Research - Interdisciplinarity- 2025

Abstract:

STOP1 from the model plant Arabidopsis thaliana is a master gene for tolerance to low-pH and Al. STOP1 (Sensitive To Proton rhizotoxicity1) is a transcription factor (TF) responsible for tolerance not only to low-pH and Al but also to several other abiotic stresses. A. thaliana mutants lacking STOP1 activity are extremely sensitive to ^H+ and ^Al3+, with this being mainly due to a deficit in the STOP1-dependent expression of root efflux transporters of small metal-chelating organic acids. pH and metals post-transcriptionally regulate the abundance of the STOP1 protein. The mechanism by which acidic pH and Al stimulate the STOP1 signaling pathway remain elusive. STOP1 contains three predicted intrinsically disordered regions (IDRs) accounting for 55 % of the STOP1 sequence. The high conformational flexibility and multivalency of IDRs render them highly prone to undergo liquid-liquid phase separation (LLPS). In eukaryotes, including A. thaliana, TFs are enriched in IDRs. Transcriptional regulators are known to sub-compartmentalize via LLPS. Their condensation can either stimulate their activity by concentrating them with components of the transcription machinery or, on the contrary, inactivates them by sequestration. Importantly, phase-separation of certain IDR-containing proteins depends on the cellular pH. In silico analysis of the STOP1 sequence indicates that its three IDRs contain multiple droplet-promoting regions that could drive LLPS. Our working hypothesis is that STOP1 undergoes LLPS in response to a decrease in pH and that the resulting condensates trigger expression of the target gene(s). The aim of this project is to experimentally ascertain the disordered nature of STOP1 IDRs and to assess their ability to undergo pH-mediated LLPS both in vitro and in planta. The three IDRs will be expressed in E. coli and the resulting purified proteins will be characterized in vitro in terms of their conformational properties and phase-separating abilities. The ability of STOP1 to undergo pH- and Al-induced LLPS in planta will be assessed by confocal microscopy using STOP1-GFP fusions. STOP1 mutants will also be designed, transiently expressed in N.benthamiana and then assessed for their ability to undergo LLPS. This project, by shedding light on the potential phase-separating abilities of STOP1, will constitute a breakthrough in our understanding of the molecular mechanisms underlying the activity of this class of plant TFs with the potential to unveil a new regulatory mechanism.

Abstract:

Protein folding is crucial for proper function, and chaperones play a key role in preventing protein aggregation and facilitating folding. Within the bacterial envelope, the periplasmic space is a site of passage and activity for many proteins, whose proper folding depends on a network of chaperones essential for maintaining proteostasis. Unexpectedly, we have discovered that one of the periplasmic chaperones of E. coli, though already well studied, binds to a heme. This observation, based on solid preliminary results, raises new questions about the role of a holo-chaperone form in periplasmic proteostasis. To this end, we have formed a multidisciplinary consortium bringing together microbiologists from the LCB, biochemists from the BIAM, and physico-chemists from the BIP.

Abstract:

Oxygenic photosynthesis was a pivotal event in Earth's history, enabling organisms to convert light into chemical energy, fixing CO2 and producing _O2, which supported the evolution of aerobic life. Cyanobacteria are unique as the only prokaryotes capable of both oxygenic photosynthesis and nitrogen fixation, making them key models for research and biotechnology. Nitrogen fixation, catalyzed by nitrogenase, is highly sensitive to O2, creating challenges for cyanobacteria, which solve this through specialized heterocysts that maintain an anaerobic environment for nitrogen fixation while neighboring vegetative cells handle photosynthesis.

In heterocysts, sugars from vegetative cells are broken down via the oxidative pentose phosphate pathway, generating NADPH and ATP for nitrogen fixation. This balance is essential for survival, but the transfer of carbon and reductants can limit efficiency. A promising bioengineering approach involves heterocysts producing hydrogen by introducing hydrogenase enzymes. Previous experiments raised bioenergetic questions about how reductants are supplied to heterocysts during hydrogen production, highlighting the need to understand carbon and electron transport in engineered strains.

The REBONC project aims to deepen understanding of bioenergetic trade-offs in cyanobacteria by studying photosynthesis and nitrogen fixation in engineered strains, with the goal of enhancing productivity for biofertilizer and bioenergy applications. The project aligns with Horizon Europe's goals of renewable nitrogen use and advancing agricultural resilience