Transcription Activation by NtcA and 2-Oxoglutarate of Three Genes Involved in Heterocyst Differentiation in the Cyanobacterium Anabaena sp. Strain PCC 7120

Abstract

In Anabaena sp. strain PCC 7120, differentiation of heterocysts takes place in response to the external cue of combined nitrogen deprivation, allowing the organism to fix atmospheric nitrogen in oxic environments. NtcA, a global transcriptional regulator of cyanobacteria, is required for activation of the expression of multiple genes involved in heterocyst differentiation, including key regulators that are specific to the process. We have set up a fully defined in vitro system, which includes the purified Anabaena RNA polymerase, and have studied the effects of NtcA and its signaling effector 2-oxoglutarate on RNA polymerase binding, open complex formation, and transcript production from promoters of the hetC, nrrA, and devB genes that are activated by NtcA at different stages of heterocyst differentiation. Both RNA polymerase and NtcA could specifically bind to the target DNA in the absence of any effector. 2-Oxoglutarate had a moderate positive effect on NtcA binding, and NtcA had a limited positive effect on RNA polymerase recruitment at the promoters. However, a stringent requirement of both NtcA and 2-oxoglutarate was observed for the detection of open complexes and transcript production at the three investigated promoters. These results support a key role for 2-oxoglutarate in transcription activation in the developing heterocyst.

SPR analysis of promoter binding of Synechocystis PCC6803 transcription factors NtcA and CRP suggests cross‐talk and sheds light on regulation by effector molecules

“…Although this overall view appears clear, the question of what makes some genes of the NtcA regulon more or less 2OG- [18] or PipX-sensitive [14] remains to be clarified. Furthermore, the possibility that there could be cross-activation of NtcA-or CRP-dependent promoters by the non-cognate factor must be clarified (Fig.…”

The Emergence of 2-Oxoglutarate as a Master Regulator Metabolite

“…As noted above, in cyanobacteria, high 2-OG levels are indicative of nitrogen deficiency (26). 2-OG binds to NtcA and acts as an allosteric effector by increasing the affinity of NtcA for specific DNA binding targets, resulting in transcriptional activation in vitro (79)(80)(81). Canonical NtcA binding sites are palindromic and, in promoters subject to activation, are frequently centered at position ϳϪ41.5 with respect to the transcriptional start site (77,82), similar to class II promoters activated by Crp and Fnr (83).…”

Nitrogen Fixation and Hydrogen Metabolism in Cyanobacteria

“…Expression of hetR is upregulated by nitrogen deprivation, and this upregulation depends on NtcA (62). Heterocyst formation is also controlled by the availability of 2-oxoglutarate, which provides the carbon skeleton for the incorporation of inorganic nitrogen and which also serves as a signal molecule of the organic carbon content in the developing heterocysts (122,161,223). NtcA is the main 2-oxoglutarate sensor for the initiation of heterocyst differentiation (239).…”