Unravelling the molecular mechanisms of action of PilC/PilY1, a key player in type 4 pilus biology found at the pilus tip in hundreds of bacterial species – T4Ptip

Type 4 pili (T4P) are unique pili because they are (1) the only known pilus type universally present in bacteria and archaea, and (2) exceptionally versatile, being involved in adhesion, motility, DNA uptake, aggregation, and mechanosensing. T4aP – the most widespread subtype of T4P, found in hundreds of bacterial species – have been studied for decades because they are critical virulence factors in many human pathogens, which makes them primary targets for the design of novel anti-virulence therapies.

T4aP are helical polymers of one major pilin, capped by a complex of four minor pilins PilHIJK, which in most species interacts with PilC/PilY1 that is presented at the pilus tip. PilC/PilY1 – a large non-pilin protein discovered more than 30 years ago, which remains poorly understood – is a key player in T4aP biology essential for piliation, adhesion and mechanosensing.

The aim of T4Ptip is to address three important questions about PilC/PilY1 – using N. meningitidis an important human pathogen and one of the most extensively used T4aP models – how this protein (1) is presented at the pilus tip and why it is required for piliation, (2) mediates adhesion to human cells, and (3) allows bacteria to sense and respond to mechanical cues.

We generated a structural model of the N. meningitidis PilC1/PilHIJK complex at pseudo-atomic resolution, which shed light on all the above questions, suggesting a series of original and unexpected molecular mechanisms. Importantly, we have produced robust preliminary data in support of some of the most critical aspects of this model, strengthening the feasibility of our T4PTip proposal. We will perform an integrative structure/function analysis of PilC/PilY1 using an approach combining structural biology, modelling, biophysics, and molecular genetics.

Our results will have a broad impact because of (1) their originality, (2) the fact that PilC/PilY1 is an extremely widespread protein, and (3) possible practical implications for human health in the longer term.