Study of the sRNA RsmY involved in the genetic regulation of the synthesis of alginate and alkyl resorcinols in Azotobacter vinelandii.

Azotobacter vineladii is a Gram-negative bacterium that produces alginate and poly-hydroxybutyrate (PHB), two polymers of biotechnological interest. This bacterium has the ability to form desiccation-resistant cysts. In the cyst the membrane phospholipids are replaced with a family of phenolic lipids called alkylresorcinols (ARs). The alginate, PHB, and ARs are controlled by the GacS/A two-component system and the small regulatory RNA (sRNA) RsmZ1, belonging to the Rsm (Csr) regulatory system. The Rsm (Csr) systems usually possess two or more sRNAs, in this regard A. vinelandii is the bacterium with the highest number of rsm-sRNAs. Originally, the presence of two sRNAs of the RsmY family (RsmY1 and RsmY2) was reported, but in a subsequent work it was suggested that they conformed to a single sRNA. In this work we provide genetic evidence confirming that rsmY1 and rsmY2 constitute a single gene. Also, it was established that rsmY mutation decreased alginate and ARs production, but did not affect the PHB synthesis. Transcriptional studies showed that rsmY has its higher expression during the stationary growth phase, and in the absence of RsmZ1, rsmY increases its transcription. Interestingly, rsmY expression was influenced by the carbon source, but its expression did not correlate with alginate production.

Routes of phosphoryl group transfer during signal transmission and signal decay in the dimeric sensor histidine kinase ArcB.

The Arc (anoxic redox control) two-component system of Escherichia coli, comprising ArcA as the response regulator and ArcB as the sensor histidine kinase, modulates the expression of numerous genes in response to respiratory growth conditions. Under reducing growth conditions, ArcB autophosphorylates at the expense of ATP, and transphosphorylates ArcA via a His292 → Asp576 → His717 → Asp54 phosphorelay, whereas under oxidizing growth conditions, ArcB catalyzes the dephosphorylation of ArcA-P by a reverse Asp54 → His717 → Asp576 → Pi phosphorelay. However, the exact phosphoryl group transfer routes and the molecular mechanisms determining their directions are unclear. Here, we show that, during signal propagation, the His292 → Asp576 and Asp576 → His717 phosphoryl group transfers within ArcB dimers occur intra- and intermolecularly, respectively. Moreover, we report that, during signal decay, the phosphoryl group transfer from His717 to Asp576 takes place intramolecularly. In conclusion, we present a mechanism that dictates the direction of the phosphoryl group transfer within ArcB dimers and that enables the discrimination of the kinase and phosphatase activities of ArcB.