The FurA regulon in Anabaena sp. PCC 7120: in silico prediction and experimental validation of novel target genes

Abstract

In the filamentous cyanobacterium Anabaena sp. PCC 7120, the ferric uptake regulator FurA functions as a global transcriptional regulator. Despite several analyses have focused on elucidating the FurA-regulatory network, the number of target genes described for this essential transcription factor is limited to a handful of examples. In this article, we combine an in silico genome-wide predictive approach with experimental determinations to better define the FurA regulon. Predicted FurA-binding sites were identified upstream of 215 genes belonging to diverse functional categories including iron homeostasis, photosynthesis and respiration, heterocyst differentiation, oxidative stress defence and light-dependent signal transduction mechanisms, among others. The probabilistic model proved to be effective at discerning FurA boxes from non-cognate sequences, while subsequent electrophoretic mobility shift assay experiments confirmed the in vitro specific binding of FurA to at least 20 selected predicted targets. Gene-expression analyses further supported the dual role of FurA as transcriptional modulator that can act both as repressor and as activator. In either role, the in vitro affinity of the protein to its target sequences is strongly dependent on metal co-regulator and reducing conditions, suggesting that FurA couples in vivo iron homeostasis and the response to oxidative stress to major physiological processes in cyanobacteria.

Ferric Uptake Regulator (FUR) protein: properties and implications in cyanobacteria

“…PCC 7120. These data suggested that Fur controls the expression of genes involved in the signal transduction cascade (Gonzalez et al 2014). FurA binding site was also predicted in the upstream region of the promoter of the transposase-encoding gene all4465 in Anabaena sp.…”

“…In Anabaena sp. PCC 7120, FurA differentially activates the transcription of psaK, amt4, hetC, alr1728, patA and asr1734, depending on the presence of the metal coregulator and a reducing environment (Gonzalez et al 2014). Gonzalez and coworkers also demonstrated the dual role of Fur protein, i.e., as both activator and repressor in the same metabolism, as observed in the regulation of tetrapyrrole biosynthesis .…”

“…FurA boxes were also located within bidirectional promoters of genes involved in several metabolisms, for example all0396-schT, all2624-alr2625, all2586-alr2587, alr2594-alr2595 involved in iron metabolism, all0949-coxB, ccmKndhF involved in photosynthesis and respiration, and znuAoprB, hupL3-xisC, and nifD3-xisA involved in other important metabolisms in Anabaena sp. PCC 7120 (Gonzalez et al 2014). These binding sites play a role in the simultaneous regulation of genes with overlapping promoters (Hunt et al 1994;Christoffersen et al 2001;Lavrrar et al 2003).…”

“…They bind to unique sequences (Fur boxes) present in the upstream promoter region of the iron responsive genes, using ferrous iron as a co-regulator (Fillat 2014). However, it has now been well established that the mechanism of regulation by Fur involves a much higher level of complexity, as Fur controls several processes, such as nitrogen metabolism, photosynthesis and respiration, intermediary metabolism (Gonzalez et al 2014), virulence factor production (Delany et al 2004;Zhou et al 2006;Gao et al 2008), and is involved in the defense against different type of stresses (Touati 2000;Cornelis et al 2011) (Fig. 1).…”

Cysteine Mutational Studies Provide Insight into a Thiol-Based Redox Switch Mechanism of Metal and DNA Binding in FurA fromAnabaenasp. PCC 7120

“…PCC 7120, three different Fur proteins have been identified: FurA (all1691), FurB (all2473), and FurC (alr0957) (21). FurA is a multifaceted protein that controls iron homeostasis and modulates an extensive regulon involved in a variety of cell functions (13,14,15). It also fulfills the features of a heme sensor protein whose interaction with this cofactor negatively affects its DNA binding ability in vitro (23,41).…”