Heat-shock Increases RpoD Dependent β-galactosidase Activity in the Escherichia coli Strains BD792 and B23

RpoD is a primary sigma factor in Escherichia coli responsible for expression of lacZ.  RpoS is an alternative sigma factor that is upregulated under environmental stress. RpoS competitively binds to RNA polymerase core proteins, displacing RpoD so that stress-related genes can be transcribed instead. This causes a decreased expression of RpoD-dependent genes. There have been contradictory findings on the relationship between heat-induced stress and RpoS concentrations in Escherichia coli.  One study found that RpoS concentrations decrease upon heat-induced stress, while another found the opposite. To address this issue, we have tested whether heat-shock causes a decrease in RpoS levels and a concomitant increase in RpoD-dependent gene expression, indirectly using a lacZ reporter system. An increase in RpoD-dependent lacZ gene expression would indicate a decrease in RpoS activity. Escherichia coli wild-type B23 and rpoS- BD792 strains were subjected to heat-shock treatment and β-galactosidase activities were measured using Miller assays as indication of lacZ expression. In the wild-type strain, β-galactosidase activity significantly increased upon heat-shock. Similarly, β-galactosidase activity significantly increased in the rpoS- strain, but the amount of β-galactosidase activity was significantly higher than that of the wild-type strain upon 15 and 60 minutes of heat-shock. Our data show that heat-shock increases the levels of RpoD-dependent β-galactosidase activity, suggesting a decrease in RpoS activity. Since an increase in β-galactosidase activity was also observed in the rpoS- strain upon heat shock, our data imply that there may be another protein involved in regulating RpoD-dependent β-galactosidase activity.