Non-Coding RNA

Carotenoid biosynthesis in the fungus Fusarium fujikuroi is regulated by environmental factors, with light being the main stimulating signal. The CarS RING-finger protein plays an important role in the downregulation of structural genes of the carotenoid pathway. A recent transcriptomic analysis on the effect of carS mutation identified a gene for a long non-coding RNA (lncRNA) upstream of carS, called carP, the deletion of which results in increased carS mRNA levels and lack of carotenoid production. We have investigated the function of carP by studying the transcriptomic effect of its deletion and the phenotypes resulting from the reintroduction of carP to a deletion strain. The RNA-seq data showed that the loss of carP affected the mRNA levels of hundreds of genes, especially after illumination. Many of these changes appeared to be cascade effects as a result of changes in carS expression, as suggested by the comparison with differentially expressed genes in a carS mutant. Carotenoid production only recovered when carP was integrated upstream of carS, but not at other genomic locations, indicating a cis-acting mechanism on carS. However, some genes hardly affected by CarS were strongly upregulated in the carP mutant, indicating that carP may have other regulatory functions as an independent regulatory element. Full article

Triple-negative breast cancers affect thousands of women in the United States and disproportionately drive mortality from breast cancer. MicroRNAs are small, non-coding RNAs that negatively regulate gene expression post-transcriptionally by inhibiting target mRNA translation or by promoting mRNA degradation. We have identified that miRNA-203, silenced by epithelial–mesenchymal transition (EMT), is a tumor suppressor and can promote differentiation of breast cancer stem cells. In this study, we tested the ability of liposomal delivery of miR-203 to reverse aspects of breast cancer pathogenesis using breast cancer and EMT cell lines. We show that translationally relevant methods for increasing miR-203 abundance within a target tissue affects cellular properties associated with cancer progression. While stable miR-203 expression suppresses LASP1 and survivin, nanoliposomal delivery suppresses BMI1, indicating that suppression of distinct mRNA target profiles can lead to loss of cancer cell migration. Full article

The Nonsense-Mediated mRNA Decay (NMD) has been classically viewed as a translation-dependent RNA surveillance pathway degrading aberrant mRNAs containing premature stop codons. However, it is now clear that mRNA quality control represents only one face of the multiple functions of NMD. Indeed, NMD also regulates the physiological expression of normal mRNAs, and more surprisingly, of long non-coding (lnc)RNAs. Here, we review the different mechanisms of NMD activation in yeast and mammals, and we discuss the molecular bases of the NMD sensitivity of lncRNAs, considering the functional roles of NMD and of translation in the metabolism of these transcripts. In this regard, we describe several examples of functional micropeptides produced from lncRNAs. We propose that translation and NMD provide potent means to regulate the expression of lncRNAs, which might be critical for the cell to respond to environmental changes. Full article

Nuclear group I introns are restricted to the ribosomal DNA locus where they interrupt genes for small subunit and large subunit ribosomal RNAs at conserved sites in some eukaryotic microorganisms. Here, the myxomycete protists are a frequent source of nuclear group I introns due to their unique life strategy and a billion years of separate evolution. The ribosomal DNA of the myxomycete Mucilago crustacea was investigated and found to contain seven group I introns, including a direct repeat-containing intron at insertion site S1389 in the small subunit ribosomal RNA gene. We collected, analyzed, and compared 72 S1389 group IC1 introns representing diverse myxomycete taxa. The consensus secondary structure revealed a conserved ribozyme core, but with surprising sequence variations in the guanosine binding site in segment P7. Some S1389 introns harbored large extension sequences in the peripheral region of segment P9 containing direct repeat arrays. These repeats contained up to 52 copies of a putative internal guide sequence motif. Other S1389 introns harbored homing endonuclease genes in segment P1 encoding His-Cys proteins. Homing endonuclease genes were further interrupted by small spliceosomal introns that have to be removed in order to generate the open reading frames. Phylogenetic analyses of S1389 intron and host gene indicated both vertical and horizontal intron transfer during evolution, and revealed sporadic appearances of direct repeats, homing endonuclease genes, and guanosine binding site variants among the myxomycete taxa. Full article

Gene expression is an essential process for cellular growth, proliferation, and differentiation. The transcription of protein-coding genes and non-coding loci depends on RNA polymerases. Interestingly, numerous loci encode long non-coding (lnc)RNA transcripts that are transcribed by RNA polymerase II (RNAPII) and fine-tune the RNA metabolism. The nucleolus is a prime example of how different lncRNA species concomitantly regulate gene expression by facilitating the production and processing of ribosomal (r)RNA for ribosome biogenesis. Here, we summarise the current findings on how RNAPII influences nucleolar structure and function. We describe how RNAPII-dependent lncRNA can both promote nucleolar integrity and inhibit ribosomal (r)RNA synthesis by modulating the availability of rRNA synthesis factors in trans. Surprisingly, some lncRNA transcripts can directly originate from nucleolar loci and function in cis. The nucleolar intergenic spacer (IGS), for example, encodes nucleolar transcripts that counteract spurious rRNA synthesis in unperturbed cells. In response to DNA damage, RNAPII-dependent lncRNA originates directly at broken ribosomal (r)DNA loci and is processed into small ncRNA, possibly to modulate DNA repair. Thus, lncRNA-mediated regulation of nucleolar biology occurs by several modes of action and is more direct than anticipated, pointing to an intimate crosstalk of RNA metabolic events. Full article

Pervasive transcription is widespread in eukaryotes, generating large families of non-coding RNAs. Such pervasive transcription is a key player in the regulatory pathways controlling chromatin state and gene expression. Here, we describe long non-coding RNAs generated from the ribosomal RNA gene promoter called UPStream-initiating transcripts (UPS). In yeast, rDNA genes are organized in tandem repeats in at least two different chromatin states, either transcribed and largely depleted of nucleosomes (open) or assembled in regular arrays of nucleosomes (closed). The production of UPS transcripts by RNA Polymerase II from endogenous rDNA genes was initially documented in mutants defective for rRNA production by RNA polymerase I. We show here that UPS are produced in wild-type cells from closed rDNA genes but are hidden within the enormous production of rRNA. UPS levels are increased when rDNA chromatin states are modified at high temperatures or entering/leaving quiescence. We discuss their role in the regulation of rDNA chromatin states and rRNA production. Full article

Exosomes are a class of small, secreted extracellular vesicles (EV) that have recently gained considerable attention for their role in normal cellular function, disease processes and potential as biomarkers. Exosomes serve as intercellular messengers and carry molecular cargo that can alter gene expression and the phenotype of recipient cells. Here, we investigated alterations of microRNA cargo in exosomes secreted by epileptogenic tissue in tuberous sclerosis complex (TSC), a multi-system genetic disorder that includes brain lesions known as tubers. Approximately 90% of TSC patients suffer from seizures that originate from tubers, and ~60% are resistant to antiseizure drugs. It is unknown why some tubers cause seizures while others do not, and the molecular basis of drug-resistant epilepsy is not well understood. It is believed that neuroinflammation is involved, and characterization of this mechanism may be key to disrupting the “vicious cycle” between seizures, neuroinflammation, and increased seizure susceptibility. We isolated exosomes from epileptogenic and non-epileptogenic TSC tubers, and we identified differences in their microRNA cargo using small RNA-seq. We identified 12 microRNAs (including miR-142-3p, miR-223-3p and miR-21-5p) that are significantly increased in epileptogenic tubers and contain nucleic acid motifs that activate toll-like receptors (TLR7/8), initiating a neuroinflammatory cascade. Exosomes from epileptogenic tissue caused induction of key pathways in cultured cells, including innate immune signaling (TLR), inflammatory response and key signaling nodes SQSTM1 (p62) and CDKN1A (p21). Genes induced in vitro were also significantly upregulated in epileptogenic tissue. These results provide new evidence on the role of exosomes and non-coding RNA cargo in the neuroinflammatory cascade of epilepsy and may help advance the development of novel biomarkers and therapeutic approaches for the treatment of drug-resistant epilepsy. Full article

Substantial evidence indicates that microRNA-155 (miR-155) plays a crucial role in the pathogenesis of diabetes mellitus (DM) and its complications. A number of clinical studies reported low serum levels of miR-155 in patients with type 2 diabetes (T2D). Preclinical studies revealed that miR-155 partakes in the phenotypic switch of cells within the islets of Langerhans under metabolic stress. Moreover, miR-155 was shown to regulate insulin sensitivity in liver, adipose tissue, and skeletal muscle. Dysregulation of miR-155 expression was also shown to predict the development of nephropathy, neuropathy, and retinopathy in DM. Here, we systematically describe the reports investigating the role of miR-155 in DM and its complications. We also discuss the recent results from in vivo and in vitro models of type 1 diabetes (T1D) and T2D, discussing the differences between clinical and preclinical studies and shedding light on the molecular pathways mediated by miR-155 in different tissues affected by DM. Full article

Neuroendocrine neoplasms (NEN) are infrequent malignant tumors of a neuroendocrine nature that arise in various organs. They occur most frequently in the lungs, intestines, stomach and pancreas. Molecular diagnostics and prognosis of NEN development are highly relevant. The role of clinical biomarkers can be played by microRNAs (miRNAs). This work is devoted to the analysis of data on miRNA expression in NENs. For the first time, a search for specificity or a community of their functional characteristics in different types of NEN was carried out. Their properties as biomarkers were also analyzed. To date, more than 100 miRNAs have been characterized as differentially expressed and significant for the development of NEN tumors. Only about 10% of the studied miRNAs are expressed in several types of NEN; differential expression of the remaining 90% was found only in tumors of specific localizations. A significant number of miRNAs have been identified as potential biomarkers. However, only a few miRNAs have values that characterized their quality as markers. The analysis demonstrates the predominant specific expression of miRNA in each studied type of NEN. This indicates that miRNA’s functional features are predominantly influenced by the tissue in which they are formed. Full article

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), a member of the genus Betacoronavirus in the family Coronaviridae, possesses an unusually large single-stranded viral RNA (ssvRNA) genome of about ~29,811 nucleotides (nt) that causes severe and acute respiratory distress and a highly lethal viral pneumonia known as COVID-19. COVID-19 also presents with multiple ancillary systemic diseases and often involves cardiovascular, inflammatory, and/or neurological complications. Pathological viral genomes consisting of ssvRNA, like cellular messenger RNA (mRNA), are susceptible to attack, destruction, neutralization, and/or modulation by naturally occurring small non-coding RNAs (sncRNAs) within the host cell, some of which are known as microRNAs (miRNAs). This paper proposes that the actions of the 2650 known human miRNAs and other sncRNAs form the basis for an under-recognized and unappreciated innate-immune regulator of ssvRNA viral genome activities and have implications for the efficiency of SARS-CoV-2 invasion, infection, and replication. Recent research indicates that both miRNA and mRNA abundance, speciation, and complexity varies widely amongst human individuals, and this may: (i) In part explain the variability in the innate-immune immunological and pathophysiological response of different human individuals to the initiation and progression of SARS-CoV-2 infection in multiple tissue types; and (ii) further support our understanding of human biochemical and genetic individuality and the variable resistance of individuals to ssvRNA-mediated viral infection and disease. This commentary will briefly address current findings and concepts in this fascinating research area of non-coding RNA and innate-immunity with special reference to natural host miRNAs, SARS-CoV-2, and the current COVID-19 pandemic. Full article

Development is a complex process regulated both by genetic and epigenetic and environmental clues. Recently, long non-coding RNAs (lncRNAs) have emerged as key regulators of gene expression in several tissues including the brain. Altered expression of lncRNAs has been linked to several neurodegenerative, neurodevelopmental and mental disorders. The identification and characterization of lncRNAs that are deregulated or mutated in neurodevelopmental and mental health diseases are fundamental to understanding the complex transcriptional processes in brain function. Crucially, lncRNAs can be exploited as a novel target for treating neurological disorders. In our review, we first summarize the recent advances in our understanding of lncRNA functions in the context of cell biology and then discussing their association with selected neuronal development and neurological disorders. Full article

MicroRNAs (miRNAs) are endogenous non-coding RNAs ~22 nucleotides in length, which have been shown to participate in various biological processes. As one of the most researched miRNAs, the miR-200 family has been found to regulate several factors that are associated with the epithelial to mesenchymal transition (EMT) and cancer stem cells (CSCs) behavior. In this review, we briefly summarize the background of the miR-200 family and their implication in various dental diseases. We focus on the expression changes, biological functions, and clinical significance of the miR-200 family in oral cancer; periodontitis; oral potentially malignant disorder; gingival overgrowth; and other periodontal diseases. Additionally, we discuss the use of the miR-200 family as molecular biomarkers for diagnosis, prognostic, and therapeutic application. Full article

Long non-coding RNAs (lncRNAs) contribute to cell fate decisions by modulating genome expression and stability. In the fission yeast Schizosaccharomyces pombe, the transition from mitosis to meiosis results in a marked remodeling of gene expression profiles, which ultimately ensures gamete production and inheritance of genetic information to the offspring. This key developmental process involves a set of dedicated lncRNAs that shape cell cycle-dependent transcriptomes through a variety of mechanisms, including epigenetic modifications and the modulation of transcription, post-transcriptional and post-translational regulations, and that contribute to meiosis-specific chromosomal events. In this review, we summarize the biology of these lncRNAs, from their identification to mechanism of action, and discuss their regulatory role in the control of gametogenesis. Full article

Phenotypes are driven by regulated gene expression, which in turn are mediated by complex interactions between diverse biological molecules. Protein–DNA interactions such as histone and transcription factor binding are well studied, along with RNA–RNA interactions in short RNA silencing of genes. In contrast, lncRNA-protein interaction (LPI) mechanisms are comparatively unknown, likely directed by the difficulties in studying LPI. However, LPI are emerging as key interactions in epigenetic mechanisms, playing a role in development and disease. Their importance is further highlighted by their conservation across kingdoms. Hence, interest in LPI research is increasing. We therefore review the current state of the art in lncRNA-protein interactions. We specifically surveyed recent computational methods and databases which researchers can exploit for LPI investigation. We discovered that algorithm development is heavily reliant on a few generic databases containing curated LPI information. Additionally, these databases house information at gene-level as opposed to transcript-level annotations. We show that early methods predict LPI using molecular docking, have limited scope and are slow, creating a data processing bottleneck. Recently, machine learning has become the strategy of choice in LPI prediction, likely due to the rapid growth in machine learning infrastructure and expertise. While many of these methods have notable limitations, machine learning is expected to be the basis of modern LPI prediction algorithms. Full article

Dictyostelium discoideum is a social amoeba, which on starvation develops from a single-cell state to a multicellular fruiting body. This developmental process is accompanied by massive changes in gene expression, which also affect non-coding RNAs. Here, we investigate how tRNAs as key regulators of the translation process are affected by this transition. To this end, we used LOTTE-seq to sequence the tRNA pool of D. discoideum at different developmental time points and analyzed both tRNA composition and tRNA modification patterns. We developed a workflow for the specific detection of modifications from reverse transcriptase signatures in chemically untreated RNA-seq data at single-nucleotide resolution. It avoids the comparison of treated and untreated RNA-seq data using reverse transcription arrest patterns at nucleotides in the neighborhood of a putative modification site as internal control. We find that nucleotide modification sites in D. discoideum tRNAs largely conform to the modification patterns observed throughout the eukaroytes. However, there are also previously undescribed modification sites. We observe substantial dynamic changes of both expression levels and modification patterns of certain tRNA types during fruiting body development. Beyond the specific application to D. discoideum our results demonstrate that the developmental variability of tRNA expression and modification can be traced efficiently with LOTTE-seq. Full article

It is now commonly accepted that most of the mammalian genome is transcribed as RNA, yet less than 2% of such RNA encode for proteins. A majority of transcribed RNA exists as non-protein-coding RNAs (ncRNAs) with various functions. Because of the lack of sequence homologies among most ncRNAs species, it is difficult to infer the potential functions of ncRNAs by examining sequence patterns, such as catalytic domains, as in the case of proteins. Added to the existing complexity of predicting the functions of the ever-growing number of ncRNAs, increasing evidence suggests that various enzymes modify ncRNAs (e.g., ADARs, METTL3, and METTL14), which has opened up a new field of study called epitranscriptomics. Here, we examine the current status of ncRNA research from the perspective of epitranscriptomics. Full article

Box C/D small nucleolar RNAs (C/D snoRNAs) represent an ancient family of small non-coding RNAs that are classically viewed as housekeeping guides for the 2′-O-methylation of ribosomal RNA in Archaea and Eukaryotes. However, an extensive set of studies now argues that they are involved in mechanisms that go well beyond this function. Here, we present these pieces of evidence in light of the current comprehension of the molecular mechanisms that control C/D snoRNA expression and function. From this inventory emerges that an accurate description of these activities at a molecular level is required to let the snoRNA field enter in a second age of maturity. Full article

RNA silencing pathways control eukaryotic gene expression transcriptionally or posttranscriptionally in a sequence-specific manner. In RNA silencing, the production of double-stranded RNA (dsRNA) gives rise to various classes of 20–24 nucleotide (nt) small RNAs (smRNAs). In Arabidopsis thaliana, smRNAs are often derived from long dsRNA molecules synthesized by one of the six genomically encoded RNA-dependent RNA Polymerase (RDR) proteins. However, the full complement of the RDR-dependent smRNAs and functions that these proteins and their RNA-binding cofactors play in plant RNA silencing has not been fully uncovered. To address this gap, we performed a global genomic analysis of all six RDRs and two of their cofactors to find new substrates for RDRs and targets of the resulting RDR-derived siRNAs to uncover new functions for these proteins in plants. Based on these analyses, we identified substrates for the three RDRγ clade proteins (RDR3–5), which had not been well-characterized previously. We also identified new substrates for the other three RDRs (RDR1, RDR2, and RDR6) as well as the RDR2 cofactor RNA-directed DNA methylation 12 (RDM12) and the RDR6 cofactor suppressor of gene silencing 3 (SGS3). These findings revealed that the target substrates of SGS3 are not limited to those solely utilized by RDR6, but that this protein seems to be a more general cofactor for the RDR family of proteins. Additionally, we found that RDR6 and SGS3 are involved in the production of smRNAs that target transcripts related to abiotic stresses, including water deprivation, salt stress, and ABA response, and as expected the levels of these mRNAs are increased in rdr6 and sgs3 mutant plants. Correspondingly, plants that lack these proteins (rdr6 and sgs3 mutants) are hypersensitive to ABA treatment, tolerant to high levels of PEG8000, and have a higher survival rate under salt treatment in comparison to wild-type plants. In total, our analyses have provided an extremely data-rich resource for uncovering new functions of RDR-dependent RNA silencing in plants, while also revealing a previously unexplored link between the RDR6/SGS3-dependent pathway and plant abiotic stress responses. Full article