Targeting the Disease Response With NlpD and LytM for Effective Nonantibiotic Treatment of Urinary Tract Infections.

Abstract

BACKGROUND: Finding new ways of treating bacterial infections is essential. The NlpD protein, which inhibits RNA polymerase II (Pol II), has shown therapeutic efficacy against urinary tract infection. This study investigated the mechanism of Pol II inhibition and protection by NlpD and its LytM peptide. METHODS: Recombinant NlpD and LytM were screened for interactions with constituents of the Pol II complex, using AlphaFold predictions and protein interaction technology. Treatment effects were quantified in infected tissues and regulated host response pathways identified by genome-wide transcriptomics analysis in models of acute pyelonephritis and acute cystitis in Irf3-/- and Asc-/- mice, respectively. RESULTS: LytM was shown to interact with constituents of the Pol II multiprotein complex, inhibiting the CDK12 kinase from phosphorylating the Pol II subunit RPB1 and disrupting Pol II complex formation by interfering with the interaction between PAF1C and RPB1. The protection by LytM against acute pyelonephritis was accompanied by a reduction in gene expression in infected kidneys from >1900 significantly regulated genes (fold change >6) in the placebo group to about 150 in LytM-treated mice. The inhibition of gene expression in infected kidneys particularly targeted the excessive innate immune response. A similar effect was observed in acute cystitis. Bacterial clearance was accelerated in both model by LytM treatment, with effects against antibiotic-sensitive and resistant Escherichia coli strains. CONCLUSIONS: The results suggest that inhibiting the disease response of the host, using NlpD or LytM, may offer an efficient alternative to antibiotics in these models.