The expression profiles of piRNAs and their interacting Piwi proteins in cellular model of renal development: Focus on Piwil1 in mitosis.

Piwi proteins and Piwi interacting RNAs, piRNAs, presented in germline cells play a role in transposon silencing during germline development. In contrast, the role of somatic Piwi proteins and piRNAs still remains obscure. Here, we characterize the expression pattern and distribution of piRNAs in human renal cells in terms of their potential role in kidney development. Further, we show that all PIWI genes are expressed at the RNA level, however, only PIWIL1 gene is detected at the protein level by western blotting in healthy and cancerous renal cells. So far, the expression of human Piwil1 protein has only been shown in testes and cancer cells, but not in healthy somatic cell lines. Since we observe only Piwil1 protein, the regulation of other PIWI genes is probably more intricated, and depends on environmental conditions. Next, we demonstrate that downregulation of Piwil1 protein results in a decrease in the rate of cell proliferation, while no change in the level of apoptotic cells is observed. Confocal microscopy analysis reveals that Piwil1 protein is located in both cellular compartments, cytoplasm and nucleus in renal cells. Interestingly, in nucleus region Piwil1 is observed close to the spindle during all phases of mitosis in all tested cell lines. It strongly indicates that Piwil1 protein plays an essential role in proliferation of somatic cells. Moreover, involvement of Piwil1 in cell division could, at least partly, explain invasion and metastasis of many types of cancer cells with upregulation of PIWIL1 gene expression. It also makes Piwil1 protein as a potential target in the anticancer therapy.

Characteristics of Transfer RNA-Derived Fragments Expressed during Human Renal Cell Development: The Role of Dicer in tRF Biogenesis.

tRNA-derived fragments participate in the regulation of many processes, such as gene silencing, splicing and translation in many organisms, ranging from bacteria to humans. We were interested to know how tRF abundance changes during the different stages of renal cell development. The research model used here consisted of the following human renal cells: hESCs, HEK-293T, HK-2 and A-489 kidney tumor cells, which, together, mimic the different stages of kidney development. The characteristics of the most abundant tRFs, tRFGly(CCC), tRFVal(AAC) and tRFArg(CCU), were presented. It was found that these parental tRNAs present in cells are the source of many tRFs, thus increasing the pool of potential regulatory RNAs. Indeed, a bioinformatic analysis showed the possibility that tRFGly(CCC) and tRRFVal(AAC) could regulate the activity of a range of kidney proteins. Moreover, the distribution of tRFs and the efficiency of their expression is similar in adult and embryonic stem cells. During the formation of tRFs, HK-2 cells resemble A-498 cancer cells more than other cells. Additionally, we postulate the involvement of Dicer nuclease in the formation of tRF-5b in all the analyzed tRNAs. To confirm this, 293T NoDice cells, which in the absence of Dicer activity do not generate tRF-5b, were used.

Long Non-Coding RNA Epigenetics.

Long noncoding RNAs exceeding a length of 200 nucleotides play an important role in ensuring cell functions and proper organism development by interacting with cellular compounds such as miRNA, mRNA, DNA and proteins. However, there is an additional level of lncRNA regulation, called lncRNA epigenetics, in gene expression control. In this review, we describe the most common modified nucleosides found in lncRNA, 6-methyladenosine, 5-methylcytidine, pseudouridine and inosine. The biosynthetic pathways of these nucleosides modified by the writer, eraser and reader enzymes are important to understanding these processes. The characteristics of the individual methylases, pseudouridine synthases and adenine-inosine editing enzymes and the methods of lncRNA epigenetics for the detection of modified nucleosides, as well as the advantages and disadvantages of these methods, are discussed in detail. The final sections are devoted to the role of modifications in the most abundant lncRNAs and their functions in pathogenic processes.

Human Long Noncoding RNA Interactome: Detection, Characterization and Function.

The application of a new generation of sequencing techniques has revealed that most of the genome has already been transcribed. However, only a small part of the genome codes proteins. The rest of the genome "dark matter" belongs to divergent groups of non-coding RNA (ncRNA), that is not translated into proteins. There are two groups of ncRNAs, which include small and long non-coding RNAs (sncRNA and lncRNA respectively). Over the last decade, there has been an increased interest in lncRNAs and their interaction with cellular components. In this review, we presented the newest information about the human lncRNA interactome. The term lncRNA interactome refers to cellular biomolecules, such as nucleic acids, proteins, and peptides that interact with lncRNA. The lncRNA interactome was characterized in the last decade, however, understanding what role the biomolecules associated with lncRNA play and the nature of these interactions will allow us to better understand lncRNA's biological functions in the cell. We also describe a set of methods currently used for the detection of lncRNA interactome components and the analysis of their interactions. We think that such a holistic and integrated analysis of the lncRNA interactome will help to better understand its potential role in the development of organisms and cancers.

tRNA-derived fragments from the Sus scrofa tissues provide evidence of their conserved role in mammalian development.

The recently discovered group of noncoding RNAs, which are fragments of tRNA molecules (tRFs), has not been fully characterized and its potential functions still require investigation. Porcine tRFs were characterized and compared to mouse and human tRFs. Two tRFs, 5' 32-33 nt and 3' 41-42 nt that are derived from the mature tRNAVal(CAC) and tRNAGly(GCC) were detected with the use of bioinformatics and the Northern blot method. The abundance of these tRFs in the case of Sus scrofa is restricted to the ovary and the kidney. The same tRFs were found in human cancer cells and in mouse sperm, circulating blood and its serum. The binding of selected sncRNAs (piRNA, 5'tRFVal(CAC) and miRNA) to the overexpressed PAZ domain of the PIWIL4 protein was also studied. It is noteworthy that porcine 5'tRFVal(CAC) and human 5'tRFVal(CAC)as well as 5'tRFGly(GCC) are bound to the PIWIL4 protein. The potential role of the analyzed tRFs in the development of mammals is also discussed.