Structural basis for Tpt1-catalyzed 2'-PO4 transfer from RNA and NADP(H) to NAD.

Tpt1 is an essential agent of fungal and plant tRNA splicing that removes an internal RNA 2'-phosphate generated by tRNA ligase. Tpt1 also removes the 2'-phosphouridine mark installed by Ark1 kinase in the V-loop of archaeal tRNAs. Tpt1 performs a two-step reaction in which the 2'-PO4 attacks NAD+ to form an RNA-2'-phospho-(ADP-ribose) intermediate, and transesterification of the ADP-ribose O2″ to the RNA 2'-phosphodiester yields 2'-OH RNA and ADP-ribose-1″,2″-cyclic phosphate. Here, we present structures of archaeal Tpt1 enzymes, captured as product complexes with ADP-ribose-1″-PO4, ADP-ribose-2″-PO4, and 2'-OH RNA, and as substrate complexes with 2',5'-ADP and NAD+, that illuminate 2'-PO4 junction recognition and catalysis. We show that archaeal Tpt1 enzymes can use the 2'-PO4-containing metabolites NADP+ and NADPH as substrates for 2'-PO4 transfer to NAD+. A role in 2'-phospho-NADP(H) dynamics provides a rationale for the prevalence of Tpt1 in taxa that lack a capacity for internal RNA 2'-phosphorylation.

Machine learning for RNA 2D structure prediction benchmarked on experimental data.

Since the 1980s, dozens of computational methods have addressed the problem of predicting RNA secondary structure. Among them are those that follow standard optimization approaches and, more recently, machine learning (ML) algorithms. The former were repeatedly benchmarked on various datasets. The latter, on the other hand, have not yet undergone extensive analysis that could suggest to the user which algorithm best fits the problem to be solved. In this review, we compare 15 methods that predict the secondary structure of RNA, of which 6 are based on deep learning (DL), 3 on shallow learning (SL) and 6 control methods on non-ML approaches. We discuss the ML strategies implemented and perform three experiments in which we evaluate the prediction of (I) representatives of the RNA equivalence classes, (II) selected Rfam sequences and (III) RNAs from new Rfam families. We show that DL-based algorithms (such as SPOT-RNA and UFold) can outperform SL and traditional methods if the data distribution is similar in the training and testing set. However, when predicting 2D structures for new RNA families, the advantage of DL is no longer clear, and its performance is inferior or equal to that of SL and non-ML methods.

preMLI: a pre-trained method to uncover microRNA-lncRNA potential interactions.

The interaction between microribonucleic acid and long non-coding ribonucleic acid plays a very important role in biological processes, and the prediction of the one is of great significance to the study of its mechanism of action. Due to the limitations of traditional biological experiment methods, more and more computational methods are applied to this field. However, the existing methods often have problems, such as inadequate acquisition of potential features of the sequence due to simple coding and the need to manually extract features as input. We propose a deep learning model, preMLI, based on rna2vec pre-training and deep feature mining mechanism. We use rna2vec to train the ribonucleic acid (RNA) dataset and to obtain the RNA word vector representation and then mine the RNA sequence features separately and finally concatenate the two feature vectors as the input of the prediction task. The preMLI performs better than existing methods on benchmark datasets and has cross-species prediction capabilities. Experiments show that both pre-training and deep feature mining mechanisms have a positive impact on the prediction performance of the model. To be more specific, pre-training can provide more accurate word vector representations. The deep feature mining mechanism also improves the prediction performance of the model. Meanwhile, The preMLI only needs RNA sequence as the input of the model and has better cross-species prediction performance than the most advanced prediction models, which have reference value for related research.

Insights into the structure and function of the RNA ligase RtcB.

To be functional, some RNAs require a processing step involving splicing events. Each splicing event necessitates an RNA ligation step. RNA ligation is a process that can be achieved with various intermediaries such as self-catalysing RNAs, 5'-3' and 3'-5' RNA ligases. While several types of RNA ligation mechanisms occur in human, RtcB is the only 3'-5' RNA ligase identified in human cells to date. RtcB RNA ligation activity is well known to be essential for the splicing of XBP1, an essential transcription factor of the unfolded protein response; as well as for the maturation of specific intron-containing tRNAs. As such, RtcB is a core factor in protein synthesis and homeostasis. Taking advantage of the high homology between RtcB orthologues in archaea, bacteria and eukaryotes, this review will provide an introduction to the structure of RtcB and the mechanism of 3'-5' RNA ligation. This analysis is followed by a description of the mechanisms regulating RtcB activity and localisation, its known partners and its various functions from bacteria to human with a specific focus on human cancer.

The Potency of Cowpea Mosaic Virus Particles for Cancer In Situ Vaccination Is Unaffected by the Specific Encapsidated Viral RNA.

Plant virus nanoparticles can be used as drug carriers, imaging reagents, vaccine carriers, and immune adjuvants in the formulation of intratumoral in situ cancer vaccines. One example is the cowpea mosaic virus (CPMV), a nonenveloped virus with a bipartite positive-strand RNA genome with each RNA packaged separately into identical protein capsids. Based on differences in their densities, the components carrying RNA-1 (6 kb) denoted as the bottom (B) component or carrying RNA-2 (3.5 kb) denoted as the middle (M) component can be separated from each other and from a top (T) component, which is devoid of any RNA. Previous preclinical mouse studies and canine cancer trials used mixed populations of CPMV (containing B, M, and T components), so it is unclear whether the particle types differ in their efficacies. It is known that the CPMV RNA genome contributes to immunostimulation by activation of TLR7. To determine whether the two RNA genomes that have different sizes and unrelated sequences cause different immune stimulation, we compared the therapeutic efficacies of B and M components and unfractionated CPMV in vitro and in mouse cancer models. We found that separated B and M particles behaved similarly to the mixed CPMV, activating innate immune cells to induce the secretion of pro-inflammatory cytokines such as IFNα, IFNγ, IL-6, and IL-12, while inhibiting immunosuppressive cytokines such as TGF-ß and IL-10. In murine models of melanoma and colon cancer, the mixed and separated CPMV particles all significantly reduced tumor growth and prolonged survival with no significant difference. This shows that the specific RNA genomes similarly stimulate the immune system even though B particles have 40% more RNA than M particles; each CPMV particle type can be used as an effective adjuvant against cancer with the same efficacy as native mixed CPMV. From a translational point of view, the use of either B or M component vs the mixed CPMV formulation offers the advantage that separated B or M alone is noninfectious toward plants and thus provides agronomic safety.

Substrate analogs that trap the 2'-phospho-ADP-ribosylated RNA intermediate of the Tpt1 (tRNA 2'-phosphotransferase) reaction pathway.

The enzyme Tpt1 removes an internal RNA 2'-PO4 via a two-step reaction in which: (i) the 2'-PO4 attacks NAD+ to form an RNA-2'-phospho-(ADP-ribose) intermediate and nicotinamide; and (ii) transesterification of the ADP-ribose O2″ to the RNA 2'-phosphodiester yields 2'-OH RNA and ADP-ribose-1″,2″-cyclic phosphate. Because step 2 is much faster than step 1, the ADP-ribosylated RNA intermediate is virtually undetectable under normal circumstances. Here, by testing chemically modified nucleic acid substrates for activity with bacterial Tpt1 enzymes, we find that replacement of the ribose-2'-PO4 nucleotide with arabinose-2'-PO4 selectively slows step 2 of the reaction pathway and results in the transient accumulation of high levels of the reaction intermediate. We report that replacing the NMN ribose of NAD+ with 2'-fluoroarabinose (thereby eliminating the ribose O2″ nucleophile) results in durable trapping of RNA-2'-phospho-(ADP-fluoroarabinose) as a "dead-end" product of step 1. Tpt1 enzymes from diverse taxa differ in their capacity to use ara-2″F-NAD+ as a substrate.

RNAlign2D: a rapid method for combined RNA structure and sequence-based alignment using a pseudo-amino acid substitution matrix.

BACKGROUND: The functions of RNA molecules are mainly determined by their secondary structures. These functions can also be predicted using bioinformatic tools that enable the alignment of multiple RNAs to determine functional domains and/or classify RNA molecules into RNA families. However, the existing multiple RNA alignment tools, which use structural information, are slow in aligning long molecules and/or a large number of molecules. Therefore, a more rapid tool for multiple RNA alignment may improve the classification of known RNAs and help to reveal the functions of newly discovered RNAs. RESULTS: Here, we introduce an extremely fast Python-based tool called RNAlign2D. It converts RNA sequences to pseudo-amino acid sequences, which incorporate structural information, and uses a customizable scoring matrix to align these RNA molecules via the multiple protein sequence alignment tool MUSCLE. CONCLUSIONS: RNAlign2D produces accurate RNA alignments in a very short time. The pseudo-amino acid substitution matrix approach utilized in RNAlign2D is applicable for virtually all protein aligners.

NAD+-dependent RNA terminal 2' and 3' phosphomonoesterase activity of a subset of Tpt1 enzymes.

The enzyme Tpt1 removes the 2'-PO4 at the splice junction generated by fungal tRNA ligase; it does so via a two-step reaction in which (i) the internal RNA 2'-PO4 attacks NAD+ to form an RNA-2'-phospho-ADP-ribosyl intermediate; and (ii) transesterification of the ribose O2″ to the 2'-phosphodiester yields 2'-OH RNA and ADP-ribose-1″,2″-cyclic phosphate products. The role that Tpt1 enzymes play in taxa that have no fungal-type RNA ligase remains obscure. An attractive prospect is that Tpt1 enzymes might catalyze reactions other than internal RNA 2'-PO4 removal, via their unique NAD+-dependent transferase mechanism. This study extends the repertoire of the Tpt1 enzyme family to include the NAD+-dependent conversion of RNA terminal 2' and 3' monophosphate ends to 2'-OH and 3'-OH ends, respectively. The salient finding is that different Tpt1 enzymes vary in their capacity and positional specificity for terminal phosphate removal. Clostridium thermocellum and Aeropyrum pernix Tpt1 proteins are active on 2'-PO4 and 3'-PO4 ends, with a 2.4- to 2.6-fold kinetic preference for the 2'-PO4 The accumulation of a terminal 3'-phospho-ADP-ribosylated RNA intermediate during the 3'-phosphotransferase reaction suggests that the geometry of the 3'-p-ADPR adduct is not optimal for the ensuing transesterification step. Chaetomium thermophilum Tpt1 acts specifically on a terminal 2'-PO4 end and not with a 3'-PO4 In contrast, Runella slithyformis Tpt1 and human Tpt1 are ineffective in removing either a 2'-PO4 or 3'-PO4 end.

The role of OncoSnoRNAs and Ribosomal RNA 2'-O-methylation in Cancer.

Ribosomes are essential nanomachines responsible for all protein production in cells. Ribosome biogenesis and function are energy costly processes, they are tightly regulated to match cellular needs. In cancer, major pathways that control ribosome biogenesis and function are often deregulated to ensure cell survival and to accommodate the continuous proliferation of tumour cells. Ribosomal RNAs (rRNAs) are abundantly modified with 2'-O-methylation (Nm, ribomethylation) being one of the most common modifications. In eukaryotic ribosomes, ribomethylation is performed by the methyltransferase Fibrillarin guided by box C/D small nucleolar RNAs (snoRNAs). Accumulating evidences indicate that snoRNA expression and ribosome methylation profiles are altered in cancer. Here we review our current knowledge on differential snoRNA expression and rRNA 2'-O methylation in the context of human malignancies, and discuss the consequences and opportunities for cancer diagnostics, prognostics, and therapeutics.

RNA Editing: A New Therapeutic Target in Amyotrophic Lateral Sclerosis and Other Neurological Diseases.

The conversion of adenosine to inosine in RNA editing (A-to-I RNA editing) is recognized as a critical post-transcriptional modification of RNA by adenosine deaminases acting on RNAs (ADARs). A-to-I RNA editing occurs predominantly in mammalian and human central nervous systems and can alter the function of translated proteins, including neurotransmitter receptors and ion channels; therefore, the role of dysregulated RNA editing in the pathogenesis of neurological diseases has been speculated. Specifically, the failure of A-to-I RNA editing at the glutamine/arginine (Q/R) site of the GluA2 subunit causes excessive permeability of α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptors to Ca2+, inducing fatal status epilepticus and the neurodegeneration of motor neurons in mice. Therefore, an RNA editing deficiency at the Q/R site in GluA2 due to the downregulation of ADAR2 in the motor neurons of sporadic amyotrophic lateral sclerosis (ALS) patients suggests that Ca2+-permeable AMPA receptors and the dysregulation of RNA editing are suitable therapeutic targets for ALS. Gene therapy has recently emerged as a new therapeutic opportunity for many heretofore incurable diseases, and RNA editing dysregulation can be a target for gene therapy; therefore, we reviewed neurological diseases associated with dysregulated RNA editing and a new therapeutic approach targeting dysregulated RNA editing, especially one that is effective in ALS.

Long noncoding RNA HOXA-AS2 mediates microRNA-106b-5p to repress sepsis-engendered acute kidney injury.

HOXA cluster antisense RNA 2 (HOXA-AS2) is a long noncoding RNA associated with the development of numerous cancers. But, whether HOXA-AS2 exhibits a certain function in sepsis-engendered acute kidney injury (AKI) remains uninvestigated. We strived to unveil the role of HOXA-AS2 in sepsis-engendered AKI. The expression of HOXA-AS2 in sepsis patients, animal models and lipopolysaccharide (LPS)-impaired HK-2 cells was primarily assessed via a real-time quantitative polymerase chain reaction. The effects of HOXA-AS2 on cell survival of HK-2 cells under LPS irritation were evaluated after overexpression of HOXA-AS2. The correlation between HOXA-AS2 and microRNA (miR)-106b-5p was forecasted via bioinformatics software and verified by using a luciferase report system. Subsequently, the functions of miR-106b-5p in LPS-damaged HK-2 cells were reassessed. Western blot was used for the determination of Wnt/ß-catenin and nuclear factor-κB (NF-κB) pathways. HOXA-AS2 expression was decreased in sepsis patients, animal operation group and LPS-irritated HK-2 cells. Overexpressed HOXA-AS2 mollified LPS-triggered impairment in HK-2 cells. In addition, a negative mediatory relation between HOXA-AS2 and miR-106b-5p was predicated. Synchronously, overexpressed miR-106b-5p counteracted the protection of HOXA-AS2 in LPS-damaged HK-2 cells. Ultimately, Wnt/ß-catenin and NF-κB pathways were hindered by HOXA-AS2 via targeting miR-106b-5p. HOXA-AS2 exhibited protection in sepsis-engendered AKI via targeting miR-106b-5p and hindering the Wnt/ß-catenin and NF-κB pathways.

Good or not good: Role of miR-18a in cancer biology.

miR-18a is a member of primary transcript called miR-17-92a (C13orf25 or MIR17HG) which also contains five other miRNAs: miR-17, miR-19a, miR-20a, miR-19b and miR-92a. This cluster as a whole shows specific characteristics, where miR-18a seems to be unique. In contrast to the other members, the expression of miR-18a is additionally controlled and probably functions as its own internal controller of the cluster. miR-18a regulates many genes involved in proliferation, cell cycle, apoptosis, response to different kinds of stress, autophagy and differentiation. The disturbances of miR-18a expression are observed in cancer as well as in different diseases or pathological states. The miR-17-92a cluster is commonly described as oncogenic and it is known as 'oncomiR-1', but this statement is a simplification because miR-18a can act both as an oncogene and a suppressor. In this review we summarize the current knowledge about miR-18a focusing on its regulation, role in cancer biology and utility as a potential biomarker.

The long noncoding RNA HIF1A-AS2 facilitates cisplatin resistance in bladder cancer.

Chemotherapy drug resistance frequently happens in more than 50% of bladder cancer patients and is the major obstacle for the bladder cancer therapy. Recent studies have shown that long noncoding RNA (lncRNA) is involved in the development of chemoresistance. In this study, we reported hypoxia inducible factor 1α-antisense RNA 2 (HIF1A-AS2), as a subtype-specific hypoxia inducible lncRNA, is upregulated in bladder cancer cells and tissue after cisplatin (Cis) treatment. The induction of HIF1A-AS2 in bladder cancer cells rendered resistance to Cis-induced apoptosis. Silencing HIF1A-AS2 in Cis-resistant bladder cancer cells was re-sensitized to Cis-induced apoptosis. Mechanically, we found that HIF1A-AS2 suppressed the transcription activity of p53 family proteins by promoting the expression of high-mobility group A1 (HMGA1). The induction of HMGA1 physically interacts with p53, p63, and p73, and therefore constrains their transcriptional activity on Bax. Knockdown of HIF1A-AS2 or HMGA1 rescued the expression of Bax, which therefore enhanced the killing effect of Cis. Furthermore, we also found that the expression of HIF1A-AS2 was higher in the human bladder tumor tissues after Cis treatment, and was positive correlated to the expression of HIF1α and HMGA1. This study suggests that upregulated HIF1A-AS2 hampers the p53 family proteins dependent apoptotic pathway to promote Cis resistance in bladder cancer. Our data suggested that HIF1A-AS2 plays oncogenic roles and can be used as a therapeutic target for treating human bladder cancer.

CERNA2: A predictor for clinical progression and poor prognosis in cervical carcinoma.

Competing long noncoding RNA 2 (lncRNA 2) for microRNA let-7b (CERNA2) has emerged as an important regulator of tumorigenesis and cancer progression but the clinical value and regulatory function of CERNA2 is yet to be investigated in cervical carcinoma. In our study, we found the CERNA2 expression was obviously increased in cervical carcinoma tissues compared with adjacent normal cervical tissues. In addition, we observed that metastatic lymph nodes exhibited high levels of CERNA2 expression in contrast to primary cervical carcinoma tissues. Furthermore, high CERNA2 expression was associated with advanced clinical stage, lymph node metastasis, distant metastasis poor histological grade, and short overall survival in cervical carcinoma patients. Moreover, high CERNA2 expression acted as an independent unfavorable predictor for overall survival in cervical carcinoma patients. The cell migration and invasion assays in vitro suggested that knockdown of CERNA2 remarkably inhibited cell migration and invasion in cervical carcinoma. In conclusion, CERNA2 functions as an oncogenic lncRNA and may be as a potential therapeutic target in cervical carcinoma.

Clinical significance of long noncoding RNA VIM-AS1 and CTBP1-AS2 expression in type 2 diabetes.

BACKGROUND/AIMS: The risk of type 2 diabetes (T2D) is determined by a combination of genetic and environmental factors. Multiple studies have proposed that long noncoding RNAs (lncRNAs) are crucial molecules in regulating several biological processes and complex diseases. The study was aimed at investigating the association between the expression levels of lncRNA VIM-AS1, lncRNA CTBP1-AS2, and T2D susceptibility. METHODS: lncRNA VIM-AS1 and lncRNA CTBP1-AS2 in the peripheral blood mononuclear cell (PBMC) of 100 healthy individuals and 100 T2D patients were collected for Quantitative Real-Time RT-PCR analysis. A logistic regression was performed to understand whether the likelihood of T2D can be predicted based on the expression levels of lncRNA VIM-AS1 and lncRNA CTBP1-AS2. Receiver operating characteristic (ROC) analysis was also performed to determine the statistical analysis of VIM-AS1 and CTBP1-AS2 levels in 200 samples. RESULTS: Our results display that decreased levels of VIM-AS1 and CTBP1-AS2 in PBMC were associated with diabetes in Iranian population. The logistic regression revealed that Systolic blood pressure (SBP), low-density lipoprotein cholesterol (LDL-C), Fasting blood glucose (FBG) and CTBP1-AS2 are substantial predictors of T2D. The ROC analysis of CTBP1-AS2 revealed the area under the ROC curve (AUC) of 0.68 with a sensitivity of 58.7% and specificity of 75.3% in distinguishing nondiabetic from diabetic subjects. The ROC analysis of VIM-AS1 determined AUC of 0.63 with a sensitivity of 56.1% and specificity of 68.37% in distinguishing the two diagnostic groups. CONCLUSION: lncRNA VIM-AS1 and lncRNA CTBP1-AS2 expression levels are associated with T2D susceptibility.

HIF1A-AS2 predicts poor prognosis and regulates cell migration and invasion in triple-negative breast cancer.

The aberrant expression of hypoxia-inducible factor 1 alpha (HIF1A)-antisense RNA 2 (HIF1A-AS2) was found in various human cancers including breast cancer. The aim of this study was to present more evidence about the role HIF1A-AS2 on triple-negative breast cancer (TNBC). In our results, HIF1A-AS2 was also found to be upregulated in TNBC tissues compared with non-TNBC tissues or adjacent normal tissues. Besides, HIF1A-AS2 expression was also elevated in TNBC cell lines compared with the normal breast epithelial cell line. Moreover, high expression of HIF1A-AS2 was associated with lymph node metastasis, distant metastasis and unfavorable histological grade in TNBC patients. Survival analysis showed a TNBC patient with high HIF1A-AS2 expression had shorter overall survival than patients with low HIF1A-AS2 expression, and HIF1A-AS2 high expression acted as an independent poor prognostic factor for overall survival in TNBC patients. The cell migration and invasion assays suggested inhibition of HIF1A-AS2 obviously depressed TNBC cell migration and invasion. In conclusion, HIF1A-AS2 serves as a novel biomarker for predicting clinical progression and prognosis in TNBC.

Archaeal fibrillarin-Nop5 heterodimer 2'-O-methylates RNA independently of the C/D guide RNP particle.

Archaeal fibrillarin (aFib) is a well-characterized S-adenosyl methionine (SAM)-dependent RNA 2'-O-methyltransferase that is known to act in a large C/D ribonucleoprotein (RNP) complex together with Nop5 and L7Ae proteins and a box C/D guide RNA. In the reaction, the guide RNA serves to direct the methylation reaction to a specific site in tRNA or rRNA by sequence complementarity. Here we show that a Pyrococcus abyssi aFib-Nop5 heterodimer can alone perform SAM-dependent 2'-O-methylation of 16S and 23S ribosomal RNAs in vitro independently of L7Ae and C/D guide RNAs. Using tritium-labeling, mass spectrometry, and reverse transcription analysis, we identified three in vitro 2'-O-methylated positions in the 16S rRNA of P. abyssi, positions lying outside of previously reported pyrococcal C/D RNP methylation sites. This newly discovered stand-alone activity of aFib-Nop5 may provide an example of an ancestral activity retained in enzymes that were recruited to larger complexes during evolution.

KCNQ1OT1, HIF1A-AS2 and APOA1-AS are promising novel biomarkers for diagnosis of coronary artery disease.

This study aimed to evaluate the potential of long noncoding RNAs (lncRNAs) as biomarkers for coronary artery disease (CAD). We measured the levels of three atherosclerosis- or cardiac-related lncRNAs in peripheral blood monocyte cells (PBMCs) from 20 CAD patients and 20 non-CAD control participants using real-time reverse transcription-polymerase chain reaction (real-time RT-PCR) methods. We found that the levels of lncRNA KCNQ1 opposite strand/antisense transcript 1 (KCNQ1OT1), hypoxia-inducible factor 1 alpha-antisense RNA 2 (HIF1A-AS2) and apolipoprotein A-1 antisense RNA (APOA1-AS) were significantly increased in CAD patients (KCNQ1OT1 increased by 2.38-fold, P = 0.00042; HIF1A-AS2 increased by 2.00-fold, P = 0.0001; APOA1-AS increased by 4.52-fold, P = 0.000048). The area under the ROC curve was 0.865 for KCNQ1OT1, 0.852 for HIF1A-AS2, and 0.967 for APOA1-AS. Furthermore, the combination of lncRNAs resulted in a much higher AUC value of 0.990 for the prediction of CAD. Spearman's correlation analysis showed that APOA1-AS was positively correlated with NT-proBNP, CKMB, MYO and HsTnT, whereas HIF1A-AS2 was correlated with NT-proBNP and HsTnT. Hence, the elevation of KCNQ1OT1, HIF1A-AS2 and APOA1-AS predicts CAD and these molecules may be considered as novel biomarkers of CAD.

Extracellular RNAs as Biomarkers of Sporadic Amyotrophic Lateral Sclerosis and Other Neurodegenerative Diseases.

Recent progress in the research for underlying mechanisms in neurodegenerative diseases, including Alzheimer disease (AD), Parkinson disease (PD), and amyotrophic lateral sclerosis (ALS) has led to the development of potentially effective treatment, and hence increased the need for useful biomarkers that may enable early diagnosis and therapeutic monitoring. The deposition of abnormal proteins is a pathological hallmark of neurodegenerative diseases, including ß-amyloid in AD, α-synuclein in PD, and the transactive response DNA/RNA binding protein of 43kDa (TDP-43) in ALS. Furthermore, progression of the disease process accompanies the spreading of abnormal proteins. Extracellular proteins and RNAs, including mRNA, micro RNA, and circular RNA, which are present as a composite of exosomes or other forms, play a role in cell-cell communication, and the role of extracellular molecules in the cell-to-cell spreading of pathological processes in neurodegenerative diseases is now in the spotlight. Therefore, extracellular proteins and RNAs are considered potential biomarkers of neurodegenerative diseases, in particular ALS, in which RNA dysregulation has been shown to be involved in the pathogenesis. Here, we review extracellular proteins and RNAs that have been scrutinized as potential biomarkers of neurodegenerative diseases, and discuss the possibility of extracellular RNAs as diagnostic and therapeutic monitoring biomarkers of sporadic ALS.

Modification of betanodavirus virulence by substitutions in the 3' terminal region of RNA2.

Betanodaviruses have bi-segmented positive-sense RNA genomes, consisting of RNAs 1 and 2. For some members of the related genus alphanodavirus, the 3' terminal 50 nucleotides (nt) of RNA2, including a predicted stem-loop structure (3'SL), are essential for replication. We investigate the possible existence and role of a similar structure in a reassortant betanodavirus strain (RGNNV/SJNNV). In this study, we developed three recombinant strains containing nucleotide changes at positions 1408 and 1412. Predictive models showed stem-loop structures involving nt 1398-1421 of the natural reassortant whereas this structure is modified in the recombinant viruses harbouring point mutations r1408 and r1408-1412, but not in r1412. Results obtained from infectivity assays showed differences between the reference strains and the mutants in both RNA1 and RNA2 synthesis. Moreover, an imbalance between the synthesis of both segments was demonstrated, mainly with the double mutant. All these results suggest an interaction between RNA1 and the 3' non-coding regions (3'NCR) of RNA2. In addition, the significant attenuation of the virulence for Senegalese sole and the delayed replication of r1408-1412 in brain tissues may point to an interaction of RNA2 with host cellular proteins.