Non-Coding RNA, Volume 8, Issue 3 (June 2022) – 15 articles

Podocytes, alternatively called glomerular epithelial cells, are terminally differentiated cells that wrap around glomerular capillaries and function as a part of the glomerular filtration barrier in the kidney. Therefore, podocyte injury with morphological alteration and detachment from glomerular capillaries leads to severe proteinuria and subsequent renal failure through glomerulosclerosis. Previous RNA sequencing analysis of primary rat podocytes exposed to puromycin aminonucleoside (PAN), a well-known experimental model of injured podocytes, identified several transcripts as being aberrantly expressed. However, how the expression of these transcripts is regulated remains unclear. MicroRNAs (miRNAs) are small noncoding RNAs that posttranscriptionally inhibit the expression of their target transcripts. In this study, using small RNA sequencing analysis, miR-217-5p was identified as the most upregulated transcript in PAN-treated rat podocytes. MiR-217-5p overexpression in E11 podocyte cells led to shrunken cells with abnormal actin cytoskeletons. Consistent with these changes in cell morphology, gene ontology (GO) enrichment analysis showed that interactive GO terms related to cell morphogenesis were enriched with the predicted targets of miR-217-5p. Of the predicted targets highly downregulated by PAN, Myosin 1d (Myo1d) is a nonmuscle myosin predicted to be involved in actin filament organization and thought to play a role in podocyte morphogenesis and injury. We demonstrated that miR-217-5p targets Myo1d by luciferase assays, qRT–PCR, and Western blotting. Furthermore, we showed that miR-217-5p was present in urine from PAN- but not saline-administrated rats. Taken together, our data suggest that miR-217-5p may serve as a therapeutic target and a biomarker for podocyte injury. Full article

Angiotensin-converting enzyme 2 (ACE2) plays a role in catalyzing angiotensin II conversion to angiotensin (1–7), which often counteracts the renin-angiotensin system. ACE2 is expressed not only in the cells of peripheral tissues such as the heart and kidney, but also in those of the central nervous system (CNS). Additionally, ACE2 acts as the receptor required for the entry of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), whose binding leads to endocytotic recycling and possible degradation of the ACE2 proteins themselves. One of the target cells for SARS-CoV-2 in the CNS is oligodendrocytes (oligodendroglial cells), which wrap neuronal axons with their differentiated plasma membranes called myelin membranes. Here, for the first time, we describe the role of ACE2 in FBD-102b cells, which are used as the differentiation models of oligodendroglial cells. Unexpectedly, RNA knockdown of ACE2 with CasRx-mediated gRNA or the cognate siRNA promoted oligodendroglial cell morphological differentiation with increased expression or phosphorylation levels of differentiation and/or myelin marker proteins, suggesting the negative role of ACE2 in morphological differentiation. Notably, ACE2′s intracellular region preferentially interacted with the active GTP-bound form of Ras. Thus, knockdown of ACE2 relatively increased GTP-bound Ras in an affinity-precipitation assay. Indeed, inhibition of Ras resulted in decreasing both morphological differentiation and expression or phosphorylation levels of marker proteins, confirming the positive role of Ras in differentiation. These results indicate the role of ACE2 itself as a negative regulator of oligodendroglial cell morphological differentiation, newly adding ACE2 to the list of regulators of oligodendroglial morphogenesis as well as of Ras-binding proteins. These findings might help us to understand why SARS-CoV-2 causes pathological effects in the CNS. Full article

This study explored the relationship between 3D genome organization and RNA–DNA triplex-forming sites of long noncoding RNAs (lncRNAs), a group of RNAs that do not code for proteins but are important factors regulating different aspects of genome activity. The triplex-forming sites of anti-sense cardiac lncRNA GATA6-AS1 derived from DBD-Capture-Seq were examined and compared to modular features of 3D genome organization called topologically associated domains (TADs) obtained from Hi-C data. It was found that GATA6-AS1 triplex-forming sites are positioned non-randomly in TADs and their boundaries. The triplex sites showed a preference for TAD boundaries over internal regions of TADs. Computational prediction analysis indicated that CTCF, the key protein involved in TAD specification, may interact with GATA6-AS1, and their binding sites correlate with each other. Examining locations of repeat elements in the genome suggests that the ability of lncRNA GATA6-AS1 to form triplex sites with many genomic locations may be achieved by the rapid expansion of different repeat elements. Some of the triplex-forming sites were found to be positioned in regions that undergo dynamic chromatin organization events such as loss/gain of TAD boundaries during cardiac differentiation. These observed associations suggest that lncRNA–DNA triplex formation may contribute to the specification of TADs in 3D genome organization. Full article

Successful immunotherapy in both solid tumors and in hematological malignancies relies on the ability of T lymphocytes to infiltrate the cancer tissue and mount an immune response against the tumor. Biomarkers able to discern the amount and the types of T lymphocytes infiltrating a given tumor therefore have high diagnostic and prognostic value. Given that lncRNAs are known to have a highly cell-type-specific expression pattern, we searched for lncRNAs specifically expressed by activated T cells and at the same time in a kind of lymphoma, follicular lymphoma, where the microenvironment is known to play a critical role in the regulation of antitumor immunity. We focused on a non-coding transcript, annotated as LINC00892, which reaches extremely high expression levels following cell activation in Jurkat cells. Interestingly LINC00892 has an expression pattern resembling that of genes involved in T cell memory. Accordingly, LINC00892 is mostly expressed by the effector memory and helper CD4+ T cell sub-types but not by naïve T cells. In situ analyses of LINC00892 expression in normal lymph nodes and in follicular lymphoma biopsies show that its expression is limited to CD4+ PD1^hi T cells, with a subcellular localization within the germinal center matching that of follicular helper T cells. Our analysis therefore suggests that the previously uncharacterized lncRNA LINC00892 could be a useful biomarker for the detection of CD4+ memory T cells in both normal and tumor tissues. Full article

In recent years, long non-coding RNAs (lncRNAs) have been shown to play important regulatory roles in cellular processes. Growth arrests specific transcript 5 (GAS5) is a lncRNA that is highly expressed during the cell cycle arrest phase but is downregulated in actively growing cells. Growth arrests specific transcript 5 was discovered to be downregulated in several cancers, primarily solid tumors, and it is known as a tumor suppressor gene that regulates cell proliferation, invasion, migration, and apoptosis via multiple molecular mechanisms. Furthermore, GAS5 polymorphism was found to affect GAS5 expression and functionality in a cell-specific manner. This review article focuses on GAS5’s tumor-suppressive effects in regulating oncogenic signaling pathways, cell cycle, apoptosis, tumor-associated genes, and treatment-resistant cells. We also discussed genetic polymorphisms of GAS5 and their association with cancer susceptibility. Full article

Circular RNAs (circRNAs) are a novel class of regulatory RNA involved in many biological, physiological and pathological processes by functioning as a molecular sponge, transcriptional/epigenetic/splicing regulator, modulator of protein–protein interactions, and a template for encoding proteins. Cells are constantly dealing with stimuli from the microenvironment, and proper responses rely on both the precise control of gene expression networks and protein–protein interactions at the molecular level. The critical roles of circRNAs in the regulation of these processes have been heavily studied in the past decades. However, how the microenvironmental stimulation controls the circRNA biogenesis, cellular shuttling, translation efficiency and degradation globally and/or individually remains largely uncharacterized. In this review, how the impact of major microenvironmental stresses on the known transcription factors, splicing modulators and epitranscriptomic regulators, and thereby how they may contribute to the regulation of circRNAs, is discussed. These lines of evidence will provide new insight into how the biogenesis and functions of circRNA can be precisely controlled and targeted for treating human diseases. Full article

Transfer RNA (tRNA)-derived small RNAs (tsRNAs) belong to a group of transfer ribonucleic acid (tRNA)-derived fragments that have recently gained interest as molecules with specific biological functions. Their involvement in the regulation of physiological processes and pathological phenotypes suggests molecular roles similar to those of miRNAs. tsRNA biogenesis under specific physiological conditions will offer new perspectives in understanding diseases, and may provide new sources for biological marker design to determine and monitor the health status of farm animals. In this review, we focus on the latest discoveries about tsRNAs and give special attention to molecules initially thought to be mainly associated with tRNA-derived stress-induced RNAs (tiRNAs). We present an outline of their biological functions, offer a collection of useful databases, and discuss future research perspectives and applications in livestock basic and applied research. Full article

Transforming growth factor-β (TGF-β) is a crucial pathogenic mediator of inflammatory diseases. In tissue fibrosis, TGF-β regulates the pathogenic activity of infiltrated immunocytes and promotes extracellular matrix production via de novo myofibroblast generation and kidney cell activation. In cancer, TGF-β promotes cancer invasion and metastasis by enhancing the stemness and epithelial mesenchymal transition of cancer cells. However, TGF-β is highly pleiotropic in both tissue fibrosis and cancers, and thus, direct targeting of TGF-β may also block its protective anti-inflammatory and tumor-suppressive effects, resulting in undesirable outcomes. Increasing evidence suggests the involvement of long non-coding RNAs (lncRNAs) in TGF-β-driven tissue fibrosis and cancer progression with a high cell-type and disease specificity, serving as an ideal target for therapeutic development. In this review, the mechanism and translational potential of TGF-β-associated lncRNAs in tissue fibrosis and cancer will be discussed. Full article

Recent advances in gene expression analysis techniques and increased access to technologies such as microarrays, qPCR arrays, and next-generation sequencing, in the last decade, have led to increased awareness of the complexity of the inflammatory responses that lead to pathology. This finding is also the case for rheumatic diseases, importantly and specifically, rheumatoid arthritis (RA). The coincidence in major genetic and epigenetic regulatory events leading to RA’s inflammatory state is now well-recognized. Research groups have characterized the gene expression profile of early RA patients and identified a group of miRNAs that is particularly abundant in the early stages of the disease and miRNAs associated with treatment responses. In this perspective, we summarize the current state of RNA-based biomarker discovery and the context of technology adoption/implementation due to the COVID-19 pandemic. These advances have great potential for clinical application and could provide preclinical disease detection, follow-up, treatment targets, and biomarkers for treatment response monitoring. Full article

Small RNA sequencing (sRNA-Seq) approaches unveiled sequences derived from longer non-coding RNAs, such as transfer RNA (tRNA) and ribosomal RNA (rRNA) fragments, known as tRFs and rRFs, respectively. However, rRNAs and RNAs shorter than 16 nt are often depleted from library preparations/sequencing analyses, although they may be functional. Here, we sought to obtain a complete repertoire of small RNAs by sequencing the total RNA from 11 samples of 6 different eukaryotic organisms, from yeasts to human, in an extended 8- to 30-nt window of RNA length. The 8- to 15-nt window essentially contained fragments of longer non-coding RNAs, such as microRNAs, PIWI-associated RNAs (piRNAs), small nucleolar RNAs (snoRNAs), tRNAs and rRNAs. Notably, unusually short RNAs < 16 nt were more abundant than those >16 nt in bilaterian organisms. A new RT-qPCR method confirmed that two unusually short rRFs of 12 and 13 nt were more overly abundant (~3-log difference) than two microRNAs. We propose to not deplete rRNA and to reduce the lower threshold of RNA length to include unusually short RNAs in sRNA-Seq analyses and datasets, as their abundance and diversity support their potential role and importance as biomarkers of disease and/or mediators of cellular function. Full article

Exosomes carry molecules of great biological and clinical interest, such as miRNAs. The contents of exosomes vary between healthy controls and cancer patients. Therefore, miRNAs and other molecules transported in exosomes are considered a potential source of diagnostic and prognostic biomarkers in cancer. Many miRNAs have been detected in recent years. Consequently, a substantial amount of miRNA-related data comparing patients and healthy individuals is available, which contributes to a better understanding of the initiation, development, malignancy, and metastasis of cancer using non-invasive sampling procedures. However, a re-analysis of available ncRNA data is rare. This study used available data about miRNAs in exosomes comparing healthy individuals and cancer patients to identify possible global changes related to the presence of cancer. A robust transcriptomic analysis identified two common miRNAs (miR-495-3p and miR-543) deregulated in five cancer datasets. They had already been implicated in different cancers but not reported in exosomes circulating in blood. The study also examined their target genes and the implications of these genes for functional processes. Full article

Cold and nutrient-activated brown adipose tissue (BAT) is capable of increasing systemic energy expenditure via the uncoupled respiration and secretion of endocrine factors, thereby protecting mice against diet-induced obesity and improving insulin response and glucose tolerance in men. Long non-coding RNAs (lncRNAs) have recently been identified as fine-tuning regulators of cellular function. While certain lncRNAs have been functionally characterised in adipose tissue, their overall contribution in the activation of BAT remains elusive. We identified lncRNAs correlating to interscapular brown adipose tissue (iBAT) function in a high fat diet (HFD) and cold stressed mice. We focused on Gm15551, which has an adipose tissue specific expression profile, is highly upregulated during adipogenesis, and downregulated by β-adrenergic activation in mature adipocytes. Although we performed comprehensive transcriptional and adipocyte physiology profiling in vitro and in vivo, we could not detect an effect of gain or loss of function of Gm15551. Full article

Transcriptomics analysis of various small RNA (sRNA) biotypes is a new and rapidly developing field. Annotations for microRNAs, tRNAs, piRNAs and rRNAs contain information on transcript sequences and loci that is vital for downstream analyses. Several databases have been established to provide this type of data for specific RNA biotypes. However, these sources often contain data in different formats, which makes the bulk analysis of several sRNA biotypes in a single pipeline challenging. Information on some transcripts may be incomplete or conflicting with other entries. To overcome these challenges, we introduce ITAS, or Integrated Transcript Annotation for Small RNA, a filtered, corrected and integrated transcript annotation containing information on several types of small RNAs, including tRNA-derived small RNA, for several species (Homo sapiens, Rattus norvegicus, Mus musculus, Drosophila melanogaster, Caenorhabditis elegans). ITAS is presented in a format applicable for the vast majority of bioinformatic transcriptomics analysis, and it was tested in several case studies for human-derived data against existing alternative databases. Full article

In recent functional genomics studies, a large number of non-coding RNAs have been identified. It has become increasingly apparent that noncoding RNAs are crucial players in a wide range of cellular and physiological functions. They have been shown to modulate gene expression on different levels, including transcription, post-transcriptional processing, and translation. This review aims to highlight the diverse mechanisms of the regulation of gene expression by small noncoding RNAs in different conditions and different types of human cells. For this purpose, various cellular functions of microRNAs (miRNAs), circular RNAs (circRNAs), snoRNA-derived small RNAs (sdRNAs) and tRNA-derived fragments (tRFs) will be exemplified, with particular emphasis on the diversity of their occurrence and on the effects on gene expression in different stress conditions and diseased cell types. The synthesis and effect on gene expression of these noncoding RNAs varies in different cell types and may depend on environmental conditions such as different stresses. Moreover, noncoding RNAs play important roles in many diseases, including cancer, neurodegenerative disorders, and viral infections. Full article