miRNA Biogenesis Research Articles

MiRNA-Based Technologies in Cancer Therapy.

The discovery of therapeutic miRNAs is one of the most exciting challenges for pharmaceutical companies. Since the first miRNA was discovered in 1993, our knowledge of miRNA biology has grown considerably. Many studies have demonstrated that miRNA expression is dysregulated in many diseases, making them appealing tools for novel therapeutic approaches. This review aims to discuss miRNA biogenesis and function, as well as highlight strategies for delivering miRNA agents, presenting viral, non-viral, and exosomic delivery as therapeutic approaches for different cancer types. We also consider the therapeutic role of microRNA-mediated drug repurposing in cancer therapy.

4-Hydroxynonenal impairs miRNA maturation in heart failure via Dicer post-translational modification.

Developing novel therapies to battle the global public health burden of heart failure remains challenging. This study investigates the underlying mechanisms and potential treatment for 4-hydroxynonenal (4-HNE) deleterious effects in heart failure. Biochemical, functional, and histochemical measurements were applied to identify 4-HNE adducts in rat and human failing hearts. In vitro studies were performed to validate 4-HNE targets. 4-HNE, a reactive aldehyde by-product of mitochondrial dysfunction in heart failure, covalently inhibits Dicer, an RNase III endonuclease essential for microRNA (miRNA) biogenesis. 4-HNE inhibition of Dicer impairs miRNA processing. Mechanistically, 4-HNE binds to recombinant human Dicer through an intermolecular interaction that disrupts both activity and stability of Dicer in a concentration- and time-dependent manner. Dithiothreitol neutralization of 4-HNE or replacing 4-HNE-targeted residues in Dicer prevents 4-HNE inhibition of Dicer in vitro. Interestingly, end-stage human failing hearts from three different heart failure aetiologies display defective 4-HNE clearance, decreased Dicer activity, and miRNA biogenesis impairment. Notably, boosting 4-HNE clearance through pharmacological re-activation of mitochondrial aldehyde dehydrogenase 2 (ALDH2) using Alda-1 or its improved orally bioavailable derivative AD-9308 restores Dicer activity. ALDH2 is a major enzyme responsible for 4-HNE removal. Importantly, this response is accompanied by improved miRNA maturation and cardiac function/remodelling in a pre-clinical model of heart failure. 4-HNE inhibition of Dicer directly impairs miRNA biogenesis in heart failure. Strikingly, decreasing cardiac 4-HNE levels through pharmacological ALDH2 activation is sufficient to re-establish Dicer activity and miRNA biogenesis; thereby representing potential treatment for patients with heart failure.

Role of miR-276-3p in the cyantraniliprole resistance mechanism of Bemisia tabaci via CYP6CX3 targeting

The sweet potato whitefly, Bemisia tabaci, is an important insect pest that transmits over 200 different plant viruses and causes serious damage to the production of cotton and Solanaceae vegetables. Cyantraniliprole is the first diamide insecticide, showing toxicity against B. tabaci. However, B. tabaci has developed resistance to this insecticide by upregulating the expressions of cytochrome P450 genes such as CYP6CX3, while there is limited information on the regulatory mechanism mediated by miRNA. In the present study, ten miRNAs were predicted to target CYP6CX3, in which miR-276-3p showed an inverse expression pattern with CYP6CX3 in two cyantraniliprole resistant strains and under cyantraniliprole exposure. A luciferase assay demonstrated that miR-276-3p suppressed CYP6CX3 expression by pairing with residues 1445–1453. Overexpression or knockdown of miR-276-3p directly impacted B. tabaci resistance to cyantraniliprole. In addition, exposure to cyantraniliprole led to a significant reduction in the expressions of five genes (drosha, dicer1, dicer2, Ago1, and Ago2A) associated with miRNA biogenesis. Suppressing genes such as drosha, dicer1, and Ago2A reduced the expression of miR-276-3p, increased CYP6CX3 expression, and decreased B. tabaci resistance to cyantraniliprole. These results improve our understanding of the role of miRNAs in P450 regulation and cyantraniliprole resistance in B. tabaci.

Malignant a-to-I RNA Editing By ADAR1 Drives T-Cell Acute Lymphoblastic Leukemia Relapse Via Attenuating dsRNA Sensing

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematological malignancy that frequently occurs in children, adolescents, and young adults. Approximately 10-20% of T-ALL patients will experience relapse months or years following remission and will often become refractory to further treatments. The survival of relapsed/refractory patients is very poor, with an overall survival rate of less than a 25% overall survival rate. Relapsed patients often have enriched pools of leukemia initiating cells (LICs) with enhanced pro-survival and self-renewal capacity, suggesting a potential vulnerable population for effective targeted therapies with less toxicity. An emerging research topic in LIC biology is the identification of RNA modifying enzymes that are important for LIC self-renewal and survival. ADAR1 enzymes catalyze the transition of adenosine (A) to inosine (I) in precursor double-stranded RNA (dsRNA). Epitranscriptomic A-to-I RNA editing events are widespread in the cancer transcriptome and are critical for the transition from pre-leukemic cells to fully functional LICs. Compared to myeloid leukemia, the role of ADAR1 in lymphoid progenitor maintenance and malignant transformation is not well understood . A-to-I RNA editing has a wide range of effects on RNA biology including gene expression, splicing, RNA degradation and translation, and miRNA biogenesis and/or 3' UTR targeting. The best documented functional roles of ADAR1 are suppression of the interferon (IFN) response and RNA editing of self-dsRNA to prevent abnormal activation of cytosolic self-dsRNA sensing. Concurrent deletion of the cytosolic dsRNA sensors melanoma differentiation-associated protein 5 (MDA5) and protein kinase R (PKR) is able to completely rescue embryo death and reverse the IFN signatures. Whether editing of immunogenic dsRNA and suppression of aberrant dsRNA sensing pathway could enhance LIC self-renewal capacities is an important question that has not been extensively addressed. In this study, we applied bioinformatic analysis on a large cohort of T-ALL samples to examine the function of ADAR1 in the context of T-ALL LIC maintenance. We found that ADAR1 is highly expressed in ~70% of all T-ALL patients and particularly within the LIC compartment. A thorough comparison of the A-to-I RNA editing landscape between non-relapsed and relapsed T-ALL patient cohorts revealed hyper-editing is associated with both increased risk of relapse and leukemia-associated mortality. A total of 338 under-edited and 1,472 over-edited sites were found in relapsed patients compared to non-relapsed samples. However, there was very little difference in ADAR1 expression and the overall A-to-I RNA editing levels, among various molecular subtypes of T-ALL, suggesting malignant A-to-I RNA editing is a common attribute of relapsed T-ALL regardless of the genetic mutation status. We also performed functional study in a three-dimensional human thymic organoid system and a T-ALL patient-derived xenograft (PDX) model. Depletion of ADAR1 showed striking effects in LIC survival and self-renewal with 50-90% reduction in leukemia growth. Mechanistically, we revealed complex dsRNA regulatory mechanisms of ADAR1 by directing hyper-editing of immunogenic dsRNA and retains unedited nuclear dsRNA to avoid detection by the innate immune sensor MDA5. Interesting, the dependency on ADAR1-MDA5 axis various among patients depending on the cell intrinsic level of MDA5. Collectively, our results show that ADAR1 functions as a self-renewal factor that limits the sensing of endogenous dsRNA. Thus, targeting ADAR1 presents a safe and effective therapeutic strategy for eliminating T-ALL LICs.

Graphene enhances artemisinin production in the traditional medicinal plant Artemisia annua via dynamic physiological processes and miRNA regulation

We investigated the effects of graphene on the model herb Artemisia annua, which is renowned for producing artemisinin, a widely used pharmacological compound. Seedling growth and biomass were promoted when A. annua was cultivated with low concentrations of graphene, an effect which was attributed to a 1.4-fold increase in nitrogen uptake, a 15%–22% increase in chlorophyll fluorescence, and greater abundance of carbon cycling–related bacteria. Exposure to 10 or 20 mg/L graphene resulted in a ∼60% increase in H2O2, and graphene could act as a catalyst accelerator, leading to a 9-fold increase in catalase (CAT) activity in vitro and thereby maintaining reactive oxygen species (ROS) homeostasis. Importantly, graphene exposure led to an 80% increase in the density of glandular secreting trichomes (GSTs), in which artemisinin is biosynthesized and stored. This contributed to a 5% increase in artemisinin content in mature leaves. Interestingly, expression of miR828 was reduced by both graphene and H2O2 treatments, resulting in induction of its target gene AaMYB17, a positive regulator of GST initiation. Subsequent molecular and genetic assays showed that graphene-induced H2O2 inhibits micro-RNA (miRNA) biogenesis through Dicers and regulates the miR828–AaMYB17 module, thus affecting GST density. Our results suggest that graphene may contribute to yield improvement in A. annua via dynamic physiological processes together with miRNA regulation, and it may thus represent a new cultivation strategy for increasing yield capacity through nanobiotechnology.

Роль генов семейства Argonaute в эффектах активатора РНК-интерференции эноксацина на продолжительность жизни Drosophila melanogaster

Epigenetic mechanisms play a leading role in the regulation of gene expression and the coordination of biological processes, influencing the aging rate and the organism’s lifespan. An important role in the implementation of these mechanisms is played by small RNAs which suppress the activity of own targets through the RNA interference and provide the antiviral protection. Enoxacin is a unique inducer of RNA interference factors with potential geroprotective activity. Its effects have been identified to be mediated by miRNAs but other types of non-coding RNAs may also be involved. In this study, we have investigated the effect of enoxacin on the Drosophila melanogaster lifespan and first analyzed the contribution of Argonaute family genes to this effect which specifically ensure the biogenesis and functioning of miRNAs, siRNAs, and piRNAs.

Regulating the regulators: understanding miRNA biogenesis and decay in adult skeletal muscles.

Maintaining cellular homeostasis necessitates precise control of gene expression, a process that molds both the transcriptome and proteome to adapt to internal and external changes effectively. MicroRNAs (miRNAs) are small RNAs (~ 22nucleotides) belonging to a broad family of non-coding RNAs and are important regulators of gene expression. While numerous studies have advanced our understanding of the common processes underlying miRNA biogenesis and function, individual cell types in diverse organisms have evolved distinct mechanisms for regulating them. In this current issue, Satoshi Oikawa and colleagues delve into the molecular dynamics of miRNAs in adult skeletal muscles. Their research introduces intriguing new inquiries for further investigations to uncover alternative mechanisms of miRNA biogenesis in skeletal muscle.

N6-Methyladenosine mRNA Modification: From Modification Site Selectivity to Neurological Functions.

The development of various chemical methods has enabled scientists to decipher the distribution features and biological functions of RNA modifications in the past decade. In addition to modifying noncoding RNAs such as tRNAs and rRNAs, N6-methyladenosine (m6A) has been proven to be the most abundant internal chemical modification on mRNAs in eukaryotic cells and is also the most widely studied mRNA modification to date. Extensive studies have repeatedly demonstrated the important functions of m6A in various biological conditions, ranging from embryonic organ development to adult organ function and pathogenesis. Unlike DNA methylation which is relatively stable, the reversible m6A modification on mRNA is highly dynamic and easily influenced by various internal or external factors, such as cell type, developmental stage, nutrient supply, circadian rhythm, and environmental stresses.In this Account, we review our previous findings on the site selectivity mechanisms regulating m6A formation, as well as the physiological roles of m6A modification in cerebellum development and long-term memory consolidation. In our initial efforts to profile m6A in various types of mouse and human cells, we surprisingly found that the sequence motifs surrounding m6A sites were often complementary with the seed sequences of miRNAs. By manipulating the abundance of the miRNA biogenesis enzyme Dicer or individual miRNAs or mutating miRNA sequences, we were able to reveal a new role of nucleus localized miRNAs, which is to guide the m6A methyltransferase METTL3 to bind to mRNAs and to promote m6A formation. As a result, we partially answered the question of why only a small proportion of m6A motifs within an mRNA could have m6A modification at a certain time point. We further explored the functions of m6A modification in regulating brain development and brain functions. We found that cerebellum had the most severe defects when Mettl3 was knocked out in developing mouse embryonic brain and revealed that the underlying mechanisms could be attributed to aberrant mRNA splicing and enhanced cell apoptosis under m6A deficit conditions. On the other hand, knocking out Mettl3 in postnatal hippocampus did not cause morphological defects in the mouse brain but impaired the efficacy of long-term memory consolidation. Under learning stimuli, formation of m6A modifications could be detected on transcripts encoding proteins related to dendrite growth, synapse formation, and other memory related functions. Loss of m6A modifications on these transcripts would result in translation deficiency and reduced protein production, particularly in the translation of early response genes, and therefore would compromise the efficacy of long-term memory consolidation. Interestingly, excessive training sessions or increased training intensity could overcome such m6A deficiency related memory defects, which is likely due to the longer turnover cycle and the cumulative abundance of proteins throughout the training process. In addition to revealing the roles of m6A modification in regulating long-term memory formation, our work also demonstrated an effective method for studying memory formation efficacy. As the lack of an appropriate model for studying memory formation efficacy has been a long-lasting problem in the field of neural science, our hippocampus-specific postnatal m6A knockout model could also be utilized to study other questions related to memory formation efficacy.

Crosstalk Between m6A RNA Methylation and miRNA Biogenesis in Cancer: An Unholy Nexus.

N6-methyladenosine (m6A) is one of the most prevalent internal reversible chemical modification of RNAs in eukaryotes, which has attracted widespread attention recently owing to its regulatory roles in a plethora of normal developmental processes and human diseases like cancer. Deposition of the m6A mark on RNAs is mediated by the dynamic interplay between m6A regulatory proteins such as m6A RNA methyltransferases (m6A writers), m6A RNA demethylases (m6A erasers) and m6A RNA binding proteins (m6A readers). m6A regulators are ectopically expressed in various cancer types, often leading to aberrant expression of tumor-suppressor and oncogenic mRNAs either directly or indirectly via regulating the biogenesis of non-coding RNAs like miRNAs. miRNAs are tiny regulators of gene expression, which often impact various hallmarks of cancer and thus influence tumorigenesis. It is becoming increasingly clear that m6A RNA modification impacts biogenesis and function of miRNAs, and recent studies have interestingly, uncovered many miRNAs whose biogenesis and function are regulated by m6A writers, erasers and readers. In this review, we discuss various mechanisms by which m6A RNA methylation regulates miRNA biogenesis, the functional crosstalk between m6A RNA methylation and miRNAs and how it modulates various aspects of tumorigenesis. The potential of m6A RNA methylation regulated miRNAs as biomarkers and novel therapeutic targets to treat various cancers is also addressed.

The Glycine-Rich RNA-Binding Protein Is a Vital Post-Transcriptional Regulator in Crops.

Glycine-rich RNA binding proteins (GR-RBPs), a branch of RNA binding proteins (RBPs), play integral roles in regulating various aspects of RNA metabolism regulation, such as RNA processing, transport, localization, translation, and stability, and ultimately regulate gene expression and cell fate. However, our current understanding of GR-RBPs has predominantly been centered on Arabidopsis thaliana, a model plant for investigating plant growth and development. Nonetheless, an increasing body of literature has emerged in recent years, shedding light on the presence and functions of GRPs in diverse crop species. In this review, we not only delineate the distinctive structural domains of plant GR-RBPs but also elucidate several contemporary mechanisms of GR-RBPs in the post-transcriptional regulation of RNA. These mechanisms encompass intricate processes, including RNA alternative splicing, polyadenylation, miRNA biogenesis, phase separation, and RNA translation. Furthermore, we offer an exhaustive synthesis of the diverse roles that GR-RBPs fulfill within crop plants. Our overarching objective is to provide researchers and practitioners in the field of agricultural genetics with valuable insights that may inform and guide the application of plant genetic engineering for enhanced crop development and sustainable agriculture.

MicroRNAs in Endometriosis: Insights into Inflammation and Progesterone Resistance.

Endometriosis, a non-malignant gynecological disorder influenced by estrogen, involves the growth of endometrial tissue outside the uterus. Its development includes processes such as inflammation, progesterone resistance, angiogenesis, and cell proliferation. Epigenetic factors, particularly the dysregulation of microRNAs (miRNAs), have emerged as key factors in these mechanisms in endometriosis. This review aims to unveil the intricate molecular processes that control inflammation, progesterone resistance, and miRNA functions in endometriosis. In addition, it provides a comprehensive overview of the current understanding regarding the involvement of miRNAs in the inflammatory aspects of this condition. This synthesis encompasses research investigating the molecular underpinnings of inflammation, along with the biogenesis and roles of miRNAs in endometriosis. Furthermore, it examines human studies and functional analyses to establish the intricate connection between miRNAs, inflammation, and progesterone resistance in the context of endometriosis. The results highlight the significant impact of dysregulated miRNAs on the inflammatory pathways and hormonal imbalances characteristic of endometriosis. Consequently, miRNAs hold promise as potential non-invasive biomarkers and targeted therapeutic agents aimed at addressing inflammation and enhancing the response to progesterone treatment in individuals with endometriosis.

FUS-dependent microRNA deregulations identify TRIB2 as a druggable target for ALS motor neurons

MicroRNAs (miRNAs) modulate mRNA expression, and their deregulation contributes to various diseases including amyotrophic lateral sclerosis (ALS). As fused in sarcoma (FUS) is a causal gene for ALS and regulates biogenesis of miRNAs, we systematically analyzed the miRNA repertoires in spinal cords and hippocampi from ALS-FUS mice to understand how FUS-dependent miRNA deregulation contributes to ALS. miRNA profiling identified differentially expressed miRNAs between different central nervous system (CNS) regions as well as disease states. Among the up-regulated miRNAs, miR-1197 targets the pro-survival pseudokinase Trib2. A reduced TRIB2 expression was observed in iPSC-derived motor neurons from ALS patients. Pharmacological stabilization of TRIB2 protein with a clinically approved cancer drug rescues the survival of iPSC-derived human motor neurons, including those from a sporadic ALS patient. Collectively, our data indicate that miRNA profiling can be used to probe the molecular mechanisms underlying selective vulnerability, and TRIB2 is a potential therapeutic target for ALS.

FRI589 MicroRNAs In Maternal Plasma At 15 Weeks Gestation As Biomarkers For Later Preterm Birth: Evidence For Dysregulated Mirna Biogenesis And Implications For Pregnancy-related Endocrine Signaling Pathways

Abstract Disclosure: F. Ramzan: None. J. Rong: None. C. Roberts: None. J. O'Sullivan: None. J.K. Perry: None. R. Taylor: None. L. McCowan: None. M.H. Vickers: None. Introduction: Preterm birth (PTB) is a leading cause of morbidity and mortality in infants, with rates of PTB increasing globally over the last 20 years. MicroRNAs (miRNAs) have been reported to have widespread involvement in the development of and protection from many diseases, there is increasing interest in their potential role in the aetiology of a range of pregnancy-related disorders, including PTB. Objectives: To investigate differential miRNA abundances in mid-pregnancy plasma samples between mothers with term infants compared with moderate to late preterm infants (spontaneous PTB) and replication across independent cohorts. Method: The Auckland and Adelaide arms of the Screening for Pregnancy Endpoints (SCOPE) study were utilised. miRNAs in plasma at 15 weeks gestation from mothers of term and moderate to late preterm infants (n=54 per group) were quantified using the NanoString nCounter human v2 miRNA expression panel. Gene expression of key mediators of miRNA biogenesis, including Drosha, Dicer and Argonaut were quantified via RT-qPCR. In-silico analysis was performed to investigate the biological pathways associated with the dysregulated miRNAs. Results: Of the 800 miRNAs examined, the top four miRNAs identified were conserved across both cohorts and were significantly downregulated in the plasma of mothers who later experienced a PTB. The gene pathways associated with the top candidate miRNAs were linked to inflammatory processes, mitochondrial biogenesis, apoptosis and regulation of key pregnancy-related hormonal signaling pathways. Moreover, miRNAs were consistently downregulated in the PTB group concomitant with reduced gene expression of key miRNA biogenesis markers. Conclusions: We identified miRNAs in maternal plasma in mid-pregnancy that were highly predictive of later spontaneous PTB and, importantly, were conserved across two independent cohorts. Moreover, in addition to identifying potential early biomarkers for subsequent PTB, we provide evidence for persistent downregulation of miRNAs in those mothers who later delivered preterm, which is likely to have widespread effects on endocrine signaling pathways during pregnancy. Presentation: Friday, June 16, 2023

Parameters of clustered suboptimal miRNA biogenesis

The nuclear cleavage of a suboptimal primary miRNA hairpin by the Drosha/DGCR8 complex ("Microprocessor") can be enhanced by an optimal miRNA neighbor, a phenomenon termed cluster assistance. Several features and biological impacts of this new layer of miRNA regulation are not fully known. Here, we elucidate the parameters of cluster assistance of a suboptimal miRNA and also reveal competitive interactions amongst optimal miRNAs within a cluster. We exploit cluster assistance as a functional assay for suboptimal processing and use this to invalidate putative suboptimal substrates, as well as identify a "solo" suboptimal miRNA. Finally, we report complexity in how specific mutations might affect the biogenesis of clustered miRNAs in disease contexts. This includes how an operon context can buffer the effect of a deleterious processing variant, but reciprocally how a point mutation can have a nonautonomous effect to impair the biogenesis of a clustered, suboptimal, neighbor. These data expand our knowledge regarding regulated miRNA biogenesis in humans and represent a functional assay for empirical definition of suboptimal Microprocessor substrates.

New insights of miRNA molecular mechanisms in breast cancer brain metastasis and therapeutic targets

Brain metastases in breast cancer (BC) patients are often associated with a poor prognosis. Recent studies have uncovered the critical roles of miRNAs in the initiation and progression of BC brain metastasis, highlighting the disease's underlying molecular pathways. miRNA-181c, miRNA-10b, and miRNA-21, for example, are all overexpressed in BC patients. It has been shown that these three miRNAs help tumors grow and metastasize by targeting genes that control how cells work. On the other hand, miRNA-26b5p, miRNA-7, and miRNA-1013p are all downregulated in BC brain metastasis patients. They act as tumor suppressors by controlling the expression of genes related to cell adhesion, angiogenesis, and invasion. Therapeutic miRNA targeting has considerable promise in treating BC brain metastases. Several strategies have been proposed to modulate miRNA expression, including miRNA-Mimics, antagomirs, and small molecule inhibitors of miRNA biogenesis. This review discusses the aberrant expression of miRNAs and metastatic pathways that lead to the spread of BC cells to the brain. It also explores miRNA therapeutic target molecular mechanisms and BC brain metastasis challenges with advanced strategies. The targeting of certain miRNAs opens a new door for the development of novel therapeutic approaches for this devastating disease.

B-377 Mirsnps as Potential Colorectal Cancer Biomarkers: An Overview

Abstract Background Colorectal cancer (CRC), also known as colon cancer, is a neoplasm that affects the colon (large intestine) or rectum. CRC stands out for being the third most frequent neoplasm in the world population, being the second cancer with the highest mortality rate. Single nucleotide polymorphisms (SNPs) located in miRNAs genes, known as miRSNPs, may be associated with dysregulated expression of miRNAs in neoplasms. The objective of this review was to highlight the SNPs that alter the biogenesis or processing of miRNAs associated with CRC as potential biomarkers of this disease, according to the literature. Methods A search of original articles in the literature was performed in PubMed, Embase and Web of Science databases. The bibliographic search included scientific articles published between 2008 and 2022, with the objective of investigating associations between miRSNPs and CRC. Exclusion criteria were brief communications and other reviews on the subject. A total of 263 articles were detected in the preliminary search, of which 133 were excluded for being replicated, 34 for being reviewers, 3 for being paid and 40 for dealing with other subjects. Thus, of the remaining 53 studies, 35 were excluded after reading because they included studies that described SNPs located in miRNA genes and SNPs located in miRNA binding sites. Finally, 18 articles that included SNPs located in the regulatory regions of miRNAs were selected for this review. The list of genes generated for each of the targets was performed using the online tool miRBase and miRDB, for prediction of miRNA targets and functional annotations. The miRTarBase online database was consulted to select target miRNAs and possible interactions. By crossing the list of genes targeted by miRNAs to identify the biological pathways, those with the greatest biological relevance were chosen using the Enrichr online tool. Results This review covered 18 studies related to SNPs located in regulatory regions of mature miRNAs that may influence the occurrence and progression of CRC. In order to characterize the metabolic profile induced by each target of the miRNAs selected in this review, it was possible to observe that miR-196, miR-211, miR-30a, miR-143, miR-27 and miR-146a are more strongly associated with positive regulation of transcription. Cytokine-mediated signaling pathway and regulation of cell migration were most strongly associated with miR-143 and miR-146a. Furthermore, it was identified that the upregulation of the apoptosis process was strongly associated with miR-30a, miR-143, miR-27a and miR-146a. MiR-211, miR-143, mi-27a and miR-146a were strongly associated with the upregulation of cell proliferation. Finally, with the objective of evaluating the correlation between the selected miRNAs. It was possible to observe a greater correlation between miR-27a and miR-30a. Conclusion Studies related to miRSNPs have been of great interest in recent years in order to find promising and less invasive new biomarkers, sensitive and specific for the early detection of CRC. Our study aims to help better understand the pathways that cause the development of CRC and possible biomarkers to predict its development and consequently reduce morbidity and mortality and improve patient survival.

B-288 The Beginning of the End: AGO2 Protein Identified and Validated as a Novel Biomarker for Multi-cancer Diagnosis Through Bioinformatics Analysis and Immunoassay

Abstract Background Despite significant progress in cancer research, the complexity and heterogeneity of the disease still pose major challenges to accurate diagnosis and effective treatment. Recent advancements in biomedical technologies have led to the identification of reliable potential cancer biomarkers. In this sense, the link between microRNAs (master regulators of gene expression) & cancer is well-established, but there has been little research on the microRNA machinery itself. We aim to investigate whether changes in the expression of genes & proteins involved in the miRNA pathway can be used for multi-cancer detection. Method We analyzed RNASeq data from the TCGA (The Cancer Genome Atlas) and GTEX (The Genotype-Tissue Expression) projects to compare the expression of core components in the miRNA biogenesis pathway, including AGO2, DGCR8, XPO5, RAN, DROSHA, DICER, and TARBP2, in normal and tumorous samples across 22 tissue types, including adrenocortical, AML, breast, colon, lung, liver, esophageal, prostate, pancreas, stomach, thyroid, rectum, uterus and ovarian cancer. We accessed and analyzed the TCGA and GTEx datasets using the UCSC Xena Browser. We also measured the protein concentrations of all these microRNA regulators in 15 normal adrenal cortex, 15 benign adenoma, and 15 adrenocortical cancer tissue homogenate samples using commercial ELISA kits. The Human Protein Argonaute-2 (EIF2C2) ELISA Kit-AE45910HU (Abebio-Co.Ltd) was used for the assay procedure, which was performed according to the manufacturer’s instructions. Statistical analysis was performed using GraphPad Prism, Version 9 (GraphPad Software, CA, USA). A P-value of less than 0.05 was considered statistically significant. Results Our RNASeq analysis results showed that among all microRNA regulators, AGO2 is most highly significantly over-expressed in samples derived from cancer patients compared with normal controls (P < 0.05). In addition, we analysed the protein expression of AGO2 in adrenocortical tissue homogenate samples. Our results demonstrated that the protein expression of AGO2 in adrenocortical carcinoma samples was significantly higher than in normal adrenal cortex and benign adrenocortical tumour samples (P < 0.001). Conclusion Our study highlights the potential role of Argonaute 2 protein (AGO2) in cancer development and progression. AGO2 is a key component of the RNA-induced silencing complex, which regulates gene expression. Our findings show that AGO2 is overexpressed in cancer samples compared to normal controls across a range of cancer types. This suggest that AGO2 could be used as a promising marker for the development of more accurate and specific methods for cancer diagnosis, in combination with other markers. Furthermore, the potential for using AGO2 protein expression in serum samples as a non-invasive diagnostic tool is a promising area for future research. This could be achieved through automated immunoassay, making it a practical diagnostic tool for use in routine clinical practice.

Dissecting key regulators of transcriptome kinetics through scalable single-cell RNA profiling of pooled CRISPR screens

We present a combinatorial indexing method, PerturbSci-Kinetics, for capturing whole transcriptomes, nascent transcriptomes and single guide RNA (sgRNA) identities across hundreds of genetic perturbations at the single-cell level. Profiling a pooled CRISPR screen targeting various biological processes, we show the gene expression regulation during RNA synthesis, processing and degradation, miRNA biogenesis and mitochondrial mRNA processing, systematically decoding the genome-wide regulatory network that underlies RNA temporal dynamics at scale.

Structure of pre-miR-31 reveals an active role in Dicer–TRBP complex processing

As an essential posttranscriptional regulator of gene expression, microRNA (miRNA) levels must be strictly maintained. The biogenesis of many miRNAs is mediated by trans-acting protein partners through a variety of mechanisms, including remodeling of the RNA structure. miR-31 functions as an oncogene in numerous cancers, and interestingly, its biogenesis is not known to be regulated by protein-binding partners. Therefore, the intrinsic structural properties of the precursor element of miR-31 (pre-miR-31) can provide a mechanism by which its biogenesis is regulated. We determined the solution structure of pre-miR-31 to investigate the role of distinct structural elements in regulating processing by the Dicer-TRBP complex. We found that the presence or absence of mismatches within the helical stem does not strongly influence Dicer-TRBP processing of the pre-miRNAs. However, both the apical loop size and structure at the Dicing site are key elements for discrimination by the Dicer-TRBP complex. Interestingly, our NMR-derived structure reveals the presence of a triplet of base pairs that link the Dicer cleavage site and the apical loop. Mutational analysis in this region suggests that the stability of the junction region strongly influences processing by the Dicer-TRBP complex. Our results enrich our understanding of the active role that RNA structure plays in regulating miRNA biogenesis, which has direct implications for the control of gene expression.

Exercise Training-Induced MicroRNA Alterations with Protective Effects in Cardiovascular Diseases.

Exercise training (ET) is an important non-drug adjuvant therapy against many human diseases, including cardiovascular diseases. The appropriate ET intensity induces beneficial adaptions and improves physiological function and cardiopulmonary fitness. The mechanisms of exercise-induced cardioprotective effects are still not fully understood. However, mounting evidence suggest that microRNAs (miRNAs) play crucial role in this process and are essential in responding to exercise-stress and mediating exercise-protective effects. Thus, this review summarizes the biogenesis of miRNAs, the mechanism of miRNA action, and specifically the miRNAs involved in exercise-induced cardio-protection used as therapeutic targets for treating cardiovascular diseases.