Genomic disorders, the syndromes with multiple manifestations, may occur sporadically due to unequal recombination in chromosomal regions with specific architecture. Therefore, each patient may carry an individual structural variant of DNA sequence (SV) with small insertions and deletions (INDELs) sometimes less than 10 bp. The transposable elements of the Tc1/mariner superfamily are often associated with hotspots for homologous recombination involved in human genetic disorders, such as Williams Beuren Syndromes (WBS) with LIM-kinase 1-dependent cognitive defects. The Drosophila melanogaster mutant agnts3 has unusual architecture of the agnostic locus harboring LIMK1: it is a hotspot of chromosome breaks, ectopic contacts, underreplication, and recombination. Here, we present the analysis of LIMK1-containing locus sequencing data in agnts3 and three D. melanogaster wild-type strains—Canton-S, Berlin, and Oregon-R. We found multiple strain-specific SVs, namely, single base changes and small INDEls. The specific feature of agnts3 is 28 bp A/T-rich insertion in intron 1 of LIMK1 and the insertion of mobile S-element from Tc1/mariner superfamily residing ~460 bp downstream LIMK1 3′UTR. Neither of SVs leads to amino acid substitutions in agnts3 LIMK1. However, they apparently affect the nucleosome distribution, non-canonical DNA structure formation and transcriptional factors binding. Interestingly, the overall expression of miRNAs including the biomarkers for human neurological diseases, is drastically reduced in agnts3 relative to the wild-type strains. Thus, LIMK1 DNA structure per se, as well as the pronounced changes in total miRNAs profile, probably lead to LIMK1 dysregulation and complex behavioral dysfunctions observed in agnts3 making this mutant a simple plausible Drosophila model for WBS.

Specific recognition of ligands by surface receptors of eukaryotic cells is a fundamental process in sensing of the exogenous environment, including cell-to-cell communication. These interactions are therefore widely probed in both basic studies and drug development to enhance or interrupt them. Here, we designed a high-throughput publicly available platform for visualization and selection of eukaryotic cells according to the specificity of surface-exposed receptors by consolidation of phage display and flow cytometry techniques. Polypeptide ligands for membrane receptors are incorporated into every copy of p3 protein of M13K07 bacteriophage, which is intracellularly biotinylated to further accept PE-Cy7-labled streptavidin. Transgenic antigen-specific B-cells expressing membrane-tethered lymphoid B-cell receptor in a single-chain format interacted with engineered bacteriophages exposing the polypeptide ligand with an unprecedented selectivity of 97% and a false-positive detection value of 2.0%. Multivalent binding of the phage bioconjugates with the receptor provided significantly better specificity and sensitivity allowing application of engineered bacteriophage bioconjugates at a concentration 3 orders of magnitude lower in comparison with synthetic biotinylated peptide. We suggest that the platform described in this work may be applied either for routine staining or characterization of orphan membrane receptors exposed on the surface of living mammalian cells in their native environment.

Using immunofluorescence with specific antibodies, we analyzed DNA hydroxymethylation in uncultured cells from 25 human uterine leiomyomas considering the menstrual cycle phase during surgery and the presence of MED12 gene mutations. It was found that each tumor node had specific DNA hydroxymethylation level that did not depend on the presence of mutations in MED12 gene, but depended on the phase of menstrual cycle. The degree of DNA hydroxymethylation was significantly lower in cells of leiomyomas excised during the luteal phase compared to the follicular phase (p=0.0431). Hormonal status changing at various phases of menstrual cycle is a factor affecting DNA hydroxymethylation in leiomyoma cells.

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