Despite their name, a number of the cold shock proteins are expressed during
normal growth, and not just during cold shock, in several species. The function of these
constitutively expressed CspA paralogues is unclear. In Salmonella Typhimurium (a
major worldwide cause of gastrointestinal disease) they have been linked to various
stress responses and the establishment of virulence. Study of the cold shock proteins as
gene regulators is therefore of great interest, and they also have potential as targets for
antimicrobial development.
CspE in Salmonella Typhimurium is constitutively expressed during normal
growth. In order to determine its function, attempts were made to identify the
interactions it forms with other cellular proteins. Initially, a proteomic investigation
attempted to identify proteins which complex with CspE by in vivo cross-linking and
affinity purification followed by mass spectrometry. Although no defined complex was
consistently identified, the results suggested a handful of proteins which might interact
with CspE in a weak or transient manner. These proteins included many from the
nucleoid and ribosomal entry site, hinting at CspE’s cellular localisation.
In order to investigate these transient interactions, a bacterial two-hybrid
system was employed. Interactions between CspE and HupA, a nucleoid protein
identified in the proteomic analysis, were probed, as were interactions between CspE
and CsdA, an RNA helicase thought to function co-operatively with CspE. The twohybrid
system also allowed investigation of CspE dimerisation, which has been
reported in vitro but not investigated in vivo until this study. CspE appears not to
interact significantly with either HupA, CsdA, or itself at 37oC.
Finally in a further attempt to identify interactions of CspE, a genomic library
was created to test CspE interactions by two-hybrid assay with random peptides
derived from the whole Salmonella genome. The library was successfully created and
screened for evidence of interaction, and revealed an association between CspE and a
transcriptional repressor, DeoT. DeoT is a repressor of several genes for catabolic
processes, suggesting a role for CspE in the regulation of central metabolism. The
findings of this work present a number of novel discoveries and several interesting
opportunities for further studies