As the gateway to translation, ribosome biogenesis is a core cellular process that is highly efficient, accurate and regulated. This is made possible in part by a suite of ancillary proteins with diverse but poorly understood functions. One such factor, the Escherichia coli GTPase YjeQ, is suspected of playing a critical role in the assembly of the 30S ribosomal subunit. Here we demonstrate that the absence of this factor in vivo leads to an accumulation of a late-stage immature 30S subunit species. While these precursors lack several ribosomal proteins and feature a number of conformational abnormalities, they are competent for maturation, suggesting that they represent an assembly intermediate. We further demonstrate that YjeQ accelerates the maturation of these precursors in vivo. In addition, we explore the role of YjeQ through genetic interaction studies and substantiate a functional connection with the putative assembly factor RbfA.
A linear correlation between growth rate and ribosomal content has been observed for multiple wild-type microbes. We have examined this relationship in the ΔyjeQ strain and found there to be a significant increase in the total cellular ribosomal material in comparison to the wild-type. This phenotype is not wholly exclusive to perturbations in biogenesis. Indeed, linear correlations and elevated levels of ribosomal content are also observed for several translation mutants. The degree of elevation, however, is marginal in comparison to that seen in the biogenesis mutant. Our work explores this phenomenon and the possibility of exploiting it to identify and further characterize perturbations in the ribosome assembly process. / Thesis / Doctor of Philosophy (PhD) / In all cells, translation is carried out by ribosomes, large molecules that mediate the interpretation of the genetic code. These cellular interpreters are absolutely required for protein synthesis in bacteria and thus, are necessary for life. Like proteins, ribosomes themselves must also be synthesized, a process known as ribosome biogenesis. The ribosome consists of myriad RNA and protein components and is perhaps the single most complex machine in cells. Nevertheless, cells can build these enormous molecules in less than two minutes. This is made possible by a team of helper proteins, such as the bacterial assembly factor YjeQ. The function of this protein has evaded researchers, but there is growing evidence that it facilitates a key stage in the assembly process. Our work provides new detail into how this protein influences ribosome biogenesis, and how this in turn affects the overall health and proliferation of bacterial cells.
| Identifer | oai:union.ndltd.org:mcmaster.ca/oai:macsphere.mcmaster.ca:11375/18157 |
| Date | January 2015 |
| Creators | Stewart, Geordie |
| Contributors | Brown, Eric, Biochemistry and Biomedical Sciences |
| Source Sets | McMaster University |
| Language | English |
| Detected Language | English |
| Type | Thesis |
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