Bacterial genetics

tmRNA is a small stable RNA present in Eubacteria. Through a mechanism called
trans-translation, tmRNA mediates ribosome rescue and quality control of proteins and
mRNA. In this study, the Escherichia coli (E. coli) mutant lacking tmRNA was
demonstrated hypersensitive to oxidative stress. The role of tmRNA-mediated
surveillance mechanism in protecting E. coli cell under oxidative stress condition was
examined. The tmRNA-mediated tagged protein levels were elevated in cells under
oxidative stress condition, demonstrating the enhanced need for tmRNA under such
condition. Our results suggest that mRNA damage by oxidative stress may cause reduced
cell viability, and that tmRNA is required to rescue cells under such condition.
Furthermore, our observations showed that tmRNA is required for the optimal growth of
E. coli under normal aeration but not under anaerobic condition, suggesting that oxidation
ofmRNA is the major reason for requirement oftmRNA during normal aeration.

The purpose of this study was to determine the effect of bacterial lipopolysaccharide (LPS) on the expression of RP105 (CD180) in human B lymphocytes from normal, leukemic, and lupus patients. The percentage of cells that express RP105 on their surface increased following a 24 hour exposure to LPS. However, continued exposure for a total of four days resulted in a marked decrease in the expression of this receptor molecule. Human B cells were purified by a combination of density gradient and negative magnetic bead selection and maintained in culture with and without LPS. Enzyme linked immunoassay for the detection of anti-dsDNA antibodies following LPS treatment of isolated B cells was negative. The percentage of RP105 positive or negative B cells from lupus patients could not be accurately determined because too few B cells were available from these lymphopenic patients following negative selection.

RNA damage occurring under oxidative stress has been shown to cause RNA dysfunction and must be detrimental to cells and organisms. We propose that damaged RNA can be removed by specific RNA surveillance activities. In this work, we investigated the role of polynucleotide phosphorylase (PNPase), a 3'->5' exoribonuclease, in protecting the cells against oxidative stress and eliminating oxidatively-damaged RNA. Previously, it was reported that E. coli PNPase has a higher affinity to poly(8-oxoG:A). We further confirmed that E. coli PNPase can specifically bind to an oxidized RNA with a high affinity. An E. coli strain deficient in PNPase (pnp) is hypersensitive to hydrogen peroxide (H2O2). Importantly, the level of H2O2-induced RNA damage, measured by the content of 8-hydroxyguanosine, increases significantly in the pnp mutant cells. Consistent with the notion that PNPase plays a direct role in these processes, introduction of the pnp gene encoding E. coli PNPase can restore the viability and RNA oxidation level of the pnp mutant cells in response to H2O2 treatment. Interestingly, degradosome-association is not required for PNPase to protect cell against oxidative stress. PNPase is evolutionary conserved in most of organisms of all domains of life. The human polynucleotide phosphorylase (hPNPase) localizes mainly in mitochondria and plays pleiotropic roles in cell differentiation and has been previously shown to bind 8- oxoG-RNA with a high affinity. Here we show that similar to E. coli PNPase, hPNPase plays an indispensable role in protecting HeLa cells against oxidative stress. The viability in HeLa cell and 8-oxoG levels in RNA are inversely correlated in response to H2O2- treatment. After removal of oxidative challenge, the elevated level of 8-oxoG in RNA decreases, suggesting the existence of surveillance mechanism(s) for cleaning up oxidized RNA.