# The World of Cyclic Dinucleotides in Bacterial Behavior
## 5 Conclusions  
Recently, our knowledge about cyclic dinucleotide second messengers has been under expansion with the discoveries of new CDNs. At the moment, three different classes of prokaryotic proteins are known to synthesize CDN molecules: (i) proteins containing GGDEF domain that synthesizes mainly c-di-GMP; (ii) CD-NTases enzymes that have the catalytic domain known as SMODS and synthesizes mainly cGAMP; and (iii) DAC proteins that have a catalytic domain called DAC domain (also described as DisA_N domain) that synthesizes mainly c-di-AMP. These CDN synthetases do not share structural similarities, use different residues for substrate binding, and probably possess different catalytic mechanisms suggesting that they probably evolved independently to catalyze similar chemical reactions.  
As evidence of the importance and ubiquity of bacterial CDNs, it is interesting to note that mammalian cells evolved to sense bacteria by detecting these molecules to stimulate the immune system to counterattack infections. Thus, the use of CDNs as adjuvants in vaccines has been considered, as they can be used as stimulators of the innate immune system [63; 195].  
The huge repertoire of CDN receptors and the complexity of CDN signaling networks in bacteria are shown in this review. Surprisingly, different CDNs share conformational similarities even in the pocket of different classes of receptors. CDNs can regulate bacterial behaviors at different speed levels, directly regulating protein function for a faster response or, more slowly, by affecting gene transcription or protein translation. Remarkably, different CDN second messenger systems may coexist in many organisms, which would imply that both the control of their synthesis and turnover and the specificity of their sensors are carefully tuned.  
Therefore, the new discoveries reviewed in this paper open up questions about how bacteria coordinate the three mains bacterial CDNs: are they interconnected to regulate the same bacterial phenotype, or do they act independently? Do bacteria use the three CDNs as second messengers or is one chosen? Are the CDN signaling pathways conserved in different bacteria? Will other CDNs be discovered to also be second messengers?  
The following are available online. Table S1: Distribution of DAC, GGDEF, and SMODS homologs across prokaryotic lineages. This data is the basis for Figure 9.  
**Author Contributions:** [PERSON] and [PERSON] contributed to the analysis of the structures and ligand conformations. [PERSON] contributed to the CDN receptors analysis. [PERSON] performed the bioinformatic analysis and [PERSON] contributed to the ligand graph preparation and analysis. All authors contributed to the writing and to the revising of the text. All authors have read and agreed to the published version of the manuscript.  
This review was supported by grants of FAPESP (project numbers: 2019/00195-2; 2018/21076-9; 2018/08996-1, 2017/10611-8, and 2016/08414-7), CNPq (project number: 403965/2016-0), and CAPES (project number: 23038.015071/2018-12).  
The authors declare that they have no conflict of interest with the contents of this article.