Glutamine synthetase mRNA releases sRNA from its 3′UTR to regulate carbon/nitrogen metabolic balance in Enterobacteriaceae

Abstract

Glutamine synthetase (GS) is the key enzyme of nitrogen assimilation induced under nitrogen limiting conditions. The carbon skeleton of glutamate and glutamine, 2-oxoglutarate, is supplied from the TCA cycle, but how this metabolic flow is controlled in response to nitrogen availability remains unknown. We show that the expression of the E1o component of 2-oxoglutarate dehydrogenase, SucA, is repressed under nitrogen limitation in Salmonella enterica and E coli. The repression is exerted at the post-transcriptional level by an Hfq-dependent sRNA GlnZ generated from the 3´UTR of the GS-encoding glnA mRNA. Enterobacterial GlnZ variants contain a conserved seed sequence and primarily regulate sucA through base-pairing far upstream of the translation initiation region. During growth on glutamine as the nitrogen source, the glnA 3´UTR deletion mutants expressed SucA at higher levels than the S. enterica and E. coli wild-type strains, respectively. In E. coli, the transcriptional regulator Nac also participates in the repression of sucA. Lastly, this study clarifies that the release of GlnZ from the glnA mRNA by RNase E is essential for the post-transcriptional regulation of sucA. Thus the mRNA coordinates the two independent functions to balance the supply and demand of the fundamental metabolites.

Global RNA interactome of nitrogen starvedEscherichia coliuncovers a conserved post-transcriptional regulatory axis required for optimal growth recovery

“…This is exemplified by GlnZ whose expression is activated by NtrC and is thus a member of the NtrC regulon. The regulatory targets of GlnZ have been explored in a number of previous studies (50,51), and many of those identified previously have also been identified in the currently study (Figure S3A). However, our data additionally uncovered that GlnZ interacts with the mRNA of nac in the N-growth state, which encodes a global transcription factor and is a member of the NtrC regulon.…”

“…For example, GlnZ could have a positive regulatory effect on nac mRNA and thereby increase Nac proteins levels in N-bacteria to repress the transcription of sucA . As sucA encodes a subunit of 2-oxoglutarate dehydrogenase enzyme, the cooperative action of GlnZ and Nac could allow efficient balancing of carbon and nitrogen metabolism at the onset of N starvation (50,51). Further, our analysis of RNA-RNA chimeras involving NtrC activated genes from different stages of N starvation in E. coli underscores that post-transcriptional regulation is clearly an important, yet relatively underexplored, facet of the adaptive response to N starvation in E. coli (Figure S3B).…”

“…In contrast, the extent to which post-transcriptional regulation contributes to the adaptive response to N starvation is not fully understood. Recently, the sRNA GlnZ was shown to promote cell survival by regulating genes linked to N and carbon flux in N starved E. coli in an Hfq-dependent manner (50,51). In MS2-affinity purification and RNA sequencing (MAPS) experiments using GlnZ as bait, conducted in short-term N-starved E. coli (i.e., a condition comparable to N-), GlnZ interacted with numerous mRNA targets (51).…”

Global Hfq-mediated RNA interactome of nitrogen starved Escherichia coli uncovers a conserved post-transcriptional regulatory axis required for optimal growth recovery

“…This is exemplified by GlnZ whose expression is activated by NtrC and is thus a member of the NtrC regulon. The regulatory targets of GlnZ have been explored in a number of previous studies ( 53 , 54 ), and many of those identified previously have also been identified in the currently study ( Supplementary Figure S3A ). However, our data additionally uncovered that GlnZ interacts with the mRNA of nac in the N- growth state, which encodes a global transcription factor and is a member of the NtrC regulon.…”

“…In contrast, the extent to which post-transcriptional regulation contributes to the adaptive response to N starvation is not fully understood. Recently, the sRNA GlnZ was shown to promote cell survival by regulating genes linked to N and carbon flux in N starved E. coli in an Hfq-dependent manner ( 53 , 54 ). In MS2-affinity purification and RNA sequencing (MAPS) experiments using GlnZ as bait, conducted in short-term N-starved E. coli (i.e.…”