Biology subjects

Ben-Zion, I.

Publications and source records attributed to Ben-Zion, I..

5 recordsLinked to original sources

bioRxiv2025

A multiplexed, target-based phenotypic screening platform using CRISPR interference in Mycobacterium abscessus

The rise of difficult-to-treat Mycobacterium abscessus infections presents a growing clinical challenge due to the immense arsenal of intrinsic, inducible and acquired antibiotic resistance mechanisms that render many existing antibiotics ineffective against this pathogen. Moreover, the limited success in discovery of novel compounds that inhibit novel pathways underscores the need for innovative drug discovery strategies. Here, we report a strategic advancement in PROSPECT (PRimary screening Of Strains to Prioritize Expanded Chemistry and Targets), which is an antimicrobial discovery strategy that measures chemical-genetic interactions between small molecules and a pool of bacterial mutants, each depleted of a different essential protein target, to identify whole-cell active compounds with high sensitivity. Applying this modified strategy to M. abscessus, in contrast to previously described versions of PROSPECT which utilized protein degradation or promoter replacement strategies for generating engineered hypomorphic strains, here we leveraged CRISPR interference (CRISPRi) to more efficiently generate mutants each depleted of a different essential gene involved in cell wall synthesis or located at the bacterial surface. We applied this platform to perform a pooled PROSPECT pilot screen of a library of 809 compounds using CRISPRi guides as mutant barcodes. We identified a range of active hits, including compounds targeting InhA, a well-known mycobacterial target but under-explored in the M. abscessus space. The unexpected susceptibility to isoniazid, traditionally considered to be ineffective in M. abscessus, suggested a complex interplay of several intrinsic resistance mechanisms. While further complementary efforts will be needed to change the landscape of therapeutic options for M. abscessus, we propose that PROSPECT with CRISPRi engineering provides an increasingly accessible, high-throughput target-based phenotypic screening platform and thus represents an important step towards accelerating early-stage drug discovery.

microbiology↗

bioRxiv2025

Reference-based chemical-genetic interaction profiling to elucidate small molecule mechanism of action in Mycobacterium tuberculosis

In an era of increasing resistance, new and effective strategies are needed for antibiotic discovery. Whole-cell active screens yield candidate compounds lacking mechanism-of-action (MOA) information and thus do not provide biological insight for prioritization. We previously reported PROSPECT (PRimary screening Of Strains to Prioritize Expanded Chemistry and Targets), an antimicrobial discovery strategy that measures chemical-genetic interactions between small molecules and a pool of Mycobacterium tuberculosis mutants, each depleted of a different essential protein target. PROSPECT facilitates efficient hit prioritization by simultaneously identifying whole-cell active compounds with high sensitivity and providing early insights into their MOA. Here, we report a reference-based approach to infer MOA from often complex PROSPECT data. For this aim, we curated a reference set of 437 compounds with published, annotated MOA and known or suspected antitubercular activity, and applied PROSPECT to it. We then developed Perturbagen CLass (PCL) analysis, a computational method that predicts MOA by comparing chemical-genetic interaction profiles of unknown compounds to those of this reference set. In leave-one-out cross-validation, PCL analysis correctly predicted MOA with 70% sensitivity and 75% precision. When applied to 75 antitubercular leads with known MOA previously reported by GlaxoSmithKline (GSK), PCL analysis similarly achieved 69% sensitivity and 87% precision. We also analyzed 98 GSK compounds lacking MOA information, predicting 60 of them to act via a reference MOA, and followed up with functional validation of 29 compounds predicted to target respiration-related MOAs. Finally, we applied PROSPECT and PCL analysis to [~]5,000 compounds from larger unbiased libraries that had not been preselected for antitubercular activity. PCL analysis identified a novel scaffold lacking wild-type activity but predicted to inhibit respiration via QcrB, and we confirmed this prediction while chemically optimizing this scaffold to achieve wild-type activity. PCL analysis of PROSPECT data thus enables rapid MOA assignment and hit prioritization, advancing the discovery of new, potent antitubercular compounds.

microbiology↗

bioRxiv2022

Recalibrating vision-for-action requires years after sight restoration from congenital blindness

Being able to perform goal-directed actions requires predictive, feed-forward control, including a mapping between the visually estimated target locations and the motor commands reaching for them. When the mapping is perturbed, e.g., due to muscle fatigue or optical distortions, we are quickly able to recalibrate the sensorimotor system to update this mapping. Here we investigated whether early visual and visuomotor experience is essential for developing the ability to recalibrate. To this end, we assessed young individuals deprived from vision due to dense congenital bilateral cataracts, who were surgically treated for sight restoration only years after birth. We compared their recalibration performance to such distortion to that of age-matched sighted controls. Their recalibration performance was impaired right after surgery. This finding cannot be explained by their still lower visual acuity, since blurring vision in controls to a matching degree did not lead to comparable behavior. Nevertheless, the recalibration ability of cataract-treated participants improved with experience, matching controls performance after around 2 years from surgery. Thus, the lack of early visual experience affects visuomotor recalibration. However, this ability is not lost but slowly develops after sight restoration, highlighting the importance of sensorimotor experience for brain plasticity late in life.

neuroscience↗

bioRxiv2020

Decoupling transcription factor expression and activity enables dimmer-switch gene regulation

Our mechanistic understanding of how gene regulatory networks can achieve complex biological responses is limited. We find that the galactose-responsive pathway in S. cerevisiae has a complex behavior - the decision to induce is controlled independently from the induction level, resembling a mechanical dimmer-switch. Surprisingly, this behavior is not achieved directly through chromatin regulation or combinatorial control at galactose-responsive promoters. Instead, this behavior is achieved by hierarchical regulation of the expression and catalytic activity of a single transcription factor. This genetic motif allows evolution to independently act on both properties, providing a means to tune both resource allocation strategies in dynamic multi-input environments on both physiological and evolutionary time-scales. Hierarchical regulation is ubiquitous and thus dimmer-switch regulation is likely a key feature of many biological systems.

systems biology↗

bioRxiv2019

Intercellular positive-feedback loops promote the evolutionary stability of microbial cooperative behaviors

Microbial cooperation enables groups of conspecific cells to perform tasks that cannot be performed efficiently by individual cells, such as utilization of various secreted public-good molecules, communication via quorum-sensing, or the formation of multicellular structures. Cooperation is often costly and therefore susceptible to exploitation by cheater cells, which enjoy the benefit of cooperation without investing in it. While population structure is key to the maintenance of cooperation, it remains unclear whether other mechanisms help in stabilizing microbial cooperation. Like other microbial traits, cooperation is often governed by complex regulatory networks, and one reoccurring motif is an intercellular positive-feedback loop, where a secreted molecule, e.g. a public-good or a quorum-sensing signaling molecule, activates its own production in all surrounding cells. Here we investigate the role of intercellular feedbacks in the maintenance of bacterial cooperation. We combine theory with a synthetic-biology approach, using swarming motility of Bacillus subtilis engineered variants, to compare the response of open-loop and feedback cooperators to the presence of cheaters. We find that positive feedbacks on cooperative behaviors - either directly or through a feedback on quorum-sensing - maintain cooperation in a broader range of environments, relieving the requirement for a strong population structure. Our results directly demonstrate the stabilizing effect of intercellular positive feedbacks on cooperative behaviors, and suggests an explanation for their abundance in regulatory networks of bacterial cooperation.

systems biology↗