Functional Analysis Of A Novel Family Of Protein Kinases

The filamentous cyanobacterium Anabaena sp. Strain PCC 7120 can fix N2 whencombined nitrogen is not available in the growth medium. It has a family of 13 genesencoding proteins with both a Ser/Thr kinase domain at N-teminal region and a Hiskinase domain at C-terminal region. So far, the function of these enzymes is unknown.Two of them are encoded by pkn41 (alr0709) and pkn42 (alr0710). This study focuses onthe function of these two genes in signal transduction. The results of RNA Dot Blot andRT-PCR show that the expression of pkn41 and pkn42 is induced by iron deprivationirrespective of the nature of the nitrogen source. pkn41 and pkn42 are separated by only72 bp on the chromosome, and our results indicate that they are cotranscribed underiron-limited conditions. Mutants in which either pkn41, pkn42, or both are inactivatedgrow similarly to the wild type under normal conditions, but their growth is impairedeither in the presence of an iron chelator, 2,2'-diopyridyl, or under conditions of nitrogenfixation and iron limitation, two situations where the demand for iron is particularlystrong. Consistent with these results, these mutants display lower intracellular ironcontent than the wild type and a higher level of expression for nifJ and nifJ-like, whichencode pyruvate: ferredoxin oxidoreductases. Both nifJ and nifJ-like are known to beinduced by iron limitation. The pigment contents of the mutants are also lower than wildtype under iron-limited conditions and the activity of catalase of the mutants are higherthan that of wild type under iron-limited conditions. These results are consistent with thefacts that the intracellular iron contents of the mutants are lower than that of WT. NtcA, aglobal regulatory factor for different metabolic pathways, binds to the putative promoterregion of pkn41, and the induction of pkn41 in response to iron limitation no longeroccurs in an ntcA mutant. Our results suggest that ntcA not only regulates the expressionof genes involved in nitrogen and carbon metabolism but also coordinates iron acquisitionand nitrogen metabolism by activating the expression of pkn41 and pkn42.