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There were many reports about the cyanobacterial PII protein (encoded by glnB). In enteric bacteria, PII is a central signal- transduction protein involved in transcriptional and post- translational regulation of nitrogen assimilation. The PII protein of Synechococcus sp. strain PCC 7942 is distinct from that of enteric bacteria in that it is subject to phosphorylation at a serine residue rather than to uridylylation at a tyrosine residue. It remains similar to PII of enteric bacteria in that its modification state is changed in response to the nitrogen status of the cell: the phosphorylation state of the PII trimer changes from a fully dephosphorylated state in ammonium-grown cells to a highly phosphorylated state in the cells subjected to nitrogen starvation. PII is dephosphorylated also when CO2 fixation is inhibited and hence is supposed to play a role in the interplay between inorganic carbon and nitrogen assimilation. While the modification of cyanobacterial PII in response to cellular nitrogen and carbon status is well established, the physiological role of the protein remains unclear.
The importance of PII in the post-translational regulation of nitrate assimilation was related by Hyun-Mi Lee (Institut Pasteur and CSIC). She found that nitrate assimilation is inhibited by ammonium in wild-type Synechococcus sp. strain PCC 7942, but it is insensitive to ammonium in a PII-null mutant (MP2). Introduction of the wild-type glnB gene into the MP2 mutant by transformation restored the ammonium-responsive inhibition of nitrate assimilation, suggesting that PII inhibits nitrate assimilation in its unphosphorylated form.
Cloning of glnB and characterization of PII from two cyanobacteria was reported: Synechocystis sp. strain PCC 6803 by Mario García-Domínguez (Universidad de Sevilla-CSIC) and Michael Hisbergues (CNRS, Marseille) and Nostoc punctiforme ATCC 29133 by Tom Hanson (University of California, Davis, now at Ohio State U.). PII was shown to be modified by phosphorylation in Synechocystis as in Synechococcus sp. strain PCC 7942 [Forchhammer & Tandeau de Marsac (1995) J Bacteriol 177:5812-5817]. A glnB-deficient mutant of Synechocystis was shown to grow photoautotrophycally and photoheterotrophically (in the light in the presence of glucose plus DCMU) but not photomixotrophically (in the light in the presence of glucose). In Nostoc punctiforme, on the other hand, Hanson could not obtain a homozygous mutant lacking the wild-type glnB gene and inferred that PII is essential for the growth of this cyanobacterium. These results demonstrate that PII is involved not only in regulation of nitrate assimilation but also in other important cellular processes in cyanobacteria.
The molecular mechanism of regulation of the phosphorylation state of PII was greatly advanced by Karl Forchhammer's group (Universität München) through biochemical studies on PII, PII kinase and PII phosphatase of Synechococcus sp. strain PCC 7942. PII binds ATP and 2-oxoglutarate (2-OG; a-ketoglutarate), and the binding of ATP and 2-OG to PII facilitates binding of the other substrate. Under physiological conditions, PII is supposed to constantly bind ATP. PII kinase phosphorylates PII-ATP-2-OG complex and the phosophorylated form of PII is resistant to the action of PII-P phosphatase as long as ATP and 2-OG is bound. Upon dissociation of 2-OG from the complex, PII-P is rapidly dephosphorylated by PII-P phosphatase. Thus, the PII functions as the sensor of cellular 2-OG level.
According to this scheme, nitrogen status of the cells indirectly regulates PII phosphorylation. When ammonium is depleted or ammonium fixation is inhibited, depletion of glutamine results in accumulation of 2-OG (the substrate of GOGAT), which in turn results in phosphorylation of PII to activate the nitrate assimilation pathway. When ammonium is added to the cells, on the other hand, rapid consumption of 2-OG would results in dephosphorylation of PII to inhibit nitrate assimilation. It is to be determined whether the intracellular level of 2-OG actually changes in response to the changes in the nitrogen status of the cell. The PII phosphatase and the PII kinase were purified separately from Synechococcus cells. Partially purified PII-P phosphatase acts on histone protein, casein, and the PII protein but not on ATP and pNPP. The phosphatase activity requires 20 mM Mg2+ and belongs to a PP2C type phosphatase family.
Another important finding related to PII was the nitrogen- responsive transcriptional regulation of glnB. In Synechococcus sp. strain PCC 7942 (described by Hyun-Mi Lee) and Synechocystis sp. strain PCC 6803 (described by Mario García-Domínguez), glnB was shown to have multiple transcription start sites, transcription from one of which is stimulated by nitrogen depletion in a NtcA-dependent manner. The PII protein level in nitrogen-starved cells of Synechocystis was shown to be three-times that of nitrogen- replete cells. Thus, the capacity of PII-mediated post-translational regulation of nitrate assimilation is increased when nitrate assimilation activity is induced.