Global Systems-Level Analysis of Hfq and SmpB Deletion Mutants in Salmonella: Implications for Virulence and Global Protein Translation

Using sample-matched transcriptomics and proteomics measurements it is now possible to begin to understand the impact of post-transcriptional regulatory programs in Enterobacteria. In bacteria post-transcriptional regulation is mediated by relatively few identified RNA-binding protein factors including CsrA, Hfq and SmpB. A mutation in any one of these three genes, csrA, hfq, and smpB, in Salmonella is attenuated for mouse virulence and unable to survive in macrophages. CsrA has a clearly defined specificity based on binding to a specific mRNA sequence to inhibit translation. However, the proteins regulated by Hfq and SmpB are not as clearly defined. Previous work identified proteins regulated by hfq using purification of the RNA-protein complex with direct sequencing of the bound RNAs and found binding to a surprisingly large number of transcripts. In this report we have used global proteomics to directly identify proteins regulated by Hfq or SmpB by comparing protein abundance in the parent and isogenic hfq or smpB mutant. From these same samples we also prepared RNA for microarray analysis to determine if alteration of protein expression was mediated post-transcriptionally. Samples were analyzed from bacteria grown under four different conditions; two laboratory conditions and two that are thought to mimic the intracellular environment. We show that mutants of hfq and smpB directly or indirectly modulate at least 20% and 4% of all possible Salmonella proteins, respectively, with limited correlation between transcription and protein expression. These proteins represent a broad spectrum of Salmonella proteins required for many biological processes including host cell invasion, motility, central metabolism, LPS biosynthesis, two-component regulatory systems, and fatty acid metabolism. Our results represent one of the first global analyses of post-transcriptional regulons in any organism and suggest that regulation at the translational level is widespread and plays an important role in virulence regulation and environmental adaptation for Salmonella

In vivo evaluation of modified gum karaya as a carrier for improving the oral bioavailability of a poorly water-soluble drug, nimodipine

This work examines the influence of modified gum karaya (MGK) on the oral bioavailability of a poorly water-soluble drug, nimodipine (NM), in comparison with that of gum karaya (GK). A cogrinding method was selected to prepare mixtures of NM and GK or MGK in a 1:9 ratio (NM:GK/MGK). Differential scanning calorimetry (DSC), Fourier transmission infrared (FT-IR) spectroscopy, X-ray diffraction (XRD), solubility studies, and in vitro release studies were performed to characterize the properties of the cogrinding mixtures. No drug-carrier interactions were found, as confirmed by DSC and FT-IR studies. The XRD study revealed that the crystallinity of NM was identical in both the cogrinding mixtures and was decreased when compared to that of physical mixtures or pure NM. The in vitro release rate of NM from both cogrinding mixtures was significantly higher than that of physical mixtures or pure NM. The in vivo study revealed that the bioavailability of NM from pure drug was significantly lower when compared to the cogrinding mixtures. The oral bioavailability was found to be NM powder < cogrinding mixtures of NM and GK < cogrinding mixtures of NM and MGK < NM solution. It can be inferred from the above results that MGK, an economical carrier, could be used for the dissolution enhancement of NM