Complete mitochondrial genome of Melia azedarach L., reveals two conformations generated by the repeat sequence mediated recombination.

Melia azedarach is a species of enormous value of pharmaceutical industries. Although the chloroplast genome of M. azedarach has been explored, the information of mitochondrial genome (Mt genome) remains surprisingly limited. In this study, we used a hybrid assembly strategy of BGI short-reads and Nanopore long-reads to assemble the Mt genome of M. azedarach. The Mt genome of M. azedarach is characterized by two circular chromosomes with 350,142 bp and 290,387 bp in length, respectively, which encodes 35 protein-coding genes (PCGs), 23 tRNA genes, and 3 rRNA genes. A pair of direct repeats (R1 and R2) were associated with genome recombination, resulting in two conformations based on the Sanger sequencing and Oxford Nanopore sequencing. Comparative analysis identified 19 homologous fragments between Mt and chloroplast genome, with the longest fragment of 12,142 bp. The phylogenetic analysis based on PCGs were consist with the latest classification of the Angiosperm Phylogeny Group. Notably, a total of 356 potential RNA editing sites were predicted based on 35 PCGs, and the editing events lead to the formation of the stop codon in the rps10 gene and the start codons in the nad4L and atp9 genes, which were verified by PCR amplification and Sanger sequencing. Taken together, the exploration of M. azedarach gap-free Mt genome provides a new insight into the evolution research and complex mitogenome architecture.

RNA editing landscape of adipose tissue in polycystic ovary syndrome provides insight into the obesity-related immune responses.

Background: Polycystic ovary syndrome (PCOS) is the most common reproductive-endocrine disorder with wide-ranging metabolic implications, including obesity. RNA editing, a post-transcriptional modification, can fine-tune protein function and introduce heterogeneity. However, the role of RNA editing and its impact on adipose tissue function in PCOS remain poorly understood. Methods: This study aimed to comprehensively analyze RNA-editing events in abdominal and subcutaneous adipose tissue of PCOS patients and healthy controls using high-throughput whole-genome sequencing (WGS) and RNA sequencing. Results: Our results revealed that PCOS patients exhibited more RNA-editing sites, with adenosine-to-inosine (A-to-I) editing being prevalent. The expression of ADAR genes, responsible for A-to-I editing, was also higher in PCOS. Aberrant RNA-editing sites in PCOS adipose tissue was enriched in immune responses, and interleukin-12 biosynthetic process. Tumor necrosis factor (TNF) signaling, nuclear factor kappa B (NF-κB) signaling, Notch signaling, terminal uridylyl transferase 4 (TUT4), hook microtubule tethering protein 3 (HOOK3), and forkhead box O1 (FOXO1) were identified to be of significant differences. Differentially expressed genes (DEGs) in PCOS adipose tissue were enriched in immune responses compared with controls, and the DEGs between subcutaneous and abdominal adipose tissue were also enriched in immune responses suggesting the important role of subcutaneous adipose tissue. Furthermore, we identified the correlations between RNA editing levels and RNA expression levels of specific genes, such as ataxia-telangiectasia mutated (ATM) and mucosa-associated lymphoid tissue lymphoma translocation protein 1 (MALT1) in inflammation pathways and ATM, TUT4, and YTH N6-methyladenosine RNA-binding protein C2 (YTHDC2) in oocyte development pathway. Conclusions: These findings suggest that RNA-editing dysregulation in PCOS adipose tissue may contribute to inflammatory dysregulations. Understanding the interplay between RNA editing and adipose tissue function may unveil potential therapeutic targets for PCOS management. However, further research and validation are required to fully elucidate the molecular mechanisms underlying these associations.

The HPE1 RNA-binding protein modulates chloroplast RNA editing to promote photosynthesis under cold stress in Arabidopsis.

Cold stress has severe negative consequences for plant growth and crop yield. Here, we report that an Arabidopsis thaliana mutant that lacks the HPE1 gene, which encodes an RNA-binding protein, maintains higher photosynthetic activity under cold stress, together with higher accumulation of thylakoid proteins. We showed that HPE1 interacts with MORF2 and MORF9 and thereby mediates RNA editing in chloroplasts. Loss of HPE1 function increased the editing efficiency at four RNA editing sites, rpoC-488, ndhB-149, ndhB-746 and matK-706, under cold stress and altered the expression of nuclear photosynthesis-related genes and cold-responsive genes. We propose that HPE1-mediated RNA editing acts as a trigger for retrograde signaling that affects photosynthesis under cold stress.

ADAR-Mediated A>I(G) RNA Editing in the Genotoxic Drug Response of Breast Cancer.

Epitranscriptomics is a field that delves into post-transcriptional changes. Among these modifications, the conversion of adenosine to inosine, traduced as guanosine (A>I(G)), is one of the known RNA-editing mechanisms, catalyzed by ADARs. This type of RNA editing is the most common type of editing in mammals and contributes to biological diversity. Disruption in the A>I(G) RNA-editing balance has been linked to diseases, including several types of cancer. Drug resistance in patients with cancer represents a significant public health concern, contributing to increased mortality rates resulting from therapy non-responsiveness and disease progression, representing the greatest challenge for researchers in this field. The A>I(G) RNA editing is involved in several mechanisms over the immunotherapy and genotoxic drug response and drug resistance. This review investigates the relationship between ADAR1 and specific A>I(G) RNA-edited sites, focusing particularly on breast cancer, and the impact of these sites on DNA damage repair and the immune response over anti-cancer therapy. We address the underlying mechanisms, bioinformatics, and in vitro strategies for the identification and validation of A>I(G) RNA-edited sites. We gathered databases related to A>I(G) RNA editing and cancer and discussed the potential clinical and research implications of understanding A>I(G) RNA-editing patterns. Understanding the intricate role of ADAR1-mediated A>I(G) RNA editing in breast cancer holds significant promise for the development of personalized treatment approaches tailored to individual patients' A>I(G) RNA-editing profiles.

F11R RNA trinucleotide over-edited by ADAR in gastric and colorectal cancers: Cross-cohort validation, gene expression regulation, and diagnostic significance.

The F11 receptor (F11R) gene encoding junctional adhesion molecule A has been associated with gastric cancer (GC) and colorectal cancer (CRC), in which its role and regulation remain to be further elucidated. Recently F11R was also identified as a potential target of adenosine-to-inosine (A-to-I) mediated by the adenosine deaminases acting on RNA (ADARs). Herein, using RNA-Seq and experimental validation, our current study revealed an F11R RNA trinucleotide over-edited by ADAR, with its regulation of gene expression and clinical significance in four GC and three CRC cohorts. Our results found an over-edited AAA trinucleotide in an AluSg located in the F11R 3'-untranslated region (3'-UTR), which showed editing levels correlated with elevated ADAR expression across all GC and CRC cohorts in our study. Overexpression and knockdown of ADAR in GC and CRC cells, followed by RNA-Seq and Sanger sequencing, confirmed the ADAR-mediated F11R 3'-UTR trinucleotide editing, which potentially disrupted an RBM45 binding site identified by crosslinking immunoprecipitation sequencing (CLIP-seq) and regulated F11R expression in luciferase reporter assays. Moreover, the F11R trinucleotide editing showed promising predictive performance for diagnosing GC and CRC across GC and CRC cohorts. Our findings thus highlight both the potential biological and clinical significance of an ADAR-edited F11R trinucleotide in GC and CRC, providing new insights into its application as a novel diagnostic biomarker for both cancers.

Assembly and evolutionary analysis of the complete mitochondrial genome of Trichosanthes kirilowii, a traditional Chinese medicinal plant.

Trichosanthes kirilowii (T. kirilowii) is a valuable plant used for both medicinal and edible purposes. It belongs to the Cucurbitaceae family. However, its phylogenetic position and relatives have been difficult to accurately determine due to the lack of mitochondrial genomic information. This limitation has been an obstacle to the potential applications of T. kirilowii in various fields. To address this issue, Illumina and Nanopore HiFi sequencing were used to assemble the mitogenome of T. kirilowii into two circular molecules with sizes of 245,700 bp and 107,049 bp, forming a unique multi-branched structure. The mitogenome contains 61 genes, including 38 protein-coding genes (PCGs), 20 tRNAs, and three rRNAs. Within the 38 PCGs of the T. kirilowii mitochondrial genome, 518 potential RNA editing sites were identified. The study also revealed the presence of 15 homologous fragments that span both the chloroplast and mitochondrial genomes. The phylogenetic analysis strongly supports that T. kirilowii belongs to the Cucurbitaceae family and is closely related to Luffa. Collinearity analysis of five Cucurbitaceae mitogenomes shows a high degree of structural variability. Interestingly, four genes, namely atp1, ccmFC, ccmFN, and matR, played significant roles in the evolution of T. kirilowii through selection pressure analysis. The comparative analysis of the T. kirilowii mitogenome not only sheds light on its functional and structural features but also provides essential information for genetic studies of the genus of Cucurbitaceae.

Base preference for inosine 3'-riboendonuclease activity of human endonuclease V: implications for cleavage of poly-A tails containing inosine.

Deamination of bases is a form of DNA damage that occurs spontaneously via the hydrolysis and nitrosation of living cells, generating hypoxanthine from adenine. E. coli endonuclease V (eEndoV) cleaves hypoxanthine-containing double-stranded DNA, whereas human endonuclease V (hEndoV) cleaves hypoxanthine-containing RNA; however, hEndoV in vivo function remains unclear. To date, hEndoV has only been examined using hypoxanthine, because it binds closely to the base located at the cleavage site. Here, we examined whether hEndoV cleaves other lesions (e.g., AP site, 6-methyladenine, xanthine) to reveal its function and whether 2'-nucleoside modification affects its cleavage activity. We observed that hEndoV is hypoxanthine-specific; its activity was the highest with 2'-OH modification in ribose. The cleavage activity of hEndoV was compared based on its base sequence. We observed that it has specificity for adenine located on the 3'-end of hypoxanthine at the cleavage site, both before and after cleavage. These data suggest that hEndoV recognizes and cleaves the inosine generated on the poly A tail to maintain RNA quality. Our results provide mechanistic insight into the role of hEndoV in vivo.

ADR-2 regulates fertility and oocyte fate in C. elegans.

RNA binding proteins play essential roles in coordinating germline gene expression and development in all organisms. Here, we report that loss of ADR-2, a member of the Adenosine DeAminase acting on RNA (ADAR) family of RNA binding proteins and the sole adenosine-to-inosine RNA editing enzyme in C. elegans, can improve fertility in multiple genetic backgrounds. First, we show that loss of RNA editing by ADR-2 restores normal embryo production to subfertile animals that transgenically express a vitellogenin (yolk protein) fusion to green fluorescent protein. Using this phenotype, a high-throughput screen was designed to identify RNA binding proteins that when depleted yield synthetic phenotypes with loss of adr-2. The screen uncovered a genetic interaction between ADR-2 and SQD-1, a member of the heterogenous nuclear ribonucleoprotein (hnRNP) family of RNA binding proteins. Microscopy, reproductive assays, and high-throughput sequencing reveal that sqd-1 is essential for the onset of oogenesis and oogenic gene expression in young adult animals, and that loss of adr-2 can counteract the effects of loss of sqd-1 on gene expression and rescue the switch from spermatogenesis to oogenesis. Together, these data demonstrate that ADR-2 can contribute to the suppression of fertility and suggest novel roles for both RNA editing-dependent and independent mechanisms in regulating embryogenesis.

ADAR1: from basic mechanisms to inhibitors.

Adenosine deaminase acting on RNA 1 (ADAR1) converts adenosine to inosine in double-stranded RNA (dsRNA) molecules, a process known as A-to-I editing. ADAR1 deficiency in humans and mice results in profound inflammatory diseases characterised by the spontaneous induction of innate immunity. In cells lacking ADAR1, unedited RNAs activate RNA sensors. These include melanoma differentiation-associated gene 5 (MDA5) that induces the expression of cytokines, particularly type I interferons (IFNs), protein kinase R (PKR), oligoadenylate synthase (OAS), and Z-DNA/RNA binding protein 1 (ZBP1). Immunogenic RNAs 'defused' by ADAR1 may include transcripts from repetitive elements and other long duplex RNAs. Here, we review these recent fundamental discoveries and discuss implications for human diseases. Some tumours depend on ADAR1 to escape immune surveillance, opening the possibility of unleashing anticancer therapies with ADAR1 inhibitors.

RNA editing sites and triplet usage in exomes of bat RNA virus genomes of the family Paramyxoviridae.

Bats contain a diverse spectrum of viral species in their bodies. The RNA virus family Paramyxoviridae tends to infect several vertebrate species, which are accountable for a variety of devastating infections in both humans and animals. Viruses of this kind include measles, mumps, and Hendra. Some synonymous codons are favoured over others in mRNAs during gene-to-protein synthesis process. Such phenomenon is termed as codon usage bias (CUB). Our research emphasized many aspects that shape the CUB of genes in the Paramyxoviridae family found in bats. Here, the nitrogenous base A occurred the most. AT was found to be abundant in the coding sequences of the Paramyxoviridae family. RSCU data revealed that A or T ending codons occurred more frequently than predicted. Furthermore, 3 overrepresented codons (CAT, AGA, and GCA) and 7 underrepresented codons (CCG, TCG, CGC, CGG, CGT, GCG and ACG) were detected in the viral genomes. Correspondence analysis, neutrality plot, and parity plots highlight the combined impact of mutational pressure and natural selection on CUB. The neutrality plot of GC12 against GC3 yielded a regression coefficient value of 0.366, indicating that natural selection had a significant (63.4 %) impact. Moreover, RNA editing analysis was done, which revealed the highest frequency of C to T mutations. The results of our research revealed the pattern of codon usage and RNA editing sites in Paramyxoviridae genomes.

The complete mitochondrial genome of Pontederia crassipes: using HiFi reads to investigate genome recombination and gene transfer from chloroplast genome.

Water hyacinth (Pontederia crassipes Mart.) is a monocotyledonous aquatic plant renowned for its rapid growth, extensive proliferation, biological invasiveness, and ecological resilience to variations in pH, nutrients, and temperature. The International Union for Conservation of Nature (IUCN) has listed P. crassipes among the top 100 invasive species. However, comprehensive genomic information, particularly concerning its mitochondrial genome (mitogenome), remains surprisingly limited. In this study, the complete mitogenome of P. crassipes was analyzed using bioinformatics approaches. The mitogenome is 399,263 bp long and contains 38 protein-coding genes (PCGs), 24 tRNA genes, and 3 rRNA genes. Sequence analysis revealed that the complete mitogenome of the species contains 3,289 dispersed repeats, and 765 RNA editing sites in protein-coding genes. The P. crassipes mitogenome possessed un-conserved structures, including extensive sequence transfer between its chloroplasts and mitochondria. Our study on the mitogenome of P. crassipes offers critical insights into its evolutionary patterns and phylogenetic relationships with related taxa. This research enhances our understanding of this invasive species, known for its significant biomass and rapid overgrowth in aquatic environments.

RNA editing regulates glutamatergic synapses in the frontal cortex of a molecular subtype of Amyotrophic Lateral Sclerosis.

BACKGROUND: Amyotrophic Lateral Sclerosis (ALS) is a highly heterogenous neurodegenerative disorder that primarily affects upper and lower motor neurons, affecting additional cell types and brain regions. Underlying molecular mechanisms are still elusive, in part due to disease heterogeneity. Molecular disease subtyping through integrative analyses including RNA editing profiling is a novel approach for identification of molecular networks involved in pathogenesis. METHODS: We aimed to highlight the role of RNA editing in ALS, focusing on the frontal cortex and the prevalent molecular disease subtype (ALS-Ox), previously determined by transcriptomic profile stratification. We established global RNA editing (editome) and gene expression (transcriptome) profiles in control and ALS-Ox cases, utilizing publicly available RNA-seq data (GSE153960) and an in-house analysis pipeline. Functional annotation and pathway analyses identified molecular processes affected by RNA editing alterations. Pearson correlation analyses assessed RNA editing effects on expression. Similar analyses on additional ALS-Ox and control samples (GSE124439) were performed for verification. Targeted re-sequencing and qRT-PCR analysis targeting CACNA1C, were performed using frontal cortex tissue from ALS and control samples (n = 3 samples/group). RESULTS: We identified reduced global RNA editing in the frontal cortex of ALS-Ox cases. Differentially edited transcripts are enriched in synapses, particularly in the glutamatergic synapse pathway. Bioinformatic analyses on additional ALS-Ox and control RNA-seq data verified these findings. We identified increased recoding at the Q621R site in the GRIK2 transcript and determined positive correlations between RNA editing and gene expression alterations in ionotropic receptor subunits GRIA2, GRIA3 and the CACNA1C transcript, which encodes the pore forming subunit of a post-synaptic L-type calcium channel. Experimental data verified RNA editing alterations and editing-expression correlation in CACNA1C, highlighting CACNA1C as a target for further study. CONCLUSIONS: We provide evidence on the involvement of RNA editing in the frontal cortex of an ALS molecular subtype, highlighting a modulatory role mediated though recoding and gene expression regulation on glutamatergic synapse related transcripts. We report RNA editing effects in disease-related transcripts and validated editing alterations in CACNA1C. Our study provides targets for further functional studies that could shed light in underlying disease mechanisms enabling novel therapeutic approaches.

Learning from the Codon Table: Convergent Recoding Provides Novel Understanding on the Evolution of A-to-I RNA Editing.

Adenosine-to-inosine (A-to-I) RNA editing recodes the genetic information. Apart from diversifying the proteome, another tempting advantage of RNA recoding is to correct deleterious DNA mutation and restore ancestral allele. Solid evidences for beneficial restorative editing are very rare in animals. By searching for "convergent recoding" under a phylogenetic context, we proposed this term for judging the potential restorative functions of particular editing site. For the well-known mammalian Gln>Arg (Q>R) recoding site, its ancestral state in vertebrate genomes was the pre-editing Gln, and all 470 available mammalian genomes strictly avoid other three equivalent ways to achieve Arg in protein. The absence of convergent recoding from His>Arg, or synonymous mutations on Gln codons, could be attributed to the strong maintenance on editing motif and structure, but the absence of direct A-to-G mutation is extremely unexpected. With similar ideas, we found cases of convergent recoding in Drosophila genus, reducing the possibility of their restorative function. In summary, we defined an interesting scenario of convergent recoding, the occurrence of which could be used as preliminary judgements for whether a recoding site has a sole restorative role. Our work provides novel insights to the natural selection and evolution of RNA editing.

Assembly and comparative analysis of the complete mitochondrial genome of Vaccinium carlesii Dunn.

Vaccinium L. is an important fruit tree with nutritional, medicinal, and ornamental values. However, the mitochondrial (mt) genome of Vaccinium L. remains largely unexplored. Vaccinium carlesii Dunn is an endemic wild resource in China, which is crucial for blueberry breeding. The V. carlesii mt genomes were sequenced using Illumina and Nanopore, which total length was 636,904 bp with 37 protein coding genes, 20 tRNA genes, and three rRNA genes. We found four pairs of long repeat fragments homologous recombination mediated the generation of substructures in the V. carlesii mt genome. We predicted 383 RNA editing sites, all converting cytosine (C) to uracil (U). According to the phylogenetic analysis, V. carlesii and V. macrocarpon of the Ericaceae exhibited the closest genetic relationship. This study provides a theoretical basis for understanding the evolution of higher plants, species classification and identification, and will also be useful for further utilization of Vaccinium germplasm resources.

Insights into structure, codon usage, repeats, and RNA editing of the complete mitochondrial genome of Perilla frutescens (Lamiaceae).

Perilla frutescens (L.) Britton, a member of the Lamiaceae family, stands out as a versatile plant highly valued for its unique aroma and medicinal properties. Additionally, P. frutescens seeds are rich in Îs-linolenic acid, holding substantial economic importance. While the nuclear and chloroplast genomes of P. frutescens have already been documented, the complete mitochondrial genome sequence remains unreported. To this end, the sequencing, annotation, and assembly of the entire Mitochondrial genome of P. frutescens were hereby conducted using a combination of Illumina and PacBio data. The assembled P. frutescens mitochondrial genome spanned 299,551 bp and exhibited a typical circular structure, involving a GC content of 45.23%. Within the genome, a total of 59 unique genes were identified, encompassing 37 protein-coding genes, 20 tRNA genes, and 2 rRNA genes. Additionally, 18 introns were observed in 8 protein-coding genes. Notably, the codons of the P. frutescens mitochondrial genome displayed a notable A/T bias. The analysis also revealed 293 dispersed repeat sequences, 77 simple sequence repeats (SSRs), and 6 tandem repeat sequences. Moreover, RNA editing sites preferentially produced leucine at amino acid editing sites. Furthermore, 70 sequence fragments (12,680 bp) having been transferred from the chloroplast to the mitochondrial genome were identified, accounting for 4.23% of the entire mitochondrial genome. Phylogenetic analysis indicated that among Lamiaceae plants, P. frutescens is most closely related to Salvia miltiorrhiza and Platostoma chinense. Meanwhile, inter-species Ka/Ks results suggested that Ka/Ks < 1 for 28 PCGs, indicating that these genes were evolving under purifying selection. Overall, this study enriches the mitochondrial genome data for P. frutescens and forges a theoretical foundation for future molecular breeding research.

APOBEC3-related mutations in the spike protein-encoding region facilitate SARS-CoV-2 evolution.

SARS-CoV-2 evolves gradually to cause COVID-19 epidemic. One of driving forces of SARS-CoV-2 evolution might be activation of apolipoprotein B mRNA editing catalytic subunit-like protein 3 (APOBEC3) by inflammatory factors. Here, we aimed to elucidate the effect of the APOBEC3-related viral mutations on the infectivity and immune evasion of SARS-CoV-2. The APOBEC3-related C > U mutations ranked as the second most common mutation types in the SARS-CoV-2 genome. mRNA expression of APOBEC3A (A3A), APOBEC3B (A3B), and APOBEC3G (A3G) in peripheral blood cells increased with disease severity. A3B, a critical member of the APOBEC3 family, was significantly upregulated in both severe and moderate COVID-19 patients and positively associated with neutrophil proportion and COVID-19 severity. We identified USP18 protein, a key molecule centralizing the protein-protein interaction network of key APOBEC3 proteins. Furthermore, mRNA expression of USP18 was significantly correlated to ACE2 and TMPRSS2 expression in the tissue of upper airways. Knockdown of USP18 mRNA significantly decreased A3B expression. Ectopic expression of A3B gene increased SARS-CoV-2 infectivity. C > U mutations at S371F, S373L, and S375F significantly conferred with the immune escape of SARS-CoV-2. Thus, APOBEC3, whose expression are upregulated by inflammatory factors, might promote SARS-CoV-2 evolution and spread via upregulating USP18 level and facilitating the immune escape. A3B and USP18 might be therapeutic targets for interfering with SARS-CoV-2 evolution.

Reovirus infection induces transcriptome-wide unique A-to-I editing changes in the murine fibroblasts.

The conversion of Adenosine (A) to Inosine (I), by Adenosine Deaminases Acting on RNA or ADARs, is an essential post-transcriptional modification that contributes to proteome diversity and regulation in metazoans including humans. In addition to its transcriptome-regulating role, ADARs also play a major part in immune response to viral infection, where an interferon response activates interferon-stimulated genes, such as ADARp150, in turn dynamically regulating host-virus interactions. A previous report has shown that infection from reoviruses, despite strong activation of ADARp150, does not influence the editing of some of the major known editing targets, while likely editing others, suggesting a potentially nuanced editing pattern that may depend on different factors. However, the results were based on a handful of selected editing sites and did not cover the entire transcriptome. Thus, to determine whether and how reovirus infection specifically affects host ADAR editing patterns, we analyzed a publicly available deep-sequenced RNA-seq dataset, from murine fibroblasts infected with wild-type and mutant reovirus strains that allowed us to examine changes in editing patterns on a transcriptome-wide scale. To the best of our knowledge, this is the first transcriptome-wide report on host editing changes after reovirus infection. Our results demonstrate that reovirus infection induces unique nuanced editing changes in the host, including introducing sites uniquely edited in infected samples. Genes with edited sites are overrepresented in pathways related to immune regulation, cellular signaling, metabolism, and growth. Moreover, a shift in editing targets has also been observed, where the same genes are edited in infection and control conditions but at different sites, or where the editing rate is increased for some and decreased for other differential targets, supporting the hypothesis of dynamic and condition-specific editing by ADARs.

Therapeutic Nonsense Suppression Modalities: From Small Molecules to Nucleic Acid-Based Approaches.

Nonsense mutations are genetic mutations that create premature termination codons (PTCs), leading to truncated, defective proteins in diseases such as cystic fibrosis, neurofibromatosis type 1, Dravet syndrome, Hurler syndrome, Beta thalassemia, inherited bone marrow failure syndromes, Duchenne muscular dystrophy, and even cancer. These mutations can also trigger a cellular surveillance mechanism known as nonsense-mediated mRNA decay (NMD) that degrades the PTC-containing mRNA. The activation of NMD can attenuate the consequences of truncated, defective, and potentially toxic proteins in the cell. Since approximately 20% of all single-point mutations are disease-causing nonsense mutations, it is not surprising that this field has received significant attention, resulting in a remarkable advancement in recent years. In fact, since our last review on this topic, new examples of nonsense suppression approaches have been reported, namely new ways of promoting the translational readthrough of PTCs or inhibiting the NMD pathway. With this review, we update the state-of-the-art technologies in nonsense suppression, focusing on novel modalities with therapeutic potential, such as small molecules (readthrough agents, NMD inhibitors, and molecular glue degraders); antisense oligonucleotides; tRNA suppressors; ADAR-mediated RNA editing; targeted pseudouridylation; and gene/base editing. While these various modalities have significantly advanced in their development stage since our last review, each has advantages (e.g., ease of delivery and specificity) and disadvantages (manufacturing complexity and off-target effect potential), which we discuss here.

Posttranscriptional Regulation by Proteins and Noncoding RNAs.

Posttranscriptional regulation comprises those mechanisms occurring after the initial copy of the DNA sequence is transcribed into an intermediate RNA molecule (i.e., messenger RNA) until such a molecule is used as a template to generate a protein. A subset of these posttranscriptional regulatory mechanisms essentially are destined to process the immature mRNA toward its mature form, conferring the adequate mRNA stability, providing the means for pertinent introns excision, and controlling mRNA turnover rate and quality control check. An additional layer of complexity is added in certain cases, since discrete nucleotide modifications in the mature RNA molecule are added by RNA editing, a process that provides large mature mRNA diversity. Moreover, a number of posttranscriptional regulatory mechanisms occur in a cell- and tissue-specific manner, such as alternative splicing and noncoding RNA-mediated regulation. In this chapter, we will briefly summarize current state-of-the-art knowledge of general posttranscriptional mechanisms, while major emphases will be devoted to those tissue-specific posttranscriptional modifications that impact on cardiac development and congenital heart disease.