Modern global analysis tools have given an unprecedented insight into the genetic circuitry of living cells. In contrast, biochemical networks so far have been poorly defined. Now researchers at the Biozentrum of the University of Basel have uncovered a fundamentally novel regulatory interaction among signaling proteins. This discovery will help to identify some of the so far hidden connectivity of the cell’s regulatory makeup. The research results have recently been published in Cell.
Two-component signal transduction systems are ubiquitous in bacteria, lower eukaryotes, and plants where they control a wide range of cellular functions including cell differentiation, stress response, and virulence. Until recently, two-component systems were believed to have a simple architecture with a kinase and a response regulator being involved in a linear, unidirectional processing of information with limited interconnections between different pathways. The sensor kinase receives an environmental stimulus and via phosphorylation passes this information on to the response regulator, which ultimately brings about a cellular response. Response regulators consist of a conserved receiver domain, which accepts the phosphoryl group from the kinase, and in turn activates the second, so-called output domain. Via the output domain activated response regulators directly control gene expression or mediate the production of secondary signaling molecules.
Dividing Caulobacter crescentus cells attached to a solid substrate via their stalk and holdfast structures. © Martin Oeggerli, www.Micronaut.ch 2008, kindly supported by FHNW, Life Sciences, Muttenz.
In the last ten years, the genomic revolution provided researchers with the full genetic information of hundreds of genomes of bacteria and higher organisms exposing two-component systems as one of the largest families of signaling proteins and the predominant signal transduction device in bacteria. Moreover, by analyzing the cell’s genetic fingerprint researchers made a puzzling observation with respect the to response regulator family. Nearly one out of five of these signaling proteins consist only of a receiver domain and completely lack the output domain necessary to elicit a cellular response. While the abundance of these single domain response regulators remained enigmatic, the new study from the Biozentrum proposes that some of these truncated proteins can play a key role in interconnecting and modulating different signaling pathways.
The experiments that led to the discovery of this novel signaling mechanism were carried out with Caulobacter crescentus, an aquatic bacterium with a characteristic asymmetric life cycle. Unlike most other bacteria, Caulobacter divides asymmetrically producing two daughter cells with distinct morphologies and developmental programs, a motile swarmer and a sessile stalked cell. One of the main interests of Urs Jenal and his colleagues is to understand how Caulobacter cells oscillate between two different cellular states and how these cells very quickly and robustly change their identity during their division cycle. The Jenal group recently identified a novel signaling molecule, cyclic di-GMP, as a key cell fate determinator during Caulobacter development. This led them to ask how the concentration of this signaling compound changes during the division cycle.
As it turned out, c-di-GMP is produced by the PleD response regulator upon activation by phosphorylation. Two kinases, DivJ and PleC, are responsible for PleD activation during the differentiation of the Caulobacter swarmer cell into a stalked cell. DivJ and PleC localize to opposite poles of the dividing asymmetric cell and by differentially partitioning into the newly formed cellular compartments, determine the individual programs of the daughter progeny. The key to understanding how these kinases control PleD activity and by that contribute to cell fate determination was when the researchers discovered that PleC functions as kinase in one cell type, but takes on the reverse phosphatase activity in the other cell type. While this was a breakthrough discovery, it did not explain how during development PleC switches from being a phosphatase to a kinase and back.
This is when DivK, a single domain response regulator, entered the picture. Several observations had already suggested a functional link between DivK and the PleC-DivJ-PleD pathway. Surprisingly, DivK dynamically localizes to both the PleC and DivJ occupied cell poles in stalked cells, but is delocalized in swarmer cells. Similar to the activity of PleD response regulator, the dynamic behavior of DivK depends on its phosphorylation by DivJ and PleC. Together, these observations raised an important question: why would this small protein move to the cell poles and what is its molecular role at this subcellular site? The experiments carried out in the Jenal lab demonstrated that DivK acts as an allosteric activator of both DivJ and PleC kinase activity. Dynamic localization of activated DivK to the cell poles switches PleC from its phosphatase into the kinase mode and at the same time stimulates the DivJ kinase. These posttranslational feedback loops very quickly and robustly lead to a change of the developmental program from a motile swarmer to a sessile stalked cell.
The results clearly showed that members of this family of response regulators not only receive information from the sensor kinase, but can also signal back to one or several kinases and by that can bring about very effective positive feedback and signal amplification. S uch response regulators can thus facilitate crosstalk and long-range communication among members of the two-component phosphorylation network. As such they might represent connecting nodes of the regulatory wiring of living cells.
Systems Biology strives after a quantitative description of the complex regulatory and metabolic interactions by unraveling of the cell's regulatory network and the connectivity of its individual components. The discovery of this novel control element sheds a new light on the complexity and versatility of cellular networks. A systematic analysis of the cell’s network will eventually allow an accurate prediction of cellular behavior and, following a synthetic biology approach, will lead to reconstruction of living systems dedicated to specialized tasks.
Source Article: Allosteric Regulation of Histidine Kinases by Their Cognate Response Regulator Determines Cell Fate. Ralf Paul, Tina Jaeger, Sören Abel, Irene Wiederkehr, Marc Folcher, Emanuele G. Biondi, Michael T. Laub, and Urs Jenal. Cell, Vol 133, 452-461, 02 May 2008.