Abstract
The sRNA Yfr1 and members of the Yfr2 sRNA family are almost universally present within cyanobacteria. The conserved motifs of these sRNAs are nearly complementary to each other, suggesting their ability to participate in crosstalk. The conserved motif of Yfr1 is shared by members of the Yfr10 sRNA family, members of which are otherwise less conserved in sequence, structure, and synteny compared to Yfr1. The different structural properties enable the discrimination of unique targets of Yfr1 and Yfr10. Unlike most studied regulatory sRNAs, Yfr1 gene expression only slightly changes under the tested stress conditions and is present at high levels at all times. In contrast, cellular levels of Yfr10 increase during the course of acclimation to darkness, and levels of Yfr2 increase when cells are shifted to high light or nitrogen limitation conditions. In this study, we investigated the targetomes of Yfr2, Yfr1, and Yfr10 in Prochlorococcus MED4, establishing CRAFD-Seq as a new method for identifying direct targets of these sRNAs that is applicable to all bacteria, including those that are not amenable to genetic modification. The results suggest that these sRNAs are integrated within a regulatory network of unprecedented complexity in the adjustment of carbon and nitrogen-related primary metabolism.
Highlights
Www.nature.com/scientificreports targets of Yfr[1] in Prochlorococcus MED411
With the exception of tmRNA, the 6 S and 4.5 S sRNAs and RNase P RNA, which are universally conserved throughout all bacterial phyla, all other sRNAs identified in Prochlorococcus are restricted to the marine picocyanobacteria (Prochlorococcus and marine Synechococcus) and do not occur outside this group[7,8]
We recently showed that the GntR family transcriptional regulator PMM1637, which is common to all cyanobacteria, binds to promotor regions of yfr[2] homologs that contain the marine picocyanobacterial CGRE1 motif[17]
Summary
Www.nature.com/scientificreports targets of Yfr[1] in Prochlorococcus MED411. In addition to yfr[1], the yfr[2] sRNA family is conserved and is present in all analysed cyanobacteria[12]. With the exception of tmRNA, the 6 S and 4.5 S sRNAs and RNase P RNA, which are universally conserved throughout all bacterial phyla, all other sRNAs identified in Prochlorococcus are restricted to the marine picocyanobacteria (Prochlorococcus and marine Synechococcus) and do not occur outside this group[7,8] While it appears that Prochlorococcus relies heavily on sRNAs to modulate gene expression, the transcription of sRNAs is coordinated by sigma factors and transcription factors. MAPS technology is based on the affinity purification of MS2-tagged sRNA and its bound targets followed by RNA-Seq, and this method can be used to study sRNAs that function without an RNA chaperone[14] Another approach, called GRIL-Seq, is based on the recombinant expression of T4 ligase and the sRNA of interest, taking advantage of proximity ligation and the subsequent enrichment of hybrid sRNA-target fragments followed by sequencing[15]. The distinct targetomes of Yfr[2], Yfr[1] and Yfr[10] coupled with their interdependency constitute a complex regulatory network comprising hundreds of genes in the Prochlorococcus MED4 genome
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