Biology subjects

Safi, H.

Publications and source records attributed to Safi, H..

5 recordsLinked to original sources

bioRxiv2025

Elucidating the interaction network of one of the largest icosahedral capsids in the virosphere

Giant viruses challenge traditional boundaries of virology with their large particle sizes, complex genomes, and unique replication strategies. Yet, despite its 750 nm diameter and incorporation of dozens of proteins, mimivirus virion retains an icosahedral symmetry, a trait often associated with smaller viruses. The functional roles and interactions of most proteins composing such complex icosahedral particles remain elusive. Here, we dissect the spatial and functional organization of mimivirus morphogenesis by integrating bioinformatics, genetics and interactomics. We performed protein clustering using a structure-informed approach integrating AlphaFold models with sequence information to classify and functionally annotate the ORFan-rich mimivirus proteome. To map the protein-protein interaction network during morphogenesis, we employed endogenous tagging and co-immunoprecipitation coupled to mass spectrometry. This strategy revealed distinct interaction modules associated with the virion membrane, nucleoid and viral factory compartments. Comparative analyses with other icosahedral and non-icosahedral giant viruses uncovered conserved assembly nodes and virion shape-specific adaptations. Our findings shed light on the global organization of mimivirus virion biogenesis and highlight the evolutionary plasticity of viral morphogenetic networks within the Nucleocytoviricota.

microbiology↗

bioRxiv2025

Mechanisms controlling the plasma membrane targeting and the nanodomain organization of the plant SPFH protein HIR2

HIR2 is a plant-specific protein belonging to the superfamily of SPFH domain-containing proteins that were proposed to play scaffolding functions in membranes. HIR2 organizes in plasma membrane (PM) nanodomains that correspond to nanometric scale structures enriched in specific lipids and proteins acting as signaling/regulation hubs. So far, how PM nanodomains are formed and maintained in plant cells remains largely unknown. Combining state of the art microscopy techniques, we investigated the mechanisms governing the trafficking and the organization into nanodomains of Arabidopsis HIR2 protein. We revealed that the mono S-acylation of HIR2 either on C6 or on C7 was required for HIR2 targeting to the PM of Arabidopsis cells, independently of the conventional secretory pathway. Investigating the mechanisms implicated in the arrangement of HIR2 into nanodomains, we provided evidences that the lipid composition in sterols and very long chain fatty acids of the PM influenced HIR2 organization. HIR2 forms oligomers and we demonstrated here that the C-terminal part of HIR2 is required for self-assembly, similarly to animal SPFH proteins. Interestingly, we highlighted that the oligomerization of HIR2 is essential for its organization in nanodomains and to ensure HIR2 lateral stability in the PM. Overall, we propose that HIR2 nanodomain organization is a complex mechanism relying on different parameters including PM lipid composition and oligomerization. HIR proteins are involved in plant immunity. Here, we revealed that HIR2 nanodomain organization is required to boost the apoplastic ROS burst induced by the bacterial peptide flg22. One sentence summaryS-acylation and oligomerization control the plasma membrane targeting and the organization into nanodomains of the Arabidopsis SPFH-domain containing protein HIR2, respectively.

plant biology↗

bioRxiv2023

Comparison of shear stress patterns by the established and advanced reconstruction method incorporating side branches to predict plaque progression

BackgroundComplete vessel reconstruction (CVR) with incorporation of side branches is essential for accurate evaluation of wall shear stress (WSS) distribution. However, CVR is time consuming and blood flow simulation is computationally expensive, while there is no evidence that WSS computed by CVR, enables better prediction of disease progression compared to WSS derived from the conventional single vessel reconstruction (SVR). We aim to compare the WSS in models reconstructed using the CVR and SVR methods and examine its ability to predict disease progression. MethodsPatients who had baseline and 13-months follow-up intravascular ultrasound (IVUS) imaging (n=19 vessels), and with neoatherosclerotic lesions (n=13 vessels) on optical coherence tomography (OCT) were included in the present analysis. All the studied vessels had at least one side branch with diameter >1mm. 3-dimensional (3D) CVR and SVR were performed and time averaged (TAWSS) and multidirectional WSS were computed using pulsatile blood flow simulation and the performance of both methods in predicting disease progression in IVUS and OCT models were assessed. ResultsThe incorporation of side branches in 3D geometry resulted in lower TAWSS in the IVUS (0.821 vs 1.698Pa, p<0.001) and OCT-based reconstructions (0.682 vs 1.325Pa, p<0.001) and influenced the multidirectional WSS distribution. In native segments, WSS metrics estimated by the CVR enabled better prediction of the lumen and plaque area and burden at follow-up than SVR and disease progression defined as decrease in lumen area and increase in plaque burden (AUC CVR 0.712 vs SVR 0.554). In stented segments, multidirectional WSS was associated with neointima area in both CVR and SVR methods, but TAWSS was only a predictor of neointima area in the CVR method. ConclusionsThe incorporation of side branches in vessel reconstruction influences WSS distribution and enables more accurate prediction of disease progression in native and stented segments than SVR modelling. HighlightsO_LIComplete vessel reconstruction (CVR) with incorporation of vessel side branches has been proposed for accurate evaluation of wall shear stress (WSS) distribution compared to the traditional single vessel reconstruction (SVR) method; however, there are no studies comparing the performance of the WSS metrics derived by these methods in predicting atherosclerotic evolution. C_LIO_LIIn vessels with large side branches, the incorporation of the side branches in the vessel geometry reconstructed from angiographic and intravascular imaging data resulted in lower time averaged wall shear stress (TAWSS) and influenced the multidirectional WSS estimations compared to the models reconstructed without the side branches. C_LIO_LIThe WSS metrics estimated in the CVR models enabled better prediction of atherosclerotic disease progression at 13-months follow-up on IVUS than the WSS derived by the SVR. C_LIO_LIIn stented vessels, all the WSS metrics in the CVR and the multidirectional WSS in SVR were associated with neointima tissue development; however, both approaches showed limited efficacy in predicting neointima proliferation. C_LI

bioengineering↗

bioRxiv2022

Origin and dynamics of Mycobacterium tuberculosis subpopulations that predictably generate drug tolerance and resistance

Initial responses to tuberculosis treatment are poor predictors of final therapeutic outcomes in drug-susceptible disease suggesting that treatment success depends on features that are hidden within a small minority of the overall infecting Mycobacterium tuberculosis (Mtb) population. We developed a multi-transwell robotic system to perform numerous parallel cultures of genetically barcoded Mtb exposed to steady-state concentrations of rifampicin to uncover these difficult to eliminate minority populations. We found that tolerance repeatedly emerged from at least two subpopulations of barcoded cells, one that could not grow on solid agar media and a second that could form colonies, but whose kill curves diverged from the general bacterial population within 4 and 16 days of drug exposure, respectively. These tolerant subpopulations reproducibly passed through a phase characterized by multiple unfixed resistance mutations followed by emergent drug resistance in some cultures. Barcodes associated with drug resistance identified an especially privileged subpopulation that was rarely eliminated despite 20 days of drug treatment even in cultures that did not contain any drug resistant mutants. The association of this evolutionary scenario with a defined subset of barcodes across multiple independent cultures suggested a transiently heritable phenotype, and indeed glpK phase variation mutants were associated with up to 16 % of the resistant cultures. Drug tolerance and resistance were eliminated in{Delta} ruvA mutant consistent with the importance of bacterial stress responses. This work provides a window into the origin and dynamics of bacterial drug tolerant subpopulations whose elimination may be critical to developing rapid and resistance free cures. ImportanceTuberculosis is unusual among bacterial diseases in that treatments which can rapidly resolve symptoms do not predictably lead to a durable cure unless treatment is continued for months after all clinical and microbiological signs of disease have been eradicated. Using a novel steady-state antibiotic exposure system combined with chromosomal barcoding, we identified small hidden Mycobacterium tuberculosis subpopulations that repeatedly enter into a state of drug tolerance with a predisposition to develop fixed drug resistance after first developing a cloud of unfixed resistance mutations. The existence of these difficult to eradicate subpopulations may explain the need for extended treatment regimen for tuberculosis. Their identification provides opportunities to test genetic and therapeutic approaches that may result in shorter and more effective TB treatments.

microbiology↗

bioRxiv2019

Phase variation in Mycobacterium tuberculosis glpK produces transiently heritable drug tolerance

The length and complexity of tuberculosis (TB) therapy, as well as the propensity of Mycobacterium tuberculosis to develop drug resistance, are major barriers to global TB control efforts. M. tuberculosis is known to have the ability to enter into a drug-tolerant state, which may explain many of these impediments to TB treatment. We have identified a novel mechanism of genetically encoded but rapidly reversible drug-tolerance in M. tuberculosis caused by transient frameshift mutations in a homopolymeric tract (HT) of seven cytosines (7C) in the glpK gene. Inactivating frameshift mutations associated with the 7C HT in glpK produce small colonies that exhibit heritable multi-drug increases in minimal inhibitory concentrations and decreases in drug-dependent killing; however, reversion back to a fully drug-susceptible large-colony phenotype occurs rapidly through the introduction of additional insertions or deletions in the same glpK HT region. These reversible frameshift mutations in the 7C HT of M. tuberculosis glpK occur in clinical isolates, accumulate in M. tuberculosis infected mice with further accumulation during drug treatment, and exhibit a reversible transcriptional profile including induction of dosR and sigH and repression of kstR regulons, similar to that observed in other in vitro models of M. tuberculosis tolerance. These results suggest that GlpK phase variation may contribute to drug-tolerance, treatment failure and relapse in human TB. Drugs effective against phase-variant M. tuberculosis may hasten TB treatment and improve cure rates.\n\nSIGNIFICANCEThe ability of M. tuberculosis to survive during prolonged treatment has been attributed to either transient stress responses or fixed heritable drug-resistance producing mutations. We show that phase-variation in the M. tuberculosis glpK gene represents a third type of resistance mechanism. The ability of these glpK mutants to grow slowly and then rapidly revert suggests that these transiently-heritable changes may also explain how a hidden population of drug-tolerant bacteria develops during TB treatment. As a genetically trackable cause of drug-tolerance, M. tuberculosis glpK mutants provides a unique opportunity to study these phenomena at a cellular and mechanistic level. These mutants could also be used for developing drugs that target tolerant populations, leading to more rapid and effective TB treatments.

microbiology↗