Recent advances in imaging flow cytometry (IFC) have revolutionized high-throughput multiparameter analyses at single-cell resolution. Although enabling the discovery of population heterogeneities and the detection of rare events, IFC generates hyperdimensional datasets that demand innovative analytical approaches. Current methods work in a supervised manner, utilize only limited information content, or require large annotated reference datasets. Dimensionality reduction algorithms, including uniform manifold approximation and projection (UMAP), have been successfully applied to analyze the large number of parameters generated in various high-throughput techniques.

RNA degradation is among the most fundamental processes that occur in living cells. The continuous decay of RNA molecules is associated not only with nucleotide turnover, but also with transcript maturation and quality control. The efficiency of RNA decay is ensured by a broad spectrum of both specific and non-specific ribonucleases. Some of these ribonucleases participate mainly in processing primary transcripts and in RNA quality control. Others preferentially digest mature, functional RNAs to yield a variety of molecules that together constitute the RNA degradome. Recently, it has become increasingly clear that the composition of the cellular RNA degradome can be modulated by numerous endogenous and exogenous factors (e.g. by stress). In addition, instead of being hydrolyzed to single nucleotides, some intermediates of RNA degradation can accumulate and function as signalling molecules or participate in mechanisms that control gene expression. Thus, RNA degradation appears to be not only a process that contributes to the maintenance of cellular homeostasis but also an underestimated source of regulatory molecules.

Identification of biomarkers that could be used for the prediction of the response to neoadjuvant radiotherapy (neo-RT) in locally advanced rectal cancer remains a challenge addressed by different experimental approaches. Exosomes and other classes of extracellular vesicles circulating in patients’ blood represent a novel type of liquid biopsy and a source of cancer biomarkers. Here, we used a combined proteomic and metabolomic approach based on mass spectrometry techniques for studying the molecular components of exosomes isolated from the serum of rectal cancer patients with different responses to neo-RT. This allowed revealing several proteins and metabolites associated with common pathways relevant for the response of rectal cancer patients to neo-RT, including immune system response, complement activation cascade, platelet functions, metabolism of lipids, metabolism of glucose, and cancer-related signaling pathways. Moreover, the composition of serum-derived exosomes and a whole serum was analyzed in parallel to compare the biomarker potential of both specimens. Among proteins that the most properly discriminated good and poor responders were GPLD1 (AUC = 0.85, accuracy of 74%) identified in plasma as well as C8G (AUC = 0.91, accuracy 81%), SERPINF2 (AUC = 0.91, accuracy 79%) and CFHR3 (AUC = 0.90, accuracy 81%) identified in exosomes. We found that the proteome component of serum-derived exosomes has the highest capacity to discriminate samples of patients with different responses to neo-RT when compared to the whole plasma proteome and metabolome. We concluded that the molecular components of exosomes are associated with the response of rectal cancer patients to neo-RT and could be used for the prediction of such response.

It is becoming increasingly evident that the RNA degradome is a crucial component of the total cellular RNA pool. Here, we present an analysis of the medium-sized RNAs (midi RNAs) that form in Arabidopsis thaliana. Our analyses revealed that the midi RNA fraction contained mostly 20–70-nt-long fragments derived from various RNA species, including tRNA, rRNA, mRNA and snRNA. The majority of these fragments could be classified as stable RNA degradation intermediates (RNA degradants). Using two dimensional polyacrylamide gel electrophoresis, we demonstrated that high copy number RNA (hcn RNA) degradants appear in plant cells not only during stress, as it was earlier suggested. They are continuously produced also under physiological conditions. The data collected indicated that the accumulation pattern of the hcn RNA degradants is organ-specific and can be affected by various endogenous and exogenous factors. In addition, we demonstrated that selected degradants efficiently inhibit translation in vitro. Thus, the results of our studies suggest that hcn RNA degradants are likely to be involved in the regulation of gene expression in plants.