The NADP+-isocitrate dehydrogenase gene (icd) is nitrogen regulated in cyanobacteria.
NADP+-isocitrate dehydrogenase (NADP+-IDH) activity and protein levels in crude extracts from the unicellular cyanobacterium Synechocystis sp. strain PCC 6803 and the filamentous, dinitrogen-fixing Anabaena sp. strain PCC 7120 were determined under different nitrogen conditions. The highest NADP+-IDH activity and protein accumulation were found under dinitrogen-fixing conditions for the Anabaena strain and under nitrogen starvation for Synechocystis sp. PCC 6803. The icd gene that encodes the NADP+-IDH from Synechocystis sp. strain PCC 6803 was cloned by heterologous hybridization with the previously isolated icd gene from Anabaena sp. strain PCC 7120. The two cyanobacterial icd genes show 81% sequence identity and share a typical 44-amino-acid region different from all the other icd genes sequenced so far. The icd gene seems to be essential for Synechocystis growth since attempts to generate a completely segregated icd mutant were unsuccessful. Transcripts of 2.0 and 1.6 kb were detected by Northern (RNA) blot analysis, for the Anabaena and Synecho-cystis icd genes, respectively. Maximal icd mRNA accumulation was reached after 5 It of nitrogen starvation in Synechocystis cells and under dinitrogen-fixing conditions in Anabaena cells. Primer extension analysis showed that the structure of the Synechocystis icd gene promoter resembles those of the NtcA-regulated promoters. In addition, mobility shift assays demonstrated that purified Synechocystis NtcA protein binds to the promoter of the icd gene. All these data suggest that the expression of the icd gene from Synechocystis sp. strain PCC 6803 may be subjected to nitrogen control mediated by the positively acting regulatory protein NtcA.
Muro-Pastor
M I
MI
Instituto de Bioquímica Vegetal y Fotosíntesis, Universidad de Sevilla-Consejo Superior de Investigaciones Científicas, Seville, Spain.
Reyes
J C
JC
Florencio
F J
FJ
D001426:Bacterial Proteins; D001483:Base Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D000458:Cyanobacteria / Q000201:enzymology / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D007521:Isocitrate Dehydrogenase / Q000235:genetics*; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D009584:Nitrogen / Q000172:deficiency*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D012329:RNA, Bacterial / Q000032:analysis; D012333:RNA, Messenger / Q000032:analysis; D017422:Sequence Analysis, DNA; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D012329:RNA, Bacterial; D012333:RNA, Messenger; D014157:Transcription Factors; C075617:ntcA protein, Synechococcus; D007521:Isocitrate Dehydrogenase; C039355:isocitrate dehydrogenase (NADP+); D009584:Nitrogen