eprintid: 31006
rev_number: 16
eprint_status: archive
userid: 6395
dir: disk0/00/03/10/06
datestamp: 2022-01-05 09:48:39
lastmod: 2022-01-19 06:57:00
status_changed: 2022-01-05 09:48:39
type: doctoralThesis
metadata_visibility: show
creators_name: Lau, Benjamin
title: Transition of the 90S Pre-Ribosome into the Primordial pre-40S Subunit involving the RNA Exosome, Utp24 Endonuclease and Dhr1 Helicase
subjects: ddc-570
divisions: i-140001
adv_faculty: af-14
abstract: Ribosomes, which are the molecular machines that synthesize the proteins of the cell, consist of ribosomal RNA (rRNA) and ribosomal proteins. To produce these ribosomes, a large precursor rRNA molecule (pre-rRNA) is transcribed by RNA-polymerase I, which in yeast is processed to form the three mature rRNAs (18S, 25S and 5.8S rRNA) accompanied by concomitant incorporation of 79 ribosomal proteins. This complicated process of ribosome assembly and maturation is driven by numerous (~200) and highly conserved assembly factors (AFs). A number of AFs are enzymes such as helicases or exo- and endonucleases, which play a vital role in remodeling and processing of the pre-rRNA, thereby allowing distinct and often irreversible transitions in the long cascade of pre-ribosome assembly.
In my PhD thesis, I have investigated the 90S-to-pre-40S ribosome transition by biochemical, genetic and structural approaches, with the goal to gain mechanistic insight into this process. One important initial observation was the sequential AF shedding of the 90S pre-ribosome after cleavage of 5’-ETS at site A1, which challenged our common view of an en bloc release of the 5’-ETS particle. Subsequently, I followed the mechanism of Utp24 endonuclease driven pre-RNA cleavage at site A1 between the 5’-ETS and 18S rRNA, which turned out to be prerequisite for the 90S-to-pre-40S transition. Moreover, I was able to isolate a 90S-exosome super-complex, revealing how the nuclear RNA exosome is docked via its co-factor Mtr4 helicase to the 90S pre-ribosome at the base of the 5’ ETS helices H9-9’ that were already dislodged in the 90S exosome super particle. This finding suggested that Mtr4 channels the free 3’ end of 5’-ETS at site A0 into the exosome for RNA degradation, thereby acting as a key driver in following 90S-to-pre-40S transition. The last transition intermediate observed in the series of discovered pre-ribosomal particles was shown to be the primordial pre-40S, which however was still decorated with a few 90S factors. One such factor was the RNA helicase Dhr1, which was seen to be directly positioned at its U3 snoRNA substrate to unwind the final hybrid between U3 and the 5’-end of the 18S rRNA. Taken together, these findings from my PhD study revealed key steps of the 90S-to-pre-40S transition, both biochemically and structurally, thereby shining light on the mechanism of pre-rRNA processing and its coupling to remodeling during eukaryotic ribosome biogenesis.
date: 2021
id_scheme: DOI
id_number: 10.11588/heidok.00031006
ppn_swb: 1786338076
own_urn: urn:nbn:de:bsz:16-heidok-310062
date_accepted: 2021-12-06
advisor: HASH(0x55a99e46cbb0)
language: eng
bibsort: LAUBENJAMITRANSITION2021
full_text_status: public
place_of_pub: Heidelberg
citation:   Lau, Benjamin  (2021) Transition of the 90S Pre-Ribosome into the Primordial pre-40S Subunit involving the RNA Exosome, Utp24 Endonuclease and Dhr1 Helicase.  [Dissertation]     
document_url: https://archiv.ub.uni-heidelberg.de/volltextserver/31006/1/Benjamin_Lau_PhD_thesis.pdf