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Expansion of CAG repeats in certain genes is a known cause of several neurodegenerative diseases, but exact mechanism behind this is not yet fully understood. It is believed that the double-stranded RNA regions formed by CAG repeats could be harmful to the cell. This study aimed to test the hypothesis that these RNA regions might potentially interfere with ADAR RNA editing enzymes, leading to the reduced A-to-I editing of RNA and activation of the interferon response. We studied induced pluripotent stem cells (iPSCs) derived from the patients with Huntington's disease or ataxia type 17, as well as midbrain organoids developed from these cells. A targeted panel for next-generation sequencing was used to assess editing in the specific RNA regions. Differentiation of iPSCs into brain organoids led to increase in the ADAR2 gene expression and decrease in the expression of protein inhibitors of RNA editing. As a result, there was increase in the editing of specific ADAR2 substrates, which allowed identification of differential substrates of ADAR isoforms. However, comparison of the pathology and control groups did not show differences in the editing levels among the iPSCs. Additionally, brain organoids with 42-46 CAG repeats did not exhibit global changes. On the other hand, brain organoids with the highest number of CAG repeats in the huntingtin gene (76) showed significant decrease in the level of RNA editing of specific transcripts, potentially involving ADAR1. Notably, editing of the long non-coding RNA PWAR5 was nearly absent in this sample. It could be stated in conclusion that in most cultures with repeat expansion, the hypothesized effect on RNA editing was not confirmed.

Adenosine-to-inosine RNA editing is a system of post-transcriptional modification widely distributed in metazoans which is catalyzed by ADAR enzymes and occurs mostly in double-stranded RNA (dsRNA) before splicing. This type of RNA editing changes the genetic code, as inosine generally pairs with cytosine in contrast to adenosine, and this expectably modulates RNA splicing. We review the interconnections between RNA editing and splicing in the context of human cancer. The editing of transcripts may have various effects on splicing, and resultant alternatively spliced isoforms may be either tumor-suppressive or oncogenic. Dysregulated RNA splicing in cancer often causes the release of excess amounts of dsRNA into cytosol, where specific dsRNA sensors provoke antiviral-like responses, including type I interferon signaling. These responses may arrest cell division, causing apoptosis and, externally, stimulate antitumor immunity. Thus, small-molecule spliceosome inhibitors have been shown to facilitate the antiviral-like signaling and are considered to be potential cancer therapies. In turn, a cytoplasmic isoform of ADAR can deaminate dsRNA in cytosol, thereby decreasing its levels and diminishing antitumor innate immunity. We propose that complete or partial inhibition of ADAR may enhance the proapoptotic and cytotoxic effects of splicing inhibitors and that it may be considered a promising addition to cancer therapies targeting RNA splicing.

RNA editing by adenosine deaminases of the ADAR family attracts a growing interest of researchers, both zoologists studying ecological and evolutionary plasticity of invertebrates and medical biochemists focusing on the mechanisms of cancer and other human diseases. These enzymes deaminate adenosine residues in the double-stranded (ds) regions of RNA with the formation of inosine. As a result, some RNAs change their three-dimensional structure and functions. Adenosine-to-inosine editing in the mRNA coding sequences may cause amino acid substitutions in the encoded proteins. Here, we reviewed current concepts on the functions of two active ADAR isoforms identified in mammals (including humans). The ADAR1 protein, which acts non-specifically on extended dsRNA regions, is capable of immunosuppression via inactivation of the dsRNA interactions with specific sensors inducing the cell immunity. Expression of a specific ADAR1 splicing variant is regulated by the type I interferons by the negative feedback mechanism. It was shown that immunosuppressing effects of ADAR1 facilitate progression of some types of cancer. On the other hand, changes in the amino acid sequences resulting from the mRNA editing by the ADAR enzymes can result in the formation of neoantigens that can activate the antitumor immunity. The ADAR2 isoform acts on RNA more selectively; its function is associated with the editing of mRNA coding regions and can lead to the amino acid substitutions, in particular, those essential for the proper functioning of some neurotransmitter receptors in the central nervous system.

Post-transcriptional RNA editing by RNA specific adenosine deaminases (ADAR) was discovered more than two decades ago. It provides additional regulation of animal and human transcriptome. In most cases, it occurs in nervous tissue, where, as a result of the reaction, adenosine is converted to inosine in particular sites of RNA. In case of messenger RNA, during translation, inosine is recognized as guanine leading to amino acid substitutions. Those substitutions are shown to affect substantially the function of proteins, e.g. subunits of the glutamate receptor. Nevertheless, most of the works on RNA editing use analysis of nucleic acids, even those which deal with a coding RNA. In this review, we propose the use of shotgun proteomics based on high resolution liquid chromatography and mass spectrometry for investigation of the effects of RNA editing at the protein level. Recently developed methods of big data processing allow combining the results of various omics techniques, being referred to as proteogenomics. The proposed proteogenomic approach for the analysis of RNA editing at the protein level will directly conduct a qualitative and quantitative analysis of protein edited sequences in the scale of whole proteome.