Expanding the Cyanobacterial Nitrogen Regulatory Network: The GntR-Like Regulator PlmA Interacts with the PII-PipX Complex

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Title: Expanding the Cyanobacterial Nitrogen Regulatory Network: The GntR-Like Regulator PlmA Interacts with the PII-PipX Complex
Authors: Labella, Jose I. | Obrebska, Anna | Espinosa Manzano, Javier | Salinas, Paloma | Forcada-Nadal, Alicia | Tremiño, Lorena | Rubio, Vicente | Contreras, Asunción
Research Group/s: Transducción de Señales en Bacterias
Center, Department or Service: Universidad de Alicante. Departamento de Fisiología, Genética y Microbiología
Keywords: PlmA | PII | PipX | Cyanobacteria | Nitrogen regulation | Signaling | GntR | Three hybrid interactions
Knowledge Area: Genética
Issue Date: 28-Oct-2016
Publisher: Frontiers Media
Citation: Frontiers in Microbiology. 2016, 7:1677. doi:10.3389/fmicb.2016.01677
Abstract: Cyanobacteria, phototrophic organisms that perform oxygenic photosynthesis, perceive nitrogen status by sensing 2-oxoglutarate levels. PII, a widespread signaling protein, senses and transduces nitrogen and energy status to target proteins, regulating metabolism and gene expression. In cyanobacteria, under conditions of low 2-oxoglutarate, PII forms complexes with the enzyme N-acetyl glutamate kinase, increasing arginine biosynthesis, and with PII-interacting protein X (PipX), making PipX unavailable for binding and co-activation of the nitrogen regulator NtcA. Both the PII-PipX complex structure and in vivo functional data suggested that this complex, as such, could have regulatory functions in addition to PipX sequestration. To investigate this possibility we performed yeast three-hybrid screening of genomic libraries from Synechococcus elongatus PCC7942, searching for proteins interacting simultaneously with PII and PipX. The only prey clone found in the search expressed PlmA, a member of the GntR family of transcriptional regulators proven here by gel filtration to be homodimeric. Interactions analyses further confirmed the simultaneous requirement of PII and PipX, and showed that the PlmA contacts involve PipX elements exposed in the PII-PipX complex, specifically the C-terminal helices and one residue of the tudor-like body. In contrast, PII appears not to interact directly with PlmA, possibly being needed indirectly, to induce an extended conformation of the C-terminal helices of PipX and for modulating the surface polarity at the PII-PipX boundary, two elements that appear crucial for PlmA binding. Attempts to inactive plmA confirmed that this gene is essential in S. elongatus. Western blot assays revealed that S. elongatus PlmA, irrespective of the nitrogen regime, is a relatively abundant transcriptional regulator, suggesting the existence of a large PlmA regulon. In silico studies showed that PlmA is universally and exclusively found in cyanobacteria. Based on interaction data, on the relative amounts of the proteins involved in PII-PipX-PlmA complexes, determined in western assays, and on the restrictions imposed by the symmetries of trimeric PII and dimeric PlmA molecules, a structural and regulatory model for PlmA function is discussed in the context of the cyanobacterial nitrogen interaction network.
Sponsor: This work was supported by grants BFU2015-66360-P to AC and BFU2014-58229-P to VR from the Spanish Ministry of Economy and Competitivity. AO was the recipient of Grisolia Fellowship from Consellería d'Educació of the Valencian Government and AF-N and LT held FPI fellowships/contracts from Ministry of Economy and Competitivity. JE and VR were supported by grants GV/2014/073 and PrometeoII/2014/029, respectively, from the Consellería d'Educació of the Valencian Government.
URI: http://hdl.handle.net/10045/59648
ISSN: 1664-302X
DOI: 10.3389/fmicb.2016.01677
Language: eng
Type: info:eu-repo/semantics/article
Rights: © 2016 Labella, Obrebska, Espinosa, Salinas, Forcada-Nadal, Tremiño, Rubio and Contreras. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
Peer Review: si
Publisher version: http://dx.doi.org/10.3389/fmicb.2016.01677
Appears in Collections:INV - TSB - Artículos de Revistas

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