A global search for high-susceptibility targets of programmable RNA antibiotics

The growing emergence of multidrug-resistant bacterial pathogens urgently demands the development of alternative antibiotics. Antisense oligomers (ASOs), such as based on peptide nucleic acid (PNA), can exert potent bactericidal effects when designed to sequester the ribosome binding site of an mRNA of an essential gene. Over the years, various essential genes have been investigated for their vulnerability to PNA targeting in enteric bacteria, with the acpP gene encoding acyl carrier protein being considered the most susceptible PNA target. However, it remains unknown what makes for a potent PNA or PNA target. We present a systematic investigation of the efficacy of antisense PNAs for 293 selected essential genes, tested in both E. coli K12 and uropathogenic E. coli (UPEC). Our screen has produced 23 and 55 potent antibacterial PNAs, corresponding to 21 and 47 essential target genes, in K12 and UPEC, respectively. In addition, we discover both strain-independent and strain-specific antibacterial PNA activities. Bioinformatic analysis will enable us to decipher common traits of the identified most efficient PNA sequences and determine core features of target susceptibility. Further, characterization and testing of UPEC-specific PNAs on a broader panel of Enterobacteriaceae may help to generate species-specific antibacterials. Overall, this study gives new insights into sequence- and target gene-dependent differences in PNA susceptibility and provides information on putative universal predictors of PNA efficacy.