In the cyanobacterium Synechococcus sp. strain PCC 7942, ammonium exerts a rapid and reversible inhibition of the nitrate and nitrite uptake, and the P ss protein (GlnB) is differentially phosphorylated depending on the intracellular N/C balance. RNA/DNA hybridizations, as well as nitrate and nitrite uptake experiments, were carried out with the wild-type strain and a P ss -null mutant. The transcriptional control by ammonium of the expression of the nir-nrtABCD-narB operon remained operative in the mutant but, in contrast to the wild-type strain, the mutant took up nitrate and nitrite even in the presence of ammonium. Moreover, the wild-type phenotype was restored by insertion of a copy of the wild-type glnB gene in the genome of the P ss -null mutant. These results indicate that the unphosphorylated form of P ss is involved in the short-term inhibition by ammonium of the nitrate and nitrite uptake in Synechococcus sp. strain PCC 7942.z 1998 Federation of European Biochemical Societies.

In the cyanobacterium Synechococcus sp. strain PCC 7942, the phosphorylation states of the signal transducer P II protein (GlnB) can change rapidly depending on the nitrogen and carbon supply. A P II -null mutant (MP2) shows no ammonium-dependent inhibition of the nitrate and nitrite uptake, in contrast to the wild-type. New mutants with different types of P II , which may mimic either the phosphorylated (GlnB S49E or GlnB S49D ) or unphosphorylated (GlnB S49A ) form of the protein, were constructed using site-directed in vitro mutagenesis. Mutant MP2-A (GlnB S49A ) grew poorly using nitrate as a nitrogen source and was unable to take up nitrate supplied at 100 mm, even in the absence of externally added ammonium. Mutants MP2-D and MP2-E (GlnB S49D and GlnB S49E , respectively), however, showed nitrate-dependent growth and regulation of nitrate uptake by ammonium, as in the wild-type. Characterization of the mutants also included an analysis of nitrite uptake and of the levels of the nir (nitrate/nitrite assimilation) operon transcripts, the presence of NrtA (nitrate/nitrite transport binding protein), and nitrate and nitrite reductase activities. In vitro, no significant difference was observed in the cooperative binding of ATP and 2-oxoglutarate between the wild-type and the unphosphorylated or phosphorylated-like forms of the mutant P II proteins. The results obtained indicate that both unphosphorylated and phosphorylated-like forms of P II are able to inhibit nitrate uptake in the presence of ammonium, but the unphosphorylated form also has a negative effect in the absence of this nitrogen source. Therefore, an additional effector, possibly 2-oxoglutarate, is required for the P II protein to relieve inhibition of nitrate uptake in the absence of ammonium.Keywords: ammonium inhibition; glnB gene product; nitrate/ nitrite uptake; regulation; Synechococcus sp. strain PCC 7942.Cyanobacteria are phototrophic bacteria that carry out oxygenevolving photosynthesis and mainly assimilate nitrate, nitrite and ammonium, and, in some species, also reduce molecular dinitrogen to fulfill their cellular nitrogen requirements. The assimilation of nitrate comprises several consecutive steps: nitrate enters the cells via a high-affinity transport system involving the NrtABCD permease (an ABC-type transporter) followed by its reduction to nitrite and then ammonium by nitrate reductase (encoded by narB) and nitrite reductase (encoded by nirA), respectively [1]. In the unicellular cyanobacterium Synechococcus sp. PCC 7942, nitrite is taken up by the same NrtABCD permease as nitrate [2±4] and, in addition, there exists a less well characterized second transport system specific for nitrite [4±6]. Ammonium, either taken up by an active transport system that depends on the membrane potential or by the diffusion of unprotonated molecules, or produced internally, is ultimately incorporated into carbon skeletons by the glutamine synthetase/glutamate synthase (GS/GOGAT) cycle to generate all the N-containing metabolites of the cell [1]....

The PII protein is encoded by a unique glnBgene in Synechococcus sp. strain PCC 7942. Its expression has been analyzed in the wild type and in NtcA-null mutant cells grown under different conditions of nitrogen and carbon supply. RNA-DNA hybridization experiments revealed the presence of one transcript species 680 nucleotides long, whatever the nutrient conditions tested. A second transcript species, 620 nucleotides long, absent in the NtcA null mutant, was observed in wild-type cells that were nitrogen starved for 2 h under both high and low CO2and in the presence of nitrate under a high CO2concentration. Primer extension analysis indicated that the two transcript species are generated from two tandem promoters, a ς70 Escherichia coli-type promoter and an NtcA-dependent promoter, located 120 and 53 nucleotides, respectively, from the glnB initiation codon. The NtcA-dependent promoter is up-regulated under the conditions mentioned above, while the ς70 E. coli-type promoter displays constitutive levels of transcripts in the NtcA null mutant and slightly different levels in the wild-type cells, depending on the nitrogen and carbon supplies. In general, a good correlation between the amounts of the two transcript species and that of the PII protein was observed, as revealed by immunodetection with specific antibodies. The phosphorylation level of PII in the wild type is inversely correlated with nitrogen availability and directly correlated with higher CO2 concentration. This regulation is correspondingly less stringent in the NtcA null mutant cells. In contrast, the dephosphorylation of PII is NtcA independent.

Background: The purpose of this study was to evaluate the prevalence of velamentous cord insertion (VCI) and the actual association between pathologically confirmed VCI and perinatal outcomes in twins based on the chorionicity. Methods: All twin pregnancies that received prenatal care at a specialty clinic for multiple pregnancies, from less than 12 weeks of gestation until delivery in a single institution between 2015 and 2018 were included in this retrospective cohort study. Results: A total of 941 twins were included in the study. The prevalence of VCI in dichorionic (DC) twins and monochorionic diamniotic (MCDA) twins was 5.8% and 7.8%, respectively (p = 0.251). In all study population, the prevalence of vasa previa and placenta accreta spectrum was higher in VCI group than that of non-VCI group (p = 0.008 and 0.022). In MCDA twins with VCI, birth weight, 1 and 5-min Apgar score were lower than DC twins with VCI (p = 0.010, 0.002 and 0.000). There was no significant association between VCI and selective fetal growth restriction (p = 0.486), twin-to-twin transfusion syndrome (p = 0.400), and birth-weight discordance (>20% and >25%) (p = 0.378 and 0.161) in MCDA twins. Conclusion: There was no difference in the incidence of VCI in twins based on the chorionicity. Moreover, VCI was not a risk factor for adverse perinatal outcomes excepting vasa previa and placenta accreta spectrum, which had a high incidence in twins with VCI.

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