Lately, new classes of PII signaling proteins have been identified and renamed as PII-like proteins. The PII-like proteins are structurally similar and clearly related to canonical PII proteins, but lacking canonical PII PROSITE signature sequences and their functions are completely unknown. Bioinformatics and structural genomics approaches expanded the members of the PII superfamily and proposed that the PII-like proteins represent an even more widespread family of regulators than classical PII proteins. PII-like proteins are universally distributed across all domains of life, including bacteria, archaea, plants, protozoa, and animals, proposing a fundamental role in cell physiology.

In our laboratory, we are exploring the molecular, structural and cellular functions of several PII-like proteins; For example: 

1) PII-like protein SbtB is linking the second messenger cAMP to carbon sensation

To cope with low atmospheric CO2concentrations that developed over the course of evolution, cyanobacteria evolved a CO2-concentrating mechanism (CCM), which elevates CO2levels in the vicinity of RubisCO, the key enzyme of CO2fixation. Recently, we characterized a unique component of cyanobacterial CCM, the PII-like signaling protein SbtB, which provided new insights into carbon sensing in cyanobacteria. We found that SbtB senses a variety of adenosine nucleotides, including the second messenger cAMP. Moreover, physiological and biochemical studies indicated importance of SbtB for acclimation to varying carbon regimes in the ecological niches of cyanobacteria. Therefore, PII-like protein SbtB represents a new principle of carbon sensing via second messengercAMP binding, to regulate the cyanobacterial carbon concentrating mechanisms. However, the whole regulatory network of SbtB remains undiscovered.

2) Evolution of PII signaling in higher plants

The ancestor of Archaeplastida inherited its PII signal transduction protein from an ancestral cyanobacterial endosymbiont. Over the course of evolution, plant PII proteins acquired a glutamine-sensing C-terminal extension, called Q-loop, subsequently present in all Chloroplastida PII proteins.

Over the past years, we systematically investigated different PII proteins from various algal strains (red, green and nonphotosynthetic algae) and from higher plant (Oryza sativa and Arabidopsis thaliana) with respect to their sensory and regulatory properties. The highly conserved role of the controlling of nitrogen metabolism via regulating enzyme of arginine biosynthesis, N-acetyl-l-glutamate kinase (NAGK), as a main PII-interactor has been demonstrated across oxygenic phototrophs of cyanobacteria and Archaeplastida. A hallmark of PII-NAGK interaction in the non-photosynthetic alga Polytomella parvais their co-evolution towards a stable hetero-oligomeric complex enzyme, irrespective of effector molecules. In addition, the PII signaling system of red algae has been identified as an evolutionary intermediate between that of Cyanobacteria and Chloroplastida.

3) PII-like protein CutA is an enigmatic protein uninvolved in heavy metal sensation

The PII-like protein CutA is annotated as being involved in heavy metal tolerance. However, the precise cellular function of CutA remains unclear. Our bioinformatic analysis reveals that CutA proteins are universally distributed across all domains of life. However, we were unable to find any involvement of cyanobacterial and E. coli CutA in metal tolerance. Therefore, we concluded that CutA is involved in yet to be identified sensory function other than metal sensing. Furthermore, we resolved high-resolution CutA structures from cyanobacterial Nostoc sp. and Synechococcus elongatus. Intriguingly, the clefts between the CutA subunits, corresponding to the binding pockets of canonical PII proteins, are formed by conserved aromatic and charged residues, suggesting a conserved binding/signaling function for CutA proteins. However, the natural CutA ligands remain to be identified.

Calcium (Ca²⁺) is one of the most important second messengers, and is known to regulate metabolic and/or differentiation processes in eukaryotic cells. Much less is known about the role of Ca²⁺ in prokaryotes and especially cyanobacteria and how they sense Ca²⁺. Therefore, we aim to uncover the role of Ca²⁺ signaling in the multicellular lifestyle of the diazotrophic cyanobacteria. Recently within the frame of an international cooperation, we identified a novel Ca²⁺-Sensor protein (CSE protein). CSE protein is important for filament integrity, cell differentiation into heterocyst cells, N₂ fixation and the regulation of photosynthesis in diazotrophic multicellular cyanobacteria.

In addition, we are currently investigating the signaling roles of the cyclic di-nucleotides c-di-AMP and c-di-GMP in unicellular cyanobacteria. We identified several of novel c-di-AMP and c-di-GMP receptors as well as RNA-riboswitches, with putative roles in Carbon/Nitrogen metabolisms and photosynthetic day/night and/or phototaxis motility lifestyles. We are currently exploring the regulatory functions of those new c-di-AMP and c-di-GMP receptors.