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Publication Abstract from PubMed

The CRP-FNR (cAMP receptor protein-fumarate/nitrate reductase regulator) superfamily of transcriptional regulators includes the cyanobacterial master regulator NtcA, which orchestrates large responses of cyanobacteria to nitrogen scarcity. NtcA uses as allosteric activator 2-oxoglutarate (2OG), a signal of nitrogen poorness and carbon richness, and binds a co-activating protein (PipX) that shuttles between the signaling protein PII and NtcA depending on nitrogen richness, thus connecting PII signaling and gene expression regulation. Here, combining structural (X-ray crystallography of six types of crystals including NtcA complexes with DNA, 2OG, and PipX), modeling, and functional [electrophoretic mobility shift assays and bacterial two-hybrid (BACTH)] studies, we clarify the reasons for the exquisite specificity for the binding of NtcA to its target DNA, its mechanisms of activation by 2OG, and its co-activation by PipX. Our crystal structures of PipX-NtcA-DNA complexes prove that PipX does not interact with DNA, although it increases NtcA-DNA contacts, and that it stabilizes the active, DNA-binding-competent conformation of NtcA. Superimposition of this complex on a very recently reported cryo-electron microscopy structure of NtcA in a transcription activity complex with RNA polymerase (RNAP), shows that PipX binding helps recruit RNAP by PipX interaction with RNAP, particularly with its gamma and sigma (region 4) subunits, a structural prediction supported here by BACTH experiments.

Structures of the cyanobacterial nitrogen regulators NtcA and PipX complexed to DNA shed light on DNA binding by NtcA and implicate PipX in the recruitment of RNA polymerase.,Forcada-Nadal A, Bibak S, Salinas P, Contreras A, Rubio V, Llacer JL Nucleic Acids Res. 2025 Feb 8;53(4):gkaf096. doi: 10.1093/nar/gkaf096. PMID:39995039^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.