In the present study, the cold shock response of <i>S. typhimurium</i> was investigated using a small custom made DNA microarray which allowed the transcriptional profiling of <i>cspA</i> and its five other paralogous genes (<i>cspB, cspC, cspD, cspE</i> and <i>cspH</i>) after a temperature downshift to 10°C or 4°C from optimal growth at 37°C. Changes in low temperature expression of the six genes were monitored in exponential and stationary phase cells. Global transcriptional analysis of <i>S. typhimurium</i> was also performed using genomic microarrays which could measure gene expression for 97.5% of all the open reading frames in the <i>S. typhimurium </i>LT2 genome. This allowed expression data from other cold induced genes as well as the <i>cspA</i> paralogues to be examined under the same conditions used for the smaller array. The main findings from the array experiments showed that <i>cspA</i> and <i>cspB</i> appeared to be the most highly induced of the paralogues genes upon cold shock, with significant levels of transcription increase also being observed for <i>cspC </i>and <i>cspE</i> at low temperature and at 37°C. The <i>cspH</i> gene was confirmed as being strictly cold-induced whilst, in contrast, <i>cspD</i> only exhibited significant levels of induction at 37°C. The genomic microarray studies detected cold shock induction of many other genes involved in a variety of cellular activities linked to the maintenance of cell envelope integrity, DNA structure and pathogenicity, among others. The overall levels and timing of global transcription up/down regulation was observed to be influenced by growth phase and the degree to which <i>Salmonella </i>cells were cold-shocked, with gene induction also being detected at temperatures below the minimum for growth (4°C). these findings suggested that the cold shock response of this pathogen can still adapt the cell for survival under such conditions until a more favourable temperature arises for cell division. Extensive mutational analysis of the <i>csp</i> genes in <i>S. typhimurium </i> indicated that two strains, harbouring only a functional <i>cspA</i> gene or, more interestingly, the constitutively expressed <i>cspE</i> gene, were fully capable of adapting the cell for growth at 10°C and for colony formation at 15°C, to a level comparable to that of the wild-type strain. A mutant strain with no functional <i>csp</i> genes was incapable of growth at temperatures below 20°C and exhibited a reduction in viability with extended cold incubation. Other phenotypic tests on <i>csp </i>mutants seemed to indicate that CspC and CspE were capable of directly or indirectly enhancing the viability of cells to freeze/thaw treatment and that CspE may also play a role in enhancing the survival of stationary phase <i>Salmonella </i>cells against oxidative stress.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:652769 |
| Date | January 2005 |
| Creators | Hutchinson, Ian Wesley |
| Publisher | University of Edinburgh |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://hdl.handle.net/1842/15077 |
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