Evolutionarily conserved function of RRP36 in early cleavages of the pre-rRNA and production of the 40S ribosomal subunit.

Ribosome biogenesis in eukaryotes is a major cellular activity mobilizing the products of over 200 transcriptionally coregulated genes referred to as the rRNA and ribosome biosynthesis regulon. We investigated the function of an essential, uncharacterized gene of this regulon, renamed RRP36. We show that the Rrp36p protein is nucleolar and interacts with 90S and pre-40S preribosomal particles.

The ATPase and helicase activities of Prp43p are stimulated by the G-patch protein Pfa1p during yeast ribosome biogenesis.

Prp43p is a RNA helicase required for pre-mRNA splicing and for the synthesis of large and small ribosomal subunits. The molecular functions and modes of regulation of Prp43p during ribosome biogenesis remain unknown. We demonstrate that the G-patch protein Pfa1p, a component of pre-40S pre-ribosomal particles, directly interacts with Prp43p.

Immature small ribosomal subunits can engage in translation initiation in Saccharomyces cerevisiae.

It is generally assumed that, in Saccharomyces cerevisiae, immature 40S ribosomal subunits are not competent for translation initiation. Here, we show by different approaches that, in wild-type conditions, a portion of pre-40S particles (pre-SSU) associate with translating ribosomal complexes. When cytoplasmic 20S pre-rRNA processing is impaired, as in Rio1p- or Nob1p-depleted cells, a large part of pre-SSUs is associated with translating ribosomes complexes.

Characterization of Saccharomyces cerevisiae Npa2p (Urb2p) reveals a low-molecular-mass complex containing Dbp6p, Npa1p (Urb1p), Nop8p, and Rsa3p involved in early steps of 60S ribosomal subunit biogenesis.

We report the characterization of the yeast Npa2p (Urb2p) protein, which is essential for 60S ribosomal subunit biogenesis. We identified this protein in a synthetic lethal screening with the rsa3 null allele. Rsa3p is a genetic partner of the putative RNA helicase Dbp6p.

Analysis of the binding of the N-terminal conserved domain of yeast Cbf5p to a box H/ACA snoRNA.

During ribosome biogenesis, the RNA precursor to mature rRNAs undergoes numerous post-transcriptional chemical modifications of bases, including conversions of uridines to pseudouridines. In archaea and eukaryotes, these conversions are performed by box H/ACA small ribonucleoprotein particles (box H/ACA RNPs), which contain a small guide RNA responsible for the selection of substrate uridines and four proteins, including the pseudouridine synthase, Cbf5p. So far, no in vitro reconstitution of eukaryotic box H/ACA RNPs from purified components has been achieved, principally due to difficulties in purifying recombinant eukaryotic Cbf5p.

hNaf1 is required for accumulation of human box H/ACA snoRNPs, scaRNPs, and telomerase.

The human telomerase ribonucleoprotein particle (RNP) shares with box H/ACA small Cajal body (sca)RNPs and small nucleolar (sno)RNPs the proteins dyskerin, hGar1, hNhp2, and hNop10. How dyskerin, hGar1, hNhp2, and hNop10 assemble with box H/ACA scaRNAs, snoRNAs, and the RNA component of telomerase (hTR) in vivo remains unknown. In yeast, Naf1p interacts with H/ACA snoRNP proteins and may promote assembly of Cbf5p (the yeast ortholog of dyskerin) with nascent pre-snoRNAs.

Structural study of the H/ACA snoRNP components Nop10p and the 3' hairpin of U65 snoRNA.

The H/ACA small nucleolar ribonucleoprotein (snoRNP) complexes guide the modification of uridine to pseudouridine at conserved sites in rRNA. The H/ACA snoRNPs each comprise a target-site-specific snoRNA and four core proteins, Nop10p, Nhp2p, Gar1p, and the pseudouridine synthase, Cbf5p, in yeast. The secondary structure of the H/ACA snoRNAs includes two hairpins that each contain a large internal loop (the pseudouridylation pocket), one or both of which are partially complementary to the target RNA(s).

The splicing ATPase prp43p is a component of multiple preribosomal particles.

Prp43p is a putative helicase of the DEAH family which is required for the release of the lariat intron from the spliceosome. Prp43p could also play a role in ribosome synthesis, since it accumulates in the nucleolus. Consistent with this hypothesis, we find that depletion of Prp43p leads to accumulation of 35S pre-rRNA and strongly reduces levels of all downstream pre-rRNA processing intermediates.

Cbf5p, the putative pseudouridine synthase of H/ACA-type snoRNPs, can form a complex with Gar1p and Nop10p in absence of Nhp2p and box H/ACA snoRNAs.

Box C/D and box H/ACA small ribonucleoprotein particles (sRNPs) are found from archaea to humans, and some of these play key roles during the biogenesis of ribosomes or components of the splicing apparatus. The protein composition of the core of both types of particles is well established and the assembly pathway of box C/D sRNPs has been extensively investigated both in archaeal and eukaryotic systems. In contrast, knowledge concerning the mode of assembly and final structure of box H/ACA sRNPs is much more limited.

Npa1p, a component of very early pre-60S ribosomal particles, associates with a subset of small nucleolar RNPs required for peptidyl transferase center modification.

We have identified a novel essential nucleolar factor required for the synthesis of 5.8S and 25S rRNAs termed Npa1p. In the absence of Npa1p, the pre-rRNA processing pathway leading to 5.

Dim2p, a KH-domain protein required for small ribosomal subunit synthesis.

Recent proteomic analyses are revealing the dynamics of preribosome assembly. Following cleavage at processing site A(2), which generates the 20S pre-rRNA (the immediate precursor to the 18S rRNA), early RRPs (ribosomal RNA processing factors) are released in bulk from the preribosomes, and the resulting pre-40S subunits are left associated with a limited set of proteins that we refer to as the SSU RRP complex. Dim2p, a core constituent of the SSU RRP complex and conserved KH-domain containing protein, is required for pre-rRNA processing and is associated with early nucleolar and late cytoplasmic pre-rRNA species.

Late cytoplasmic maturation of the small ribosomal subunit requires RIO proteins in Saccharomyces cerevisiae.

Numerous nonribosomal trans-acting factors involved in pre-rRNA processing have been characterized, but few of them are specifically required for the last cytoplasmic steps of 18S rRNA maturation. We have recently demonstrated that Rrp10p/Rio1p is such a factor. By BLAST analysis, we identified the product of a previously uncharacterized essential gene, YNL207W/RIO2, called Rio2p, that shares 43% sequence similarity with Rrp10p/Rio1p.

Naf1p, an essential nucleoplasmic factor specifically required for accumulation of box H/ACA small nucleolar RNPs.

Box H/ACA small nucleolar ribonucleoprotein particles (H/ACA snoRNPs) play key roles in the synthesis of eukaryotic ribosomes. The ways in which these particles are assembled and correctly localized in the dense fibrillar component of the nucleolus remain largely unknown. Recently, the essential Saccharomyces cerevisiae Naf1p protein (encoded by the YNL124W open reading frame) was found to interact in a two-hybrid assay with two core protein components of mature H/ACA snoRNPs, Cbf5p and Nhp2p (T.