TYPE 4 FILAMENTS

Our primary research objectives are to progress towards a better understanding of the molecular mechanisms of filament assembly and of different T4F-mediated functions. The two distinctive features of our experimental approach are (1) its multi-disciplinarity, allowing us to bridge the scales from bacterial cells/communities down to atomic-resolution structures of T4F components, and (2) the use of several bacterial models, one which is often better suited to answer a particular research question. For the past 20 years – an effort still ongoing – we have been tackling many aspects of T4F biology and have made significant contributions to the field. The following selected examples do not represent an exhaustive list.

T4F assembly

1. We have shown that only a fraction of T4F components required for filament biogenesis in complex systems such as N. meningitidis T4aP – which stands for type 4a pili – are required for filament assembly.
2. We have confirmed this by reconstituting a smaller machinery composed only of those components, which is capable of assembling T4aP in a non-piliated heterologous host.
3. By studying new T4F models – notably the Com pilus widespread in monoderms – we found that only four “core” proteins universally conserved in T4F – several type 4 pilins, a prepilin peptidase, an extension ATPase and a so-called platform protein are sufficient for filament assembly. We have recently been able to produce a model of a complete T4F machinery, which serves as a blueprint for ongoing studies.

T4F-mediated functions

1. We have shed light on the role of T4F in DNA uptake during natural transformation by identifying the first type 4 pilin with intrinsic DNA-binding ability, i.e., the ComP minor pilin in N. meningitidis T4aP. We showed that ComP binds DNA in a novel way, with a specificity for a highly repeated motif in the meningococcal genome. This illuminated the unusual propensity of Neisseria species to preferentially take up their own DNA.
2. We have shown how T4aP mediate adhesion of S. sanguinis to human cells. By determining the structures of all the subunits of this pilus (major and minor) and of the filament itself – a first in the T4F field – we found that S. sanguinis T4aP are capped by a complex of minor pilins mediating adhesion either to host proteins or glycans. Two of these minor pilins have extra functional modules, “grafted” onto pilins and directly binding the above receptors. We named these “modular pilins” and showed that they are widespread in bacteria, with an astonishing variety of architectures.
3. Using M. xanthus as a model, we have shown that twitching motility – a widespread type of motility powered exclusively by T4aP – is modulated by a complex at the tip of the filaments. This complex consists of four minor pilins, capped by the large non-pilin protein PilY1. The presence of three PilY1 paralogs in M. xanthus genome – exhibiting a modular architecture with conserved C-termini and highly variable N-termini – suggests that this species can accessorise the tip of its T4P to allow twitching motility in changing environmental conditions by binding to different substrates.

Many of the above projects are currently being further developed to gain additional insight in T4F biology. Additional projects are also under way to address important unanswered research questions in the field.