An RpaA-Dependent Sigma Factor Cascade Sets the Timing of Circadian Transcriptional Rhythms in Synechococcus elongatus.

The circadian clock of the cyanobacterium Synechococcus elongatus PCC 7942 drives oscillations in global mRNA abundances with 24-hr periodicity under constant light conditions. The circadian clock-regulated transcription factor RpaA controls the timing of circadian gene expression, but the mechanisms underlying this control are not well understood. Here, we show that four RpaA-dependent sigma factors-RpoD2, RpoD6, RpoD5, and SigF2-are sequentially activated downstream of active RpaA and are required for proper expression of circadian mRNAs. By measuring global gene expression in strains modified to individually lack rpoD2, rpoD6, rpoD5, and sigF2, we identify how expression of circadian mRNAs, including sigma factor mRNAs, is altered in the absence of each sigma factor. Broadly, our findings suggest that a single transcription factor, RpaA, is sufficient to generate complex circadian expression patterns in part by regulating an interdependent sigma factor cascade.

Controlling the release of peptide antimicrobial agents from surfaces.

Medical conditions are often exacerbated by the onset of infection caused by hospital dwelling bacteria such as Staphylococcus aureus. Antibiotics taken orally or intravenously can require large and frequent doses, further contributing to the sharp rise in resistant bacteria observed over the past several decades. These existing antibiotics are also often ineffective in preventing biofilm formation, a common cause of medical device failure. Local delivery of new therapeutic agents that do not allow bacterial resistance to occur, such as antimicrobial peptides, could alleviate many of the problems associated with current antibacterial treatments. By taking advantage of the versatility of layer-by-layer assembly of polymer thin films, ponericin G1, an antimicrobial peptide known to be highly active against S. aureus, was incorporated into a hydrolytically degradable polyelectrolyte multilayer film. Several film architectures were examined to obtain various drug loadings that ranged from 20 to 150 microg/cm2. Release was observed over approximately ten days, with varying release profiles, including burst as well as linear release. Results indicated that film-released peptide did not suffer any loss in activity against S. aureus and was able to inhibit bacteria attachment, a necessary step in preventing biofilm formation. Additionally, all films were found to be biocompatible with the relevant wound healing cells, NIH 3T3 fibroblasts and human umbilical vein endothelial cells. These films provide the level of control over drug loading and release kinetics required in medically relevant applications including coatings for implant materials and bandages, while eliminating susceptibility to bacterial resistance.