Utp14 Recruits and Activates the RNA Helicase Dhr1 To Undock U3 snoRNA from the Preribosome

Abstract

bIn eukaryotic ribosome biogenesis, U3 snoRNA base pairs with the pre-rRNA to promote its processing. However, U3 must be removed to allow folding of the central pseudoknot, a key feature of the small subunit. Previously, we showed that the DEAH/ RHA RNA helicase Dhr1 dislodges U3 from the pre-rRNA. DHR1 can be linked to UTP14, encoding an essential protein of the preribosome, through genetic interactions with the rRNA methyltransferase Bud23. Here, we report that Utp14 regulates Dhr1. Mutations within a discrete region of Utp14 reduced interaction with Dhr1 that correlated with reduced function of Utp14. These mutants accumulated Dhr1 and U3 in a pre-40S particle, mimicking a helicase-inactive Dhr1 mutant. This similarity in the phenotypes led us to propose that Utp14 activates Dhr1. Indeed, Utp14 formed a complex with Dhr1 and stimulated its unwinding activity in vitro. Moreover, the utp14 mutants that mimicked a catalytically inactive dhr1 mutant in vivo showed reduced stimulation of unwinding activity in vitro. Dhr1 binding to the preribosome was substantially reduced only when both Utp14 and Bud23 were depleted. Thus, Utp14 is bifunctional; together with Bud23, it is needed for stable interaction of Dhr1 with the preribosome, and Utp14 activates Dhr1 to dislodge U3. Ribosomes are complex ribonucleoprotein (RNP) particles composed of an intricate assembly of folded rRNA interdigitated with ribosomal proteins (r-proteins). Faithful assembly of this RNP is critical for efficient and accurate cellular translation. Eukaryotic cells devote more than 200 trans-acting factors (1-3) to the assembly of ribosomes. This highly dynamic process (4) involves extensive structural rearrangements of RNA-RNA, RNA-protein, and protein-protein interactions (5, 6). To investigate how structural rearrangements are regulated to occur at the proper stage of assembly, our studies centered on the RNA helicase Dhr1, which dissociates a key RNA-RNA interaction, dislodging U3 snoRNA from the pre-rRNA of the preribosome (1, 3, 7).U3 is instrumental in orchestrating early pre-rRNA processing. Ribosome assembly in Saccharomyces cerevisiae begins with the cotranscriptional assembly of the 90S preribosome, a large complex of r-proteins and trans-acting factors, including proteins and snoRNAs, on the growing 5= end of the nascent 35S transcript (8-10). The pre-rRNA undergoes extensive modification and processing (11) to liberate the mature 18S, 5.8S, and 25S rRNAs that are embedded between transcribed spacer elements. U3 snoRNA is required for the early cleavages at sites A0 and A1 to generate the mature 5= end of 18S and cleavage at A2 to separate the pre-40S (containing 20S pre-rRNA) from the pre-60S subunits (12, 13). Under conditions in which cleavage at A2 is blocked or reduced, cleavage at A3 can generate the pre-60S precursor. Thus, U3 is required for biogenesis of the small but not the large subunit.U3 is also expected to promote proper rRNA folding. U3 hybridizes with the pre-rRNA at multiple sites within the 35S precursor: at sites ...

A Puzzle of Life: Crafting Ribosomal Subunits

“…Dismantling of the 90S preribosome is likely to depend on the RNA helicase Dhr1, since pre-40S intermediates trapped by catalytically inactive Dhr1 still contain the U3 snoRNP [67]. Moreover, Dhr1, stimulated by its recruiting cofactor Utp14, can unwind U3 rRNA duplexes in vitro [67,68]…”

Overexpression of U three protein 14a (UTP14a) is associated with poor prognosis of esophageal squamous cell carcinoma

“…UTP14 is a subunit of the U3 containing small subunit (SSU) processome complex, and interacts with the preribosome. As a member of the UTP14 family, UTP14a is also involved in ribosome biogenesis and 18S rRNA synthesis . Some studies found that the expression of UTP14a is upregulated in human hepatocellular carcinoma and colorectal carcinoma, and it may be associated with poor prognosis …”

“…As a member of the UTP14 family, UTP14a is also involved in ribosome biogenesis and 18S rRNA synthesis. 7,8 Some studies found that the expression of UTP14a is upregulated in human hepatocellular carcinoma and colorectal carcinoma, and it may be associated with poor prognosis. 12,13 In this study, UTP14a was positively expressed in 88.1% of the ESCC samples, and 30% of CDM samples exhibited positive expression; Western blot analysis further confirmed that the protein levels of UTP14a were significantly higher in ESCC.…”

“…UTP14a is an essential component of a large ribonucleoprotein complex bound to the U3 small nucleolar RNA. It plays a key role in ribosome biogenesis and 18S rRNA synthesis . Hu et al .…”

“…It plays a key role in ribosome biogenesis and 18S rRNA synthesis. 7,8 Hu et al 9 reported that UTP14a is overexpressed in various kinds of tumor cell lines, leading to cell cycle arrest and apoptosis through p53 protein degradation, and moreover, ectopically expressed UTP14a could protect tumor cells from chemotherapeutic drug-induced apoptosis. 10 However, the transcription of UTP14a is not regulated by p53.…”