Open Access
Regulation of gene expression by non-phosphorylated response regulators
(Institut d'Estudis Catalans, 2021) Gómez Arrebola, Carmen; Solano Goñi, Cristina; Lasa Uzcudun, Íñigo; Ciencias de la Salud; Osasun Zientziak
Two-component systems (TCSs) are a prominent sensory system in bacteria. A prototypical TCS comprises a membrane-bound sensor histidine kinase (HK) responsible for sensing the signal and a cytoplasmic response regulator (RR) that controls target gene expression. Signal binding activates a phosphotransfer cascade from the HK to the RR. As a result, the phosphorylated RR undergoes a conformational change that leads to activation of the response. Growing experimental evidence indicates that unphosphorylated RRs may also have regulatory functions, and thus, the classical view that the RR is only active when it is phosphorylated needs to be revisited. In this review, we highlight the most recent findings showing that RRs in the non-phosphorylated state control critical bacterial processes that range from secretion of factors to the host, antibiotic resistance, iron transport, stress response, and cell-wall metabolism to biofilm development.
Open Access
Systematic reconstruction of the complete two-component sensorial network in staphylococcus aureus
(American Society for Microbiology, 2020) Rapún Araiz, Beatriz; Haag, Andreas F.; Gil Puig, Carmen; Dorado Morales, Pedro; Lasa Uzcudun, Íñigo; Ciencias de la Salud; Osasun Zientziak
In bacteria, adaptation to changes in the environment is mainly controlled through two-component signal transduction systems (TCSs). Most bacteria contain dozens of TCSs, each of them responsible for sensing a different range of signals and controlling the expression of a repertoire of target genes (regulon). Over the years, identification of the regulon controlled by each individual TCS in different bacteria has been a recurrent question. However, limitations associated with the classical approaches used have left our knowledge far from complete. In this report, using a pioneering approach in which a strain devoid of the complete nonessential TCS network was systematically complemented with the constitutively active form of each response regulator, we have reconstituted the regulon of each TCS of S. aureus in the absence of interference between members of the family. Transcriptome sequencing (RNA-Seq) and proteomics allowed us to determine the size, complexity, and insulation of each regulon and to identify the genes regulated exclusively by one or many TCSs. This gain-of-function strategy provides the first description of the complete TCS regulon in a living cell, which we expect will be useful to understand the pathobiology of this important pathogen. IMPORTANCE Bacteria are able to sense environmental conditions and respond accordingly. Their sensorial system relies on pairs of sensory and regulatory proteins, known as two-component systems (TCSs). The majority of bacteria contain dozens of TCSs, each of them responsible for sensing and responding to a different range of signals. Traditionally, the function of each TCS has been determined by analyzing the changes in gene expression caused by the absence of individual TCSs. Here, we used a bacterial strain deprived of the complete TC sensorial system to introduce, one by one, the active form of every TCS. This gain-of-function strategy allowed us to identify the changes in gene expression conferred by each TCS without interference of other members of the family.
Open Access
The role of ArlRS and VraSR in regulating ceftaroline hypersusceptibility in methicillin-resistant Staphylococcus aureus
(MDPI, 2021) Villanueva San Martín, Maite; Roch, Melanie; Lasa Uzcudun, Íñigo; Renzoni, Adriana; Kelley, William L.; Ciencias de la Salud; Osasun Zientziak
Methicillin-resistant Staphylococcus aureus infections are a global health problem. New control strategies, including fifth-generation cephalosporins such as ceftaroline, have been developed, however rare sporadic resistance has been reported. Our study aimed to determine whether disruption of two-component environmental signal systems detectably led to enhanced susceptibility to ceftaroline in S. aureus CA-MRSA strain MW2 at sub-MIC concentrations where cells normally continue to grow. A collection of sequential mutants in all fifteen S. aureus non-essential two-component systems (TCS) was first screened for ceftaroline sub-MIC susceptibility, using the spot population analysis profile method. We discovered a role for both ArlRS and VraSR TCS as determinants responsible for MW2 survival in the presence of sub-MIC ceftaroline. Subsequent analysis showed that dual disruption of both arlRS and vraSR resulted in a very strong ceftaroline hypersensitivity phenotype. Genetic complementation analysis confirmed these results and further revealed that arlRS and vraSR likely regulate some common pathway(s) yet to be determined. Our study shows that S. aureus uses particular TCS environmental sensing systems for this type of defense and illustrates the proof of principle that if these TCS were inhibited, the efficacy of certain antibiotics might be considerably enhanced.

Lasa Uzcudun, Íñigo

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