ruffle fungi differentiate into subterranean fruiting bodies bearing spores sequestered in an inconspicuous globouslike mass of hyphae. Truffle-forming species have evolved in nearly every major group of fleshy fungi over 100 times independently within the Ascomycota and Basidiomycota and the majority of evolutionary transitions to a truffle morphology have occurred in lineages establishing ectomycorrhizal mutualistic symbiosis with plants 1. This pattern suggests that symbiosis has been a major driver in the evolution of truffle diversity. The evolution of the hypogeous lifestyle across a diversity of truffle lineages also suggests that the transition from epigeous to hypogeous fruiting is driven by strong selection for traits, for example pungent odours, that promote

Summary Developmental transitions associated with the life cycle of plant‐symbiotic fungi, such as the ascomycete Tuber melanosporum, are likely to require an extensive reprogramming of gene expression brought about by transcription factors (TFs). To date, little is known about the transcriptome alterations that accompany developmental shifts associated with symbiosis or fruiting body formation. Taking advantage of the black truffle genome sequence, we used a bioinformatic approach, coupled with functional analysis in yeast and transcriptome profiling, to identify and catalogue T. melanosporum TFs, the so‐called ‘regulome’. The T. melanosporum regulome contains 102 homologs of previously characterized TFs, 57 homologs of hypothetical TFs, and 42 putative TFs apparently unique to Tuber. The yeast screen allowed the functional discovery of four TFs and the validation of about one‐fifth of the in silico predicted TFs. Truffle proteins apparently unrelated to transcription were also identified as potential transcriptional regulators, together with a number of plant TFs. Twenty‐nine TFs, some of which associated with particular developmental stages, were found to be up‐regulated in ECMs or fruiting bodies. About one‐quarter of these up‐regulated TFs are expressed at surprisingly high levels, thus pointing to a striking functional specialization of the different stages of the Tuber life cycle.

Ribosome biogenesis in Saccharomyces cerevisiae involves a regulon of >200 genes (Ribi genes) coordinately regulated in response to nutrient availability and cellular growth rate. Two cis-acting elements called PAC and RRPE are known to mediate Ribi gene repression in response to nutritional downshift. Here, we show that most Ribi gene promoters also contain binding sites for one or more General Regulatory Factors (GRFs), most frequently Abf1 and Reb1, and that these factors are enriched in vivo at Ribi promoters. Abf1/Reb1/Tbf1 promoter association was required for full Ribi gene expression in rich medium and for its modulation in response to glucose starvation, characterized by a rapid drop followed by slow recovery. Such a response did not entail changes in Abf1 occupancy, but it was paralleled by a quick increase, followed by slow decrease, in Rpd3L histone deacetylase occupancy. Remarkably, Abf1 site disruption also abolished Rpd3L complex recruitment in response to starvation. Extensive mutational analysis of the DBP7 promoter revealed a complex interplay of Tbf1 sites, PAC and RRPE in the transcriptional regulation of this Ribi gene. Our observations point to GRFs as new multifaceted players in Ribi gene regulation both during exponential growth and under repressive conditions.

Bacterial resistance represents a major health threat worldwide, and the development of new therapeutics, including innovative antibiotics, is urgently needed. We describe a discovery platform, centered on in silico screening and in vivo bioluminescence resonance energy transfer in yeast cells, for the identification of new antimicrobials that, by targeting the protein–protein interaction between the β′-subunit and the initiation factor σ70 of bacterial RNA polymerase, inhibit holoenzyme assembly and promoter-specific transcription. Out of 34 000 candidate compounds, we identified seven hits capable of interfering with this interaction. Two derivatives of one of these hits proved to be effective in inhibiting transcription in vitro and growth of the Gram-positive pathogens Staphylococcus aureus and Listeria monocytogenes. Upon supplementation of a permeability adjuvant, one derivative also effectively inhibited Escherichia coli growth. On the basis of the chemical structures of these inhibitors, we generated a ligand-based pharmacophore model that will guide the rational discovery of increasingly effective antibacterial agents.

The bioluminescence resonance energy transfer (BRET) technology is a widely used live cell-based method for monitoring protein-protein interactions as well as conformational changes within proteins or molecular complexes. Considering the emergence of protein-protein interactions as a new promising class of therapeutic targets, we have adapted the BRET method in budding yeast. In this technical note, we describe the advantages of using this simple eukaryotic model rather than mammalian cells to perform high-throughput screening of chemical compound collections: genetic tractability, tolerance to solvent, rapidity, and no need of expensive robotic systems. Here, the HDM2/p53 interaction, related to cancer, is used to highlight the interest of this technology in yeast. Sharing the protocol of this BRET-based assay with the scientific community will extend its application to other protein-protein interactions, even though it is toxic for mammalian cells, in order to discover promising therapeutic candidates.