## 2 sRNA as Orchestrators of Bacterial Pathogenesis
### Adherence to the Host
#### 2.1.1 PapR  
A large-scale phylogenomic analysis of 907 _E. coli_ isolates, including 722 UPEC strains, identified the _papGII_ locus, encoding P-fimbirae, as a key genetic determinant distinguishing invasive UPEC associated with severe UTI pathogenesis, such as pyclonephritis and urinary-source bacteremia, from non-invasive UPEC strains linked to asymptomatic bacteriuria orystitis [76]. Notably, the number of _pap_-operon copies (_papGEEKJDCHABI_) varies among UPEC strains, with UTI89 harboring a single copy and CFT073 possessing two copies. Beyond their role in adhesion, P-fimbirae contribute to biofilm formation, enhancing bacterial survival by providing protection against host immune responses and antimicrobial agents [77]. The phase variation in P-fimbirae is tightly regulated through an antagonistic interaction between PapI and the small RNA PapR (Figure 1). PapR, a 180-nt long sRNA was 6-fold upregulated in the UPEC strain UTI89 during infection [75]. This small RNA is found in widely studied UPEC strains CFT073 and UTI89 and in some extraintestinal pathogenic _E. coli_ (ExPEC) strains but is missing from _E. coli_ K-12 strains. PapR expression was shown to be regulated by Lrp as deletion of _lrp_ in UTI89 abrogated the expression of PapR [75]. Agglutination assays with yeast and human erythrocytes indicated that _papR_ deletion increased mannose-resistant fimbial expression, such as P-fimbirae, which was restored to wild-type level in the complemented strain [75]. Similarly, adhesion assays using bladder and kidney epithelial cells showed increased mannose-resistant adhesion in UTI89_papR_, reinforcing PapR's role in regulating fimbial expression [75]. Flow cytometry further confirmed that _papR_ deletion doubled the fraction of P-fimbirated cells, while type-1 fimbiae levels remained unchanged [75]. These findings suggest that PapR negatively regulates P-fimbiral expression at both the population and single-cell levels. To achieve this, PapR turns the pap-operon to the OFF state by interacting with PapI mRNA, thereby preventing unnecessary fimbial expression. While PapI promotes the ON state by interacting with the Lrp-DNA complex, PapR inhibits PapI by binding to a specific region (+74 to +96 nucleotides downstream of the PapI translational start site) (Figure 1a). This likely facilitates RNase recruitment or modulates ribosomal loading to repress PapI translation. Consequently, this inhibition disrupts the PapI-mediated translocation of the Lrp-DNA complex, thereby stabilizing the OFF phase [75]. Figure 1: **The role of bacterial sRNAs in virulence. ** (**a**) **The regulation of P-pili expression by PapR:** The expression of P-pili in UPEC is controlled by a phase switch that togles between the OFF and ON states. **Upper left panel:** In the OFF state, the phase switch is regulated by the interaction of Lrp, PapL, PapB, and Dam methylase. The intergenic region between _papB_ and _papI_ contains two sets of methylation sites: proximal and distal. Binding of Lrp to the proximal site prevents Dam methylation, leading to methylation of the distal site and the suppression of _pap_-operon transcription, thereby preventing P-fimbirae expression. **Upper right panel:** In the ON state, Lrp, in association with PapL, binds to the proximal site, facilitating the activation of the phase switch and allowing transcription of the _pap_-operon, leading to P-fimbirae expression. **Lower left panel:** The small RNA PapR posttranscriptionally represses PapL expression by directly interacting with its transcript, preventing ribosome binding. In the absence of PapL, Lrp cannot activate the phase switch, thereby repressing P-fimbirae expression. A few components of this figure were created using Biorender. **(b) The structural components of P-fimbirae:** A schematic representation of the major structural components of P-fimbirae, which facilitate bacterial adherence to host epithelial cells.