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MicroRNAs (miRNAs) are small, non-coding RNAs about 22 nucleotides in length that regulate the expression of target genes post-transcriptionally, and are highly involved in cancer progression. They are able to impact a variety of cell processes such as proliferation, apoptosis and differentiation and can consequently control tumor initiation, tumor progression and metastasis formation. miRNAs can regulate, at the same time, metabolic gene expression which, in turn, influences relevant traits of malignancy such as cell adhesion, migration and invasion. Since the interaction between metabolism and adhesion or cell movement has not, to date, been well understood, in this review, we will specifically focus on miRNA alterations that can interfere with some metabolic processes leading to the modulation of cancer cell movement. In addition, we will analyze the signaling pathways connecting metabolism and adhesion/migration, alterations that often affect cancer cell dissemination and metastasis formation.

Adhesion between cells and the extracellular matrix (ECM) is one of the prerequisites for multicellularity, motility, and tissue specialization. Focal adhesions (FAs) are defined as protein complexes that mediate signals from the ECM to major components of the cytoskeleton (microtubules, actin, and intermediate filaments), and their mutual communication determines a variety of cellular processes. In this study, human cytoskeletal crosstalk proteins were identified by comparing datasets with experimentally determined cytoskeletal proteins. The spectraplakin dystonin was the only protein found in all datasets. Other proteins (FAK, RAC1, septin 9, MISP, and ezrin) were detected at the intersections of FAs, microtubules, and actin cytoskeleton. Homology searches for human crosstalk proteins as queries were performed against a predefined dataset of proteomes. This analysis highlighted the importance of FA communication with the actin and microtubule cytoskeleton, as these crosstalk proteins exhibit the highest degree of evolutionary conservation. Finally, phylogenetic analyses elucidated the early evolutionary history of spectraplakins and cortical microtubule stabilization complexes (CMSCs) as model representatives of the human cytoskeletal crosstalk. While spectraplakins probably arose at the onset of opisthokont evolution, the crosstalk between FAs and microtubules is associated with the emergence of metazoans. The multiprotein complexes contributing to cytoskeletal crosstalk in animals gradually gained in complexity from the onset of metazoan evolution.

Sun or therapy-related ultraviolet B (UVB) irradiation induces different cell death modalities such as apoptosis, necrosis/necroptosis and autophagy. Understanding of mechanisms implicated in regulation and execution of cell death program is imperative for prevention and treatment of skin diseases. An essential component of death-inducing complex is Fas-associated protein with death domain (FADD), involved in conduction of death signals of different death modalities. The purpose of this study was to enlighten the role of FADD in the selection of cell death mode after narrow-band UVB (NB-UVB) irradiation using specific cell death inhibitors (carbobenzoxy-valyl-alanyl-aspartyl-[O-methyl]- fluoromethylketone (zVAD-fmk), Necrostatin-1 and 3-Methyladenine) and FADD-deficient (FADD−/−) mouse embryonic fibroblasts (MEFs) and their wild type (wt) counterparts. The results imply that lack of FADD sensitized MEFs to induction of receptor–interacting protein 1 (RIPK1)-dependent apoptosis by the generation of reactive oxygen species (ROS), but without activation of the proteins p53, Bax and Bcl-2 as well as without the enrolment of calpain-2. Autophagy was established as a contributing factor to NB-UVB-induced death execution. By contrast, wt cells triggered intrinsic apoptotic pathway that was resistant to the inhibition by zVAD-fmk and Necrostatin-1 pointing to the mechanism overcoming the cell survival. These findings support the role of FADD in prevention of autophagy-dependent apoptosis.

The expression of Ikaros family transcription factors and consequently their signalling pathway is limiting for hematopoietic and lymphocyte development in mice and human. Due to their importance, these transcription factors are highly homologous between species. As an initial approach to examining the possible involvement of Ikaros transcription factors in pathogenesis of rat lymphoid development, we analyzed the expression of all known Ikaros family members, Ikaros, Aiolos, Helios, Eos and Pegasus in the rat thymus. We established a semi-quantitative RT-PCR to detect mRNA of each transcription factor. For the first time we give evidence of the expression of Ikaros family transcription factors in the rat thymus. Further, we evaluated whether their mRNA expression was succumbed to changes when the rats were exposed to ethanol, as a known debilitating agent during development. Therefore we analyzed the thymus of adult rats whose mothers were forced to drink ethanol during gestation, to detect possible changes in thymus mRNA expression levels of Ikaros, Aiolos, Helios, Eos and Pegasus. We found that rats prenatally exposed to ethanol show a slightly higher expression of Ikaros family transcription factors in the adult thymus when compared to control rats, but these differences were not statistically significant. We further studied the distribution of the major lymphocyte subpopulations in the rat thymus according to CD3, CD4 and CD8 expression by four color flow cytometry. We found a higher incidence of CD3 positive cells in the double positive, CD4+CD8+ thymic subpopulation of rats prenatally exposed to ethanol when compared to non-exposed animals. Our findings indicate that ethanol exposure of pregnant rats might influence the development of CD3 positive cells in the thymus of the offspring but this result should be further tackled at the level of transcription factor expression.

Elucidation of molecular pathways involved in development of human lymphoma requires efficient methods for tackling gene expression in lymph nodes. Expression studies of transcription factors in these malignancies facilitate understanding the changes occurring in neoplastic transformation and lymphoma development. Excised lymph nodes are routinely fixed in formalin and embedded in paraffin for diagnosis and stored in many hospitals' pathology archives. These tissues represent a precious resource for research since they allow retrospective studies to cover a broad range of human lymphoma even the less frequent types. Reverse transcription polymerase chain reaction (RT-PCR) is a commonly used method for gene expression analysis and a reproducible protocol for RNA isolation from lymph nodes is an inevitable requirement for these studies. However, formalin fixation and paraffin-embedding interfere with the quality of RNA especially when isolated from lymph nodes being the most fragile lymphytic tissues. We present here a simple and fast method for RNA isolation from formalin-fixed paraffin- embedded lymph nodes that can be successfully applied for RT-PCR as well as for quantitative RT- PCR analysis. We tested diverse isolation reagents and combined a range of factors in order to get a high quality RNA for retrospective studies of gene expression in human lymphoma samples. Our modified method of RNA extraction from FFPE provide superior yields and purity based on qPCR data.

Transcription factors from the Ikaros family are involved in lymphocyte differentiation and have a critical role at specific check points of the haemopoietic pathway. However, how developmentally regulated changes are reflected in gene expression programs of lymphocyte differentiation is not well understood. It has been suggested that disregulation of transcription factors from the Ikaros family is associated with the development of different human leukemias. In this work we analyzed the state of Ikaros family members in different leukemic cells with the aim to explore the transcriptional control of human hematopoietic lineages and shed some new light on our understanding of transcription factor significance in human leukemias. By means of RT-PCR and specific primers we investigated the expression of Ikaros, Aiolos and Helios transcription factors and their splicing variants in seven leukemia cell lines derived from different types of leukemia (ALL, CML, AML) and lymphoma (histiocytic lymphoma, Burkitt lymphoma and anaplastic large cell lymphoma). In all of the cell lines examined Ikaros was present in dominant Ik1 to Ik4 isoforms and small Ik6 isoform was absent. Aiolos was expressed in the majority of the cell lines, of both, B and T origin, in the form of the full length Aio1. Helios was also present only in two long isoforms Hel1 and Hel2, and was absent in one third of the lines. Similar distribution of positive and negative expression of Aiolos and Helios found in various types of leukemias could implicate common pathways of their regulation.