Poster abstracts

Poster number 41 submitted by Jason Talkish

Does the DEAD-box protein Drs1 drive the chronology of ribosome assembly?

Jason Talkish (Department of Biological Sciences, Carnegie Mellon University), Jelena Jakovljevic (Department of Biological Sciences, Carnegie Mellon University), Jingyu Zhang (Department of Biological Sciences, Carnegie Mellon University), Philip C. Andrews (Department of Biological Chemistry, Michigan Proteome Consortium, University of Michigan), Janine Maddock (Department of Molecular, Cellular, and Developmental Biology, University of Michigan), John R. Strahler (Department of Biological Chemistry, Michigan Proteome Consortium, University of Michigan)

Abstract:
Ribosome assembly requires 180 assembly factor proteins not found in mature ribosomes. These assembly factors are thought to enter and exit the ribosome biogenesis pathway at various times, but a largely unanswered question is what drives the ordered assembly of ribosomes. One hypothesis is that ATPases regulate this process using nucleotide hydrolysis to cause structural rearrangements within the preribosome, driving maturation forward. To investigate the mechanisms that govern ordered assembly, we are studying the DEAD-box protein Drs1 in Saccharomyces cerevisiae. We believe that Drs1, through ATP hydrolysis and helicase activity, functions by either removing early assembly factors from preribosomes or causing a structural rearrangement, to create an environment in which later assembly factors can bind, thus regulating the timing of ribosome biogenesis. We show that Drs1 is required for three steps of pre-rRNA processing. Depletion of Drs1 blocks processing of 27SA₂ and 27SA₃ pre-rRNAs and traps preribosomes in the nucleolus. Partial loss of function mutants however block processing of 27SB pre-rRNA. While it is known that some assembly factors enter and exit the assembly pathway early and others enter and exit late, we show that depletion of Drs1 causes the formation of a preribosome containing both “early” and “late” assembly factors. In the absence of Drs1, “late” assembly factors copurify with increased amounts of “early” assembly factors and pre-rRNA intermediates. One model is that depletion of Drs1 causes “late” assembly factors to enter the nucleolus and associate with preribosomes prematurely. Another scenario is that preribosomes are very dynamic early in assembly, possibly a result of ATP hydrolysis by Drs1 and other ATPases, causing “late” proteins to be weakly associated with preribosomes. Depletion of Drs1 could cause these early particles to become less dynamic and more stable, therefore allowing detection of these “late” proteins.

References:
Henras, A.K., Soudet, J., Gerus, M., Lebaron, S., Caizergues-Ferrer, M., Mougin, A., and Y. Henry. 2008. The post-transcriptional steps of eukaryotic ribosome biogenesis. Cell. Mol. Life. Sci. 65:2334-2359.

Ripmaster, T.L., G.P. Vaughn, and J.L. Woolford, Jr. 1992. A putative ATP-dependent RNA helicase involved in Saccharomyces cerevisiae ribosome assembly. Proc. Natl. Acad. Sci. USA 89:11131-11135.

Adams, C.C., J. Jakovljevic, J. Roman, P. Harnpicharnchai, and J.L. Woolford, Jr. 2002. Saccharomyces cerevisiae nucleolar protein Nop7 is necessary for biogenesis of 60S ribosomal subunits. RNA 8:150-165.

Keywords: Ribosome, ATPase, preribosomal RNA processing