Acute myeloid leukemia (AML) is a rapidly progressing heterogeneous disease with a high mortality rate, which is characterized by hyperproliferation of atypical immature myeloid cells. The number of AML patients is expected to increase in the near future, due to the old-age-associated nature of AML and increased longevity in the human population. RUNX1 and CEBPA, key transcription factors (TFs) of hematopoiesis, are frequently and independently mutated in AML. RUNX1 and CEBPA can bind TET2 demethylase and attract it to their binding sites (TFBS) in cell lines, leading to DNA demethylation of the regions nearby. Since TET2 does not have a DNA-binding domain, TFs are crucial for its guidance to target genomic locations. In this paper, we show that RUNX1 and CEBPA mutations in AML patients affect the methylation of important regulatory sites that resulted in the silencing of several RUNX1 and CEBPA target genes, most likely in a TET2-dependent manner. We demonstrated that hypermethylation of TFBS in AML cells with RUNX1 mutations was associated with resistance to anticancer chemotherapy. Demethylation therapy restored expression of the RUNX1 target gene, BIK, and increased sensitivity of AML cells to chemotherapy. If our results are confirmed, mutations in RUNX1 could be an indication for prescribing the combination of cytotoxic and demethylation therapies.

Fertilized teleost fish eggs are a complex formation, in which dividing cells are located in a small point in the entire volume of eggs. Isolating embryonic cells can be considered a necessary step in the research of developmental peculiarities of fish cells at the earliest stages of embryogenesis before embryo formation. The main advantages of the offered protocol are rapid isolation, no enzymes, and overall low cost compared to other protocols. The protocol is suitable for the isolation of embryonic cells from medium-sized eggs at the stages of blastula or gastrula, for studies in a variety of applications (e.g., microscopy, flow cytometry, and other methods). Fertilized nelma eggs (Stenodus leucichthys nelma) are used in the protocol as a model. Key features Fast and cheap isolation of cells from fish eggs at early stages (blastula or gastrula). Applicable for most of the methods for cell study (any staining, microscopy, flow cytometry, etc.). Can be applied to other teleost fish eggs with medium egg diameter of 3-4mm.

OmpF porin from the outer membrane ofYersinia pseudotuberculosiswas cloned into pET-40b(+) plasmid. UsingE. coliRosetta (DE3) strain, MX medium, IPTG concentration of 0.2 mmand post-induction cultivation at 14°C overnight allowed us to obtain a water-soluble form of the recombinant protein (rs-OmpF). Rs-OmpF was shown to have the ordered spatial structure at the levels of secondary and tertiary structure. Rs-OmpF was found to be effective as diagnostic antigen in ELISA for pseudotuberculosis diagnostics.