Biology subjects

Veksler-Lublinsky, I.

Publications and source records attributed to Veksler-Lublinsky, I..

6 recordsLinked to original sources

bioRxiv2026

Context-dependent determinants of CRISPR-Cas9 editing efficiency revealed through cross-species endogenous editing analysis

Accurate prediction of CRISPR-Cas9 guide RNA (gRNA) editing efficiency remains limited, particularly outside human systems, where models trained on exogenous human datasets show poor generalization. We analyzed Cas9 efficiency and repair outcomes using novel endogenous editing data from four human cell types, two tomato cell types, and cells from giant river prawn and black soldier fly. While integrating publicly available predictors via ensemble frameworks improved performance, our analysis revealed hundreds of novel features affecting activity. Crucially, dominant features related to sites competition for gRNA, and local geometric properties varied across systems, highlighting the strong context dependence of Cas9 efficiency and arguing against a universal model. Interestingly, codon usage bias-based features also emerged as informative predictors, as they are proxies for chromatin accessibility. In contrast, trends in repair outcomes remained conserved. This work provides essential resources for more generalizable CRISPR guide design.

synthetic biology↗

bioRxiv2025

Defending Synthetic DNA Orders Against Splitting-Based Obfuscation

Biosecurity screening of synthetic DNA orders is a key defense against malicious actors and careless enthusiasts producing dangerous pathogens or toxins. It is important to evaluate biosecurity screening tools for potential vulnerabilities and to work responsibly with providers to ensure that vulnerabilities can be patched before being publicly disclosed. Here, we consider a class of potential vulnerabilities in which a DNA sequence is obfuscated by splitting it into two or more fragments that can be readily joined via routine biological mechanisms such as restriction enzyme digestion or splicing. We evaluated this potential vulnerability by developing a test set of obfuscated sequences based on controlled venoms, sharing these materials with the biosecurity screening community, and collecting test results from open source and commercial biosecurity screening tools, as well as a novel Gene Edit Distance algorithm specifically designed to be robust against splitting-based obfuscations.

bioinformatics↗

bioRxiv2024

Identification and developmental profiling of microRNAs in the acoel worm Hofstenia miamia

The acoel worm Hofstenia miamia (H. miamia) has recently emerged as a model organism for studying whole-body regeneration and embryonic development, providing an opportunity to probe post-transcriptional mechanisms in both processes in the same organism. Here, we establish a resource for studying H. miamia microRNA-mediated gene regulation, a major aspect of post-transcriptional control in animals. We generated a PacBio long-read sequencing-based genome assembly. We developed a stringent microRNA annotation framework for H. miamia using small RNA-sequencing samples spanning key developmental stages. Our analysis uncovered 545 microRNA loci, including 154 highest-confidence loci based on structural features, expression levels, and prediction quality metrics. Comparison of microRNA seed sequences with those in other bilaterian species revealed that H. miamia encodes many known conserved bilaterian microRNA families and that several microRNA families previously reported only in protostomes or deuterostomes likely have ancient bilaterian origins. We profiled and characterized the expression dynamics and strand preference of microRNAs in H. miamia embryonic and post-embryonic development. An intron that is spliced in the primary transcript of co-transcribed let-7 and mir-125 microRNAs. To generate hypotheses for microRNA function, we annotated the 3 UTRs of H. miamia protein-coding genes and performed microRNA target site predictions. Focusing on genes that are known to function in the wound response, posterior patterning, and neural differentiation in H. miamia, we found that these processes may be under substantial microRNA regulation. Notably, we found that microRNAs in MIR-7 and MIR-9 families, which have target sites in the posterior genes fz-1, wnt-3, and sp5 are indeed expressed in the anterior of the animal, consistent with an anterior-biased repressive effect on their corresponding target genes. Our annotation provides a resource for future studies of post-transcriptional regulation of gene expression during development and regeneration.

genomics↗

bioRxiv2021

Comprehensive machine-learning-based analysis of microRNA-target interactions reveals variable transferability of interaction rules across species

BackgroundMicroRNAs (miRNAs) are small non-coding RNAs that regulate gene expression post-transcriptionally via base-pairing with complementary sequences on messenger RNAs (mRNAs). Due to the technical challenges involved in the application of high-throughput experimental methods, datasets of direct bona-fide miRNA targets exist only for a few model organisms. Machine learning (ML) based target prediction methods were successfully trained and tested on some of these datasets. There is a need to further apply the trained models to organisms where experimental training data is unavailable. However, it is largely unknown how the features of miRNA-target interactions evolve and whether there are features that have been fixed during evolution, questioning the general applicability of these ML methods across species. ResultsIn this paper, we examined the evolution of miRNA-target interaction rules and used data science and ML approaches to investigate whether these rules are transferable between species. We analyzed eight datasets of direct miRNA-target interactions in four organisms (human, mouse, worm, cattle). Using ML classifiers, we achieved high accuracy for intra-dataset classification and found that the most influential features of all datasets significantly overlap. To explore the relationships between datasets we measured the divergence of their miRNA seed sequences and evaluated the performance of cross-datasets classification. We showed that both measures coincide with the evolutionary distance of the compared organisms. ConclusionsOur results indicate that the transferability of miRNA-targeting rules between organisms depends on several factors, the most associated factors being the composition of seed families and evolutionary distance. Furthermore, our feature importance results suggest that some miRNA-target features have been evolving while some have been fixed during evolution. Our study lays the foundation for the future developments of target prediction tools that could be applied to "non-model" organisms for which minimal experimental data is available. Availability and implementation The code is freely available at https://github.com/gbenor/TPVOD

bioinformatics↗

bioRxiv2021

Critical contribution of 3' non-seed base pairing to the in vivo function of the evolutionarily conserved let-7a microRNA

Base-pairing of the seed region (g2-g8) is essential for microRNA targeting, however, the in vivo function of the 3 non-seed region (g9-g22) are less well understood. Here we report the first systematic investigation of the in vivo roles of 3 non-seed nucleotides in microRNA let-7a, whose entire g9-g22 region is conserved among bilaterians. We found that the 3 non-seed sequence functionally distinguishes let-7a from its family paralogs. The complete pairing of g11-g16 is essential for let-7a to fully repress multiple key targets, including evolutionarily conserved lin-41, daf-12 and hbl-1. Nucleotides at g17-g22 are less critical but may compensate for mismatches in the g11-g16 region. Interestingly, the 3 non-seed pairing of let-7a can be critically required with certain minimal complementarity for sites with perfect seed pairing. These results provide evidence that the specific configurations of both seed and 3 non-seed base-pairing can critically influence microRNA-mediated gene regulation in vivo.

molecular biology↗

bioRxiv2019

HRPK-1, a conserved KH-domain protein, modulates microRNA activity during Caenorhabditis elegans development

microRNAs (miRNAs) are potent regulators of gene expression that function in diverse developmental and physiological processes. Argonaute proteins loaded with miRNAs form the miRNA Induced Silencing Complexes (miRISCs) that repress gene expression at the post-transcriptional level. miRISCs target genes through partial sequence complementarity between the miRNA and the target mRNAs 3 UTR. In addition to being targeted by miRNAs, these mRNAs are also extensively regulated by RNA-binding proteins (RBPs) through RNA processing, transport, stability, and translation regulation. While the degree to which RBPs and miRISCs functionally interact to regulate gene expression is likely extensive, we have only begun to unravel these functional interactions. An RNAi-based screen of putative ALG-1 Argonaute interactors has identified a role for a conserved RNA binding protein, HRPK-1, in modulating miRNA activity during C. elegans development. Here, we report the physical and genetic interaction between HRPK-1 and ALG-1/miRNAs. Specifically, we report the genetic and molecular characterizations of hrpk-1 and its role in C. elegans development and miRNA-mediated target repression. We show that loss of hrpk-1 causes numerous developmental defects and enhances the mutant phenotypes associated with reduction of miRNA activity, including those of lsy-6, mir-35-family, and let-7-family miRNAs. In addition to hrpk-1 genetic interaction with these miRNA families, hrpk-1 is required for efficient regulation of lsy-6 target cog-1. We report that hrpk-1 may play a role in miRNA processing but is not globally required for mature miRNA biogenesis or ALG-1/AIN-1 miRISC assembly and confirm HRPK-1 ability to co-precipitate with ALG-1. We suggest that HRPK-1 may functionally interact with miRNAs on multiple levels to enhance miRNA/miRISC gene regulatory activity and present several models for its activity.\n\nAuthor summarymicroRNAs are small non-coding RNAs that regulate gene expression at the post-transcriptional level. The core microRNA Induced Silencing Complex (miRISC), composed of Argonaute, mature microRNA, and GW182 protein effector, assembles on the target messenger RNA and inhibits translation or leads to messenger RNA degradation. RNA binding proteins interface with miRNA pathways on multiple levels to coordinate gene expression regulation. Here, we report identification and characterization of HRPK-1, a conserved RNA binding protein, as a physical and functional interactor of miRNAs. We confirm the physical interaction between HRPK-1, an hnRNPK homolog, and Argonaute ALG-1. We report characterizations of hrpk-1 role in development and its functional interactions with multiple miRNA families. We suggest that HRPK-1 promotes miRNA activity on multiple levels in part by contributing to miRNA processing and by coordinating with miRISC at the level of target RNAs. This work contributes to our understanding of how RNA binding proteins and auxiliary miRNA cofactors may interface with miRNA pathways to modulate miRNA gene regulatory activity.

genetics↗