The enzymatic dissociation of human solid tissues is a critical process for disaggregating extracellular matrix and the isolation of individual cells for various applications, including the immortalizing primary cells, creating novel cell lines, and performing flow cytometry and its specialized type, FACS, as well as conducting scRNA-seq studies. Tissue dissociation procedures should yield intact, highly viable single cells that preserve morphology and cell surface markers. However, endocrine tissues, such as adrenal gland tumors, thyroid carcinomas, and pituitary neuroendocrine tumors, present unique challenges due to their complex tissue organization and morphological features. Our study conducted a morphological examination of these tissues, highlighting the intricate structures and secondary degenerative changes that complicate the dissociation process. We investigated the effects of various dissociation parameters, including the types of enzymes, incubation duration, and post-dissociation purification procedures, such as debris removal and nontarget blood cell lysis, on the viability of cells derived from different tumor types. The findings emphasize the importance of optimizing tissue digestion protocols to preserve cell viability and integrity, ensuring reliable outcomes for downstream analyses.

Single-cell RNA sequencing (scRNA-seq) has revolutionized transcriptomic analysis, enabling high-resolution profiling of gene expression across individual cells. However, technical limitations, particularly dropout events, still hinder the accurate quantification of gene expression heterogeneity, resulting in an underestimation of gene-expressing cell proportions. Numerous computational methods have been developed to impute missing expression signals. In this study, we systematically evaluated eight prominent imputation algorithms—MAGIC, SAVER, scVI, DCA, scBiG, kNN-smoothing, scImpute, and ALRA—using two simulated datasets and a deeply sequenced reference dataset. We defined conditions that make it possible to improve estimates of cell population sizes and demonstrate the utility of imputation in biologically relevant contexts, including identifying cell populations susceptible to SARS-CoV-2 infection and detecting CFTR-expressing cells. Our findings highlight the potential and limitations of current imputation strategies and offer practical recommendations for improving the interpretability and accuracy of scRNA-seq data.

Eukaryotic cells are characterized by a high degree of compartmentalization of their internal contents, which ensures precise and controlled regulation of intracellular processes. During many processes, including different stages of transcription, dynamic membraneless compartments termed biomolecular condensates are formed. Transcription condensates contain various transcription factors and RNA polymerase and are formed by high- and low-specificity interactions between the proteins, DNA, and nearby RNA. This review discusses recent data demonstrating important role of nonspecific multivalent protein–protein and RNA–protein interactions in organization and regulation of transcription.

Patients infected with human immunodeficiency virus‐1 (HIV‐1) have an increased incidence of B‐cell lymphoma, even though HIV‐1 does not infect B cells. The development of B‐cell lymphomas appears to be related to the action of the HIV‐1 transactivator protein (Tat), which is released from HIV‐infected cells and penetrates uninfected B cells, affecting host cell gene expression. Upon chronic HIV‐1 infection, Tat acts on the cells for a long time, probably allowing the cells to adapt to the presence of the viral protein. The aim of this work was to identify and study the mechanism of adaptation of cells to prolonged (chronic) exposure to HIV‐1 Tat. We performed a comparative analysis of cells expressing Tat under the action of either an inducible promoter or a constitutive promoter, allowing us to model acute and chronic Tat effects, respectively. We found that the acute action of Tat leads to the suppression of cell proliferation, probably due to the downregulation of genes associated with replication and protein synthesis. In the case of chronic action of Tat, cell proliferation was restored and the expression of genes associated with the implementation of protective (antiviral) functions of the cell was increased. Analysis using proteasome inhibitors showed that in the case of chronic action, intense Tat proteolysis occurred, which could be the main mechanism of B‐cell adaptation. Thus, B cells have a powerful mechanism to adapt to the entry of HIV‐1 Tat, the efficiency of which may determine the frequency of lymphomagenesis in HIV‐1‐infected patients.