PipY, a Member of the Conserved COG0325 Family of PLP-Binding Proteins, Expands the Cyanobacterial Nitrogen Regulatory Network

Abstract

Synechococcus elongatus PCC 7942 is a paradigmatic model organism for nitrogen regulation in cyanobacteria. Expression of genes involved in nitrogen assimilation is positively regulated by the 2-oxoglutarate receptor and global transcriptional regulator NtcA. Maximal activation requires the subsequent binding of the co-activator PipX. PII, a protein found in all three domains of life as an integrator of signals of the nitrogen and carbon balance, binds to PipX to counteract NtcA activity at low 2-oxoglutarate levels. PII-PipX complexes can also bind to the transcriptional regulator PlmA, whose regulon remains unknown. Here we expand the nitrogen regulatory network to PipY, encoded by the bicistronic operon pipXY in S. elongatus. Work with PipY, the cyanobacterial member of the widespread family of COG0325 proteins, confirms the conserved roles in vitamin B6 and amino/keto acid homeostasis and reveals new PLP-related phenotypes, including sensitivity to antibiotics targeting essential PLP-holoenzymes or synthetic lethality with cysK. In addition, the related phenotypes of pipY and pipX mutants are consistent with genetic interactions in the contexts of survival to PLP-targeting antibiotics and transcriptional regulation. We also showed that PipY overexpression increased the length of S. elongatus cells. Taken together, our results support a universal regulatory role for COG0325 proteins, paving the way to a better understanding of these proteins and of their connections with other biological processes.

Studies on cyanobacterial protein PipY shed light on structure, potential functions, and vitamin B6‐dependent epilepsy

“…In addition, we have characterized some PipY traits in solution, including its mass, its spectroscopic properties, and both its ability to bind PLP and its sensitivity to the PLP‐targeting antibiotic DCS. The toxicity of this antibiotic for S. elongatus was previously shown to be increased by pipY inactivation .…”

“…Very recently, the synteny in cyanobacteria of the mutually adjacent pipX and pipY genes led to the identification in S. elongatus of functional interactions between these two genes . pipY is a member of the highly widespread and intriguing COG0325 gene family that includes Escherichia coli yggS , a gene that when inactivated causes a phenotype of pyridoxine toxicity and pleiotropic changes related to amino acid–related metabolism .…”

“…Very recently, the synteny in cyanobacteria of the mutually adjacent pipX and pipY genes led to the identification in S. elongatus of functional interactions between these two genes . pipY is a member of the highly widespread and intriguing COG0325 gene family that includes Escherichia coli yggS , a gene that when inactivated causes a phenotype of pyridoxine toxicity and pleiotropic changes related to amino acid–related metabolism . Inactivation of pipY in S. elongatus also enhanced pyridoxine toxicity and led to increased sensitivity to antibiotics that target pyridoxal phosphate (PLP) such as D‐cycloserine (DCS) ; these two traits were influenced by various amino acids .…”

“…Striving toward delineation of the roles of PipY, we decided to characterize physically and structurally PipY from S. elongatus . Such characterization appeared pertinent, given the extensive in vivo functional data collected for its corresponding gene . Furthermore, there was only one published analysis of the structure of another COG0325 protein, YBL036C from yeast , an organism that is phylogenetically distant from cyanobacteria.…”

“…To try to explain these puzzling observations, we have introduced into PipY the mutations corresponding to Pro87Leu and Arg241Gln of PROSC, studying relevant characteristics of the mutant PipY forms. This approach has exploited the apparent universality of COG0325 functions (for which the recent studies on S. elongatus provided strong confirmation ) and the sequence conservation between S. elongatus PipY and human PROSC (Fig. S1).…”