Helicobacter pylori is one of the most common human pathogens worldwide, recognized as the causative agent of severe diseases such as peptic ulcer and malignant gastric cancer, ranking this microorganism a class 1 carcinogen. Although H. pylori infection can be treated with antibiotics, the currently available therapies are beginning to lose efficacy because of an increasing rate of antibiotic resistance. H. pylori was included in the 2017 WHO list of 12 pathogens that pose a major threat for humankind, to stimulate the development of novel antimicrobial strategies.
The H. pylori gene hp1043 encodes the essential transcriptional regulator HP1043 for which some structural data are available. However, the impossibility of generating an hp1043-deletion mutant, or to overexpress HP1043 in the cell, has hindered its functional characterization. Using ChIP-seq methodology, we demonstrated that HP1043 binds in vivo the promoter of genes involved in fundamental cellular processes, including replication, transcription, translation, energy production and conversion. In addition, HP1043 targets the promoter of CncR1, a conserved small RNA encoded by the cag pathogenicity island. Although the regulatory function of CncR1 and its target mRNAs are still unknown, it appears that this sRNA plays a pivotal role in the opposite modulation of bacterial motility and adhesion to host cells, a crucial aspect of H. pylori pathogenesis. Besides emerging as a unique example of a single transcriptional regulator that orchestrates all important cellular processes, HP1043 represents an appealing antibacterial drug target.
We aim to tackle the regulatory role of HP1043 and its involvement in the pathogenesis through the control of CncR1. To achieve this goal, an hp1043 conditional mutant will be constructed and whole transcriptome analysis will be performed. We will define the full interactome of CncR1 sRNA through the implementation of a recently developed functional genomics approach called MAPS (MS2 affinity purification coupled with RNA sequencing).
A second line of research aims at the identification and characterization of molecules able to disrupt HP1043 function and block H. pylori infection. To pursue this objective, we have generated and validated a structural model of HP1043 interacting with DNA to carry out virtual screening of libraries of molecules to identify ligands predicted to bind HP1043 dimerization and/or DNA-binding interface, thereby blocking its regulatory activity. Identified molecules will be experimentally validated both in vitro on the purified protein and in vivo by using E. coli reporter strains and on H. pylori cell cultures.
Obtained results will offer an in-depth understanding of HP1043 regulatory mechanism and of its role in pathogenesis, as well as a further step towards the design of novel molecules able to block H. pylori infection.
