Function of CikA in the cyanobacterial circadian system: the pseudo-receiver domain of CikA regulates the circadian input pathway
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The circadian input kinase gene (cikA) was first identified from a Tn5 mutant of Synechococcus elongatus PCC 7942. A cikA null strain shows a striking phenotype related to circadian gene regulation: all sampled loci show a shortened circadian period and reduced amplitude of oscillation and a failure to exhibit a wild-type resetting of the phase of the rhythm after an environmental signal. This global defect in response to the environment suggests a key role for CikA in the circadian input pathways. Bioinformatics results classify CikA as a divergent member of the bacteriophytochrome family, suggesting a role in light signal transduction. In vitro analysis previously showed that CikA is a bona fide histidine protein kinase (HPK), and its kinase activity is regulated by the presence of other domains. Its own pseudo-receiver (PsR) domain is not the cognate receiver domain of its kinase HPK domain, and its GAF domain does not likely bind a bilin chromophore as do photoreceptive phytochromes. Recent results suggested that CikA may function as a redox-sensor. In this study, we examined the function of each domain of CikA using different mutant cikA alleles, and determined their phenotypes with respect to complementation of a null mutant and overexpression in both wild type and cikA null strains. All domains except the featureless N-terminus were required for CikA function. Overexpression of all mutant alleles that encoded the PsR domain, whether or not the HPK was functional, caused a dominant arrhythmia phenotype. In the absence of PsR, overexpressed variants did not cause arrhythmia, but affected the amplitude and period of oscillation. The results suggest a model in which the PsR domain regulates kinase activity and mediates interaction with other input pathway components to allow CikA to reach the correct cellular position to fulfill its function. Cellular localization assays showed CikA can interact with a complex and showed a polar localization pattern, whereas its variant without PsR showed uniform distribution in the cell. In summary, CikA is an autoregulated kinase in which the PsR domain regulates activity of the HPK domain and also serves as an interaction module to lead the CikA to a specific cellular position.