Biology subjects

Girard, M.

Publications and source records attributed to Girard, M..

28 records · Page 2Linked to original sources

bioRxiv2020

Finite-size transitions in complex membranes

The lipid raft hypothesis postulates that cell membranes possess some degree of lateral organization. The last decade has seen a large amount of experimental evidence for rafts. Yet, the underlying mechanism remains elusive. One hypothesis that supports rafts relies on the membrane to lie near a critical point. While supported by experimental evidence, the role of regulation is unclear. Using both a lattice model and molecular dynamics simulations, we show that lipid regulation of a many-component membrane can lead to critical behavior over a large temperature range. Across this range, the membrane displays a critical composition due to finite-size effects. This mechanism provides a rationale as to how cells tune their composition without the need for specific sensing mechanisms. It is robust and reproduces important experimentally verified biological trends: membrane-demixing temperature closely follows cell growth temperature, and the composition evolves along a critical manifold. The simplicity of the mechanism provides a strong argument in favor of the critical membrane hypothesis. SIGNIFICANCEWe show that biological regulation of a large amount of phospholipids in membranes naturally leads to a critical composition for finite-size systems. This suggests that regulating a system near a critical point is trivial for cells. These effects vanish logarithmically and therefore can be present in micron-sized systems.

biophysics↗

bioRxiv2020

Moleculewise semi-grand canonical ensembles

The plasma membrane is the interface between cells and exterior media. While its existence has been known for a long time, organization of its constituent lipids remain a challenge. Recently, we have proposed that lipid populations may be controlled by chemical potentials of different lipid species, resulting in semi-grand canonical thermodynamic ensembles. However, the currently available molecular dynamics software packages do not allow for molecule-based chemical potentials. Here, we propose a variation on existing algorithms that allow defining chemical potentials for molecules. Additionally, we allow coupling with collective variables and show that it can be used to dynamically create asymmetric membranes. We release an implementation of the algorithm for the HOOMD-Blue molecular dynamics engine. SIGNIFICANCEWe demonstrate an algorithm that allows for simulations of molecules in the semi-grand canonical ensemble. It also allows coupling the chemical potential values to collective variable and create asymmetric membranes.

biophysics↗

bioRxiv2020

An Arf/Rab cascade controls the growth and invasiveness of glioblastoma

Glioblastoma is the most common and deadly malignant brain cancer. We now demonstrate that loss of function of the endosomal GTPase Rab35 in human brain tumor initiating cells (BTICs) increases glioblastoma growth and decreases animal survival following BTIC implantation in mouse brain. Mechanistically, we identify that the GTPase Arf5 interacts with the guanine nucleotide exchange factor (GEF) for Rab35, DENND1/connecdenn and allosterically enhances its GEF activity towards Rab35. Knockdown of either Rab35 or Arf5 increases cell migration, invasiveness and self-renewal in culture and enhances the growth and invasiveness of BTIC-initiated brain tumors in mice. RNAseq of the tumors reveals upregulation of the tumor-promoting transcription factor SPOCD1, and disruption of the Arf5/Rab35 axis in glioblastoma cells leads to strong activation of the epidermal growth factor receptor with resulting enhancement of SPOCD1 levels. These discoveries reveal an unexpected cascade between an Arf and a Rab and indicate a role for the cascade, and thus endosomal trafficking, in brain tumors.

cancer biology↗

bioRxiv2019

Regulating lipid composition rationalizes acyl-tail saturation homeostasis in ectotherms

Cell membranes mainly consist of lipid bilayers with an actively regulated composition. The underlying processes are still poorly understood, in particular how the hundreds of components are controlled. Surprisingly, in recent experiments on ectotherms, the cholesterol fraction, along with un- and mono-saturated acyl tail fractions and demixing temperatures, was shown to increase with body temperature. We establish a model based on chemical reaction networks to study regulation of membranes, resulting in multiple semi-grand canonical ensembles. By running computer simulations, we show that higher cholesterol fractions correlate with lower degrees of unsaturation, ultimately controlling the composition of lipid tails. Cholesterol also dictates membrane viscosity and regulation of the latter implies that cholesterol must increase with temperature. Overall, our model proposes a different picture of lipid regulation, where components can be passively, instead of actively, regulated. SIGNIFICANCEIn this article, we propose a regulation model where only some of the components are actively regulated between membranes, while others are naturally balanced by chemical potentials. This model provides a rationale to recently measured puzzling trends in ectotherms, that is, increased plasma membrane cholesterol fraction with temperature. Here, we show that it is directly correlated with with acyl tail saturation and order parameter correlation length. Furthermore, we highlight the relation between cholesterol and membrane viscosity.

biophysics↗

bioRxiv2019

RpiRc regulates RsbU to modulate eDNA-dependent biofilm formation and in vivo virulence of Staphylococcus aureus in a mouse model of catheter infection

Staphylococcus aureus is a major human pathogen. Despite high incidence and morbidity, molecular mechanisms occurring during infection remain largely unknown. Under defined conditions, biofilm formation contributes to the severity of S. aureus related infections. Extracellular DNA (eDNA), a component of biofilm matrix released from apoptotic bacteria, is involved in biofilm structure and stability. In many bacterial biofilms, eDNA originates from cell lysis although eDNA can also be actively secreted or exported by bacterial membrane vesicles. By screening the Nebraska transposon library, we identified rpiRc as a biofilm regulator involved in eDNA regulation. RpiRc is a transcription factor from the pentose phosphate pathway (PPP) whose product is a polysaccharide intercellular adhesin (PIA) precursor. However, rpiRc mutant strain showed neither susceptibility to DispersinB(R) (a commercially available enzyme disrupting PIA biofilms) nor alteration of ica transcription (the operon regulating PIA production). Decreased biofilm formation was linked to Sln, an extracellular compound degrading eDNA in an autolysis independent pathway. Biofilm susceptibility to antibiotics in wt and mutant strains was tested using a similar protocol as the Calgary biofilm device. Involvement of RpiRc in S. aureus virulence was assessed ex vivo by internalization experiments into HEK293 cells and in vivo in a mouse model of subcutaneous catheter infection. While minimum inhibitory concentrations (MICs) of planktonic cells were not affected in the mutant strain, we observed increased biofilm susceptibility to almost all tested antibiotics, regardless of their mode of action. More importantly, the rpiRc mutant showed reduced virulence in both ex vivo and in vivo experiments related to decreased fnbpA-B transcription and eDNA production. RpiRc is an important regulator involved in eDNA degradation inside the matrix of mature PIA independent biofilms. These results illustrate that RpiRc contributes to increased antibiotic tolerance in mature bacterial biofilm and also to S. aureus cell adhesion and virulence during subcutaneous infection.\n\nAuthor summaryBiofilm formation contributes to the severity of Staphylococcus aureus related infections. Biofilm matrix is mainly composed by polysaccharide intercellular adhesion (PIA), proteins and extracellular DNA (eDNA). By screening a mutant library of S. aureus, RpiRc was identified as a new regulator of eDNA dependent biofilm formation. How RpiRc regulates biofilm and its role in S. aureus virulence was studied in four different S. aureus strains. Deletion of RpiRc resulted in a pronounced decreased eDNA dependent biofilm formation, but not PIA dependent biofilm formation. Decreased biofilm formation was not related to increased autolysis, but was linked to extracellular compounds found in the supernatant of mutant biofilms. Sln was identified as one of this compound. RpiRc deletion also decreased biofilm recalcitrance (resistance) to selected antibiotics. Involvement of RpiRc in S. aureus pathogenesis was investigated ex vivo by internalization into HEK293 cells and in vivo in a mouse model of catheter infection. RpiRc deletion resulted in decreased virulence related to decreased expression of surface proteins like the fibronectin binding proteins A and B (FnbpA-B). These results illustrate that RpiRc contributes to increased antibiotic tolerance in mature bacterial biofilm and also to S. aureus cell adhesion and virulence during subcutaneous infection.

microbiology↗

bioRxiv2019

Immunotherapy prevents long-term disability in relapsing multiple sclerosis over 15 years

ObjectiveWhether immunotherapy improves long-term disability in multiple sclerosis has not been satisfactorily demonstrated. This study examined the effect of immunotherapy on long-term disability outcomes in relapsing-remitting multiple sclerosis.\n\nMethodsWe studied patients from MSBase followed for [≥]1 year, with [≥]3 visits, [≥]1 visit per year and exposed to a multiple sclerosis therapy, and a subset of patients with [≥]15-year follow-up. Marginal structural models were used to compare the hazard of 12-month confirmed increase and decrease in disability, EDSS step 6 and the incidence of relapses between treated and untreated periods. Marginal structural models were continuously re-adjusted for patient age, sex, pregnancy, date, disease course, time from first symptom, prior relapse history, disability and MRI activity.\n\nResults14,717 patients were studied. During the treated periods, patients were less likely to experience relapses (hazard ratio 0.60, 95% confidence interval 0.43-0.82, p=0.0016), worsening of disability (0.56, 0.38-0.82, p=0.0026) and progress to EDSS step 6 (0.33, 0.19-0.59, p=0.00019). Among 1085 patients with [≥]15-year follow-up, the treated patients were less likely to experience relapses (0.59, 0.50-0.70, p=10-9) and worsening of disability (0.81, 0.67-0.99, p=0.043).\n\nConclusionsContinued treatment with multiple sclerosis immunotherapies reduces disability accrual (by 19-44%), the risk of need of a walking aid by 67% and the frequency of relapses (by 40-41%) over 15 years. A proof of long-term effect of immunomodulation on disability outcomes is the key to establishing its disease modifying properties.

neuroscience↗

bioRxiv2019

Chestnut extract but not sodium salicylate decrease the severity of diarrhea and enterotoxigenic Escherichia coli F4 shedding in artificially infected piglets

The development of alternatives to antibiotics is crucial to limiting the incidence of antimicrobial resistance, especially in prophylactic and metaphylactic use to control post-weaning diarrhea (PWD). Feed additives, including bioactive compounds, could be a promising alternative. This study aimed to test two bioactive compounds, sodium salicylate (SA) and a chestnut extract (CE) containing hydrolysable tannins, on the occurrence of PWD. At weaning, 72 piglets were assigned to four treatments that combined two factors: CE supplementation (with 2% of CE (CE+) or without (CE-)) and SA supplementation (with 35 mg/kg BW of SA (SA+) or without (SA-)). Then, 4 days after weaning, all piglets were infected with a suspension at 108 CFU/ml of enterotoxigenic Escherichia coli (ETEC F4ac). Each piglet had free access to an electrolyte solution containing, or not, SA. This SA supplementation was administered for 5 days (i.e., from the day of infection (day 0) to 4 days post-infection (day 4). During the 2 weeks post-infection, supplementation with SA had no effect (P > 0.05) on growth performances nor on fecal scores. A significant SA x time interaction (P < 0.01) for fecal scores and the percentage of diarrhea indicated that piglets with SA did not recover faster and did have a second episode of diarrhea. In contrast to SA treatment, inclusion of CE increased (P < 0.05) growth performances and feed intake. In the first week post-infection, CE decreased (P < 0.001) the overall fecal scores, the percentage of piglets with diarrhea, the days in diarrhea, and ETEC shedding in the feces. There was a SAxCE interaction (P < 0.05) for ETEC shedding, suggesting a negative effect of combining SA with CE. This study highlighted that, in contrast to SA, CE could represent a promising alternative to antibiotics immediately after weaning for improving growth performance and reducing PWD.

physiology↗

bioRxiv2018

From genotype to antibiotic susceptibility phenotype in Enterobacteriaceae: a clinical perspective

Predicting the antibiotic susceptibility phenotype from genomic data is challenging, especially for some specific antibiotics in Enterobacteriaceae. Here we aimed to assess the performance of whole genomic sequencing (WGS) for predicting the antibiotic susceptibility in various Enterobacteriaceae species using the detection of antibiotic resistance genes (ARGs), specific mutations and a knowledge-based decision algorithm. We sequenced (Illumina MiSeq 2x250b) 187 clinical isolates from species possessing (n=98) or not (n=89) an intrinsic AmpC-type cephalosporinase. Antibiotic susceptibility was performed by the disc diffusion method. Reads were assembled by A5-miseq and ARGs were identified from the ResFinder database using Diamond. Mutations on GyrA and ParC topoisomerases were studied. We assessed the prediction rates for amoxicillin, co-amoxiclav, piperacillin, piperacillin-tazobactam, ceftazidime, cefepime, meropenem, amikacin, gentamicin and fluoroquinolones. A total of 1,870 isolate/antibiotic combinations were considered. In 295 cases (15.8%) no attempt of prediction was made. Among the 1,575 attempts, 1,537 (97.6%) were correct (1,011 for predicting susceptibility and 526 for predicting resistance), 15 (0.9%) were major errors (MEs) and 23 (1.5%) were very major errors (VMEs). The concordance rates were similar between non-AmpC and AmpC-producing Enterobacteriaceae (907/935 [97.0%] vs 630/640 [98.4%], Chi2 test p=0.07), but more VMEs were observed in non-AmpC producing strains than in those producing an AmpC (20/935 [2.1%] vs 3/640 [0.5%], Chi2 test p=0.007). The majority of VMEs were putatively due to the overexpression of chromosomal genes. In conclusion, the inference of antibiotic susceptibility from genomic data showed good performances for non-AmpC and AmpC-producing Enterobacteriaceae species.

microbiology↗

bioRxiv2018

The intestinal microbiota predisposes to traveller’s diarrhoea and to the carriage of multidrug-resistant Enterobacteriaceae after travelling to tropical regions

The risk of acquisition of multidrug-resistant Enterobacteriaceae (MRE) and of occurrence of diarrhoea is high when travelling to tropical regions. The relationships between these phenomena and the composition of human gut microbiota have not yet been assessed. Here, we investigated the dynamics of changes of metabolically active microbiota by sequencing total RNA from faecal samples taken before and after travel to tropical regions. We found that the occurrence of diarrhoea during the travel was associated with a higher relative abundance of Prevotella copri before departure and after return. The composition of microbiota, before travel as well as at return, was not correlated with the acquisition of MRE. However, the clearance of MRE one month after return was linked to a specific pattern of bacterial species that was also found before and after return.

microbiology↗

bioRxiv2016

Clinical metagenomics of bone and joint infections: a proof of concept study

BackgroundBone and joint infections (BJI) are severe infections that require a tailored and protracted antibiotic treatment. The diagnostic of BJI relies on the culture of surgical specimens, yet some bacteria would not grow because of extreme oxygen sensitivity or fastidious growth. Hence, metagenomic sequencing could potentially address those limitations. In this study, we assessed the performances of metagenomic sequencing of BJI samples for the identification of pathogens and the prediction of antibiotic susceptibility.\n\nMethodsA total of 179 samples were considered. The DNA was extracted with a kit aiming to decrease the amount of human DNA (Molzym), and sequenced on an Illumina HiSeq2500 in 2x250 paired-end reads. The taxonomy was obtained by MetaPhlAn2, the bacterial reads assembled with MetaSPAdes and the antibiotic resistance determinants (ARDs) identified using a database made of Resfinder+ARDs from functional metagenomic studies.\n\nResultsWe could sequence the DNA from 24 out of 179 samples. For monomicrobial samples (n=8), the presence of the pathogen was confirmed by metagenomics in all cases. For polymicrobial samples (n=16), 32/55 bacteria (58.2%) were found at the species level (41/55 [74.5%] at the genus level). Conversely, a total of 273 bacteria not found in culture were identified, 182 being possible pathogens undetected in culture and 91 contaminants. A correct antibiotic susceptibility could be inferred in 94.1% cases for monomicrobial samples and in 76.5% cases in polymicrobial samples.\n\nConclusionsWhen sufficient amounts of DNA can be extracted from samples, we found that clinical metagenomics is a potential tool to support conventional culture.

microbiology↗