Frontal theta oscillations during emotion regulation in people with borderline personality disorder.

BACKGROUND: Borderline personality disorder (BPD) is a severe psychiatric disorder conceptualised as a disorder of emotion regulation. Emotion regulation has been linked to a frontolimbic network comprising the dorsolateral prefrontal cortex and the amygdala, which apparently synchronises its activity via oscillatory coupling in the theta frequency range. AIMS: To analyse whether there are distinct differences in theta oscillatory coupling in frontal brain regions between individuals with BPD and matched controls during emotion regulation by cognitive reappraisal. METHOD: Electroencephalogram (EEG) recordings were performed in 25 women diagnosed with BPD and 25 matched controls during a cognitive reappraisal task in which participants were instructed to downregulate negative emotions evoked by aversive visual stimuli. Between- and within-group time-frequency analyses were conducted to analyse regulation-associated theta activity (3.5-8.5 Hz). RESULTS: Oscillatory theta activity differed between the participants with BPD and matched controls during cognitive reappraisal. Regulation-associated theta increases were lower in frontal regions in the BPD cohort compared with matched controls. Functional connectivity analysis for regulation-associated changes in the theta frequency band revealed a lower multivariate interaction measure (MIM) increase in frontal brain regions in persons with BPD compared with matched controls. CONCLUSIONS: Our findings support the notion of alterations in a frontal theta network in BPD, which may be underlying core symptoms of the disorder such as deficits in emotion regulation. The results add to the growing body of evidence for altered oscillatory brain dynamics in psychiatric populations, which might be investigated as individualised treatment targets using non-invasive stimulation methods.

rTMS in mental health disorders.

Transcranial magnetic stimulation (TMS) is an innovative and non-invasive technique used in the diagnosis and treatment of psychiatric and neurological disorders. Repetitive TMS (rTMS) can modulate neuronal activity, neuroplasticity and arousal of the waking and sleeping brain, and, more generally, overall mental health. Numerous studies have examined the predictors of the efficacy of rTMS on clinical outcome variables in various psychiatric disorders. These predictors often encompass the stimulated brain region's location, electroencephalogram (EEG) activity patterns, potential morphological and neurophysiological anomalies, and individual patient's response to treatment. Most commonly, rTMS is used in awake patients with depression, catatonia, and tinnitus. Interestingly, rTMS has also shown promise in inducing slow-wave oscillations in insomnia patients, opening avenues for future research into the potential beneficial effects of these oscillations on reports of non-restorative sleep. Furthermore, neurophysiological measures emerge as potential, disease-specific biomarkers, aiding in predicting treatment response and monitoring post-treatment changes. The study posits the convergence of neurophysiological biomarkers and individually tailored rTMS treatments as a gateway to a new era in psychiatric care. The potential of rTMS to induce slow-wave activity also surfaces as a significant contribution to personalized treatment approaches. Further investigations are called for to validate the imaging and electrophysiological biomarkers associated with rTMS. In conclusion, the potential for rTMS to significantly redefine treatment strategies through personalized approaches could enhance the outcomes in neuropsychiatric disorders.

Depletion of METTL3 alters cellular and extracellular levels of miRNAs containing m6A consensus sequences.

Extracellular vesicles (EVs) are capable of transferring cargo from donor to recipient cells, but precisely how cargo content is regulated for export is mostly unknown. For miRNA cargo, we previously showed that when compared to isogenic colorectal cancer (CRC) cells expressing wild-type KRAS, a distinct subset of miRNAs are differentially enriched in EVs from KRAS mutant active CRC cells, with miR-100 being one of the most enriched. The mechanisms that could explain how miR-100 and other miRNAs are differentially exported into EVs have not been fully elucidated. Here, we tested the effect of N6-methyladenosine (m6A) modification on miRNA export into EVs by depletion of METTL3 and ALKBH5, a writer and eraser of m6A modification, respectively. While the effects of ALKBH5 knockdown were quite modest, decreased levels of METTL3 led to reduced cellular and extracellular levels of a subset of miRNAs that contain consensus sequences for m6A modification. Functional testing of EVs prepared from cells expressing shRNAs against METTL3 showed that they were less capable of conferring colony growth in 3D to wild-type KRAS cells and were also largely incapable of conferring the spread of cetuximab resistance. Our data support a role for METTL3 modification on cellular miRNA levels and export of specific miRNAs.

Analysis of the epitranscriptome with ion-pairing reagent free oligonucleotide mass spectrometry.

RNA modifications gain growing attention as a new frontier in the life sciences but with the rise of RNA vaccines also in biomedical drug development. Impeccable characterization of RNA modifications within their sequence context remains an analytical challenge. Oligonucleotide mass spectrometry (ON-MS), an approach similar to bottom-up proteome analysis, is capable of defining a short 5-15 nucleotide sequence context of an RNA modification while delivering information on the chemical character of the modified nucleotide. Commonly, ON-MS requires the use of ion pairing reagents for ON separation which is not compatible with most other MS-based applications and only few laboratories run dedicated MS instruments for the task. Here, we present an ON-MS technique which is independent of ion pairing reagents and can be used on any available mass spectrometer without risking its sensitivity for other analytes. In this chapter, we describe the experiments necessary for ON-MS method development, ON-MS application to native and synthetic RNAs and finally a guideline for data analysis.

["Coronasomnia"-promoting resilience through insomnia treatment]. / "Coronasomnia" ­ Resilienzförderung durch Insomniebehandlung.

Background: The term "coronasomnia" is used in popular science to describe sleep disorders associated with the COVID-19 pandemic. These disorders may also affect part of the population in the aftermath of the pandemic. Early scientific evidence suggests that COVID-19-associated insomnia and insomniac symptoms can become chronic and will continue to preoccupy the sleep medicine community even after the pandemic has ended. Methods: A literature review was conducted in Medline and Google Scholar using the following combination of keywords: "insomnia and COVID-19", "insomnia and long COVID", "insomnia, PTSD and COVID-19", and "fatigue and insomnia in long COVID". In addition, the authors reviewed several recent articles published by members of the European Insomnia Network. Results: Studies on insomnia and COVID-19 show significant associations between acute infection and insomnia in affected individuals. The prevalence of insomnia symptoms in COVID-19-affected individuals was 36 to 88%, which is significantly higher than the estimated 10 to 40% prevalence of insomnia in the general population. Conclusion: Digital therapy as a current treatment option for insomnia can be offered to patients regardless of physical distance. Accordingly, not only early approval of therapy apps, but also person-led, digital therapy options for insomnia would be recommended. The inclusion of personalised and sleep-coaching measures in the area of occupational health management is encouraged.

Strategies to Avoid Artifacts in Mass Spectrometry-Based Epitranscriptome Analyses.

In this report, we perform structure validation of recently reported RNA phosphorothioate (PT) modifications, a new set of epitranscriptome marks found in bacteria and eukaryotes including humans. By comparing synthetic PT-containing diribonucleotides with native species in RNA hydrolysates by high-resolution mass spectrometry (MS), metabolic stable isotope labeling, and PT-specific iodine-desulfurization, we disprove the existence of PTs in RNA from E. coli, S. cerevisiae, human cell lines, and mouse brain. Furthermore, we discuss how an MS artifact led to the initial misidentification of 2'-O-methylated diribonucleotides as RNA phosphorothioates. To aid structure validation of new nucleic acid modifications, we present a detailed guideline for MS analysis of RNA hydrolysates, emphasizing how the chosen RNA hydrolysis protocol can be a decisive factor in discovering and quantifying RNA modifications in biological samples.

Sleep quality and COVID-19 outcomes: the evidence-based lessons in the framework of predictive, preventive and personalised (3P) medicine.

Sleep quality and duration play a pivotal role in maintaining physical and mental health. In turn, sleep shortage, deprivation and disorders are per evidence the risk factors and facilitators of a broad spectrum of disorders, amongst others including depression, stroke, chronic inflammation, cancers, immune defence insufficiency and individual predisposition to infection diseases with poor outcomes, for example, related to the COVID-19 pandemic. Keeping in mind that COVID-19-related global infection distribution is neither the first nor the last pandemic severely affecting societies around the globe to the costs of human lives accompanied with enormous economic burden, lessons by predictive, preventive and personalised (3P) medical approach are essential to learn and to follow being better prepared to defend against global pandemics. To this end, under extreme conditions such as the current COVID-19 pandemic, the reciprocal interrelationship between the sleep quality and individual outcomes becomes evident, namely, at the levels of disease predisposition, severe versus mild disease progression, development of disease complications, poor outcomes and related mortality for both - population and healthcare givers. The latter is the prominent example clearly demonstrating the causality of severe outcomes, when the long-lasting work overload and shift work rhythm evidently lead to the sleep shortage and/or deprivation that in turn causes immune response insufficiency and strong predisposition to the acute infection with complications. This article highlights and provides an in-depth analysis of the concerted risk factors related to the sleep disturbances under the COVID-19 pandemic followed by the evidence-based recommendations in the framework of predictive, preventive and personalised medical approach.

Quantification of Modified Nucleosides in the Context of NAIL-MS.

Recent progress in epitranscriptome research shows an interplay of enzymes modifying RNAs and enzymes dedicated for RNA modification removal. One of the main techniques to study RNA modifications is liquid chromatography-coupled tandem mass spectrometry (LC-MS/MS) as it allows sensitive detection of modified nucleosides. Although RNA modifications have been found to be highly dynamic, state-of-the-art LC-MS/MS analysis only gives a static view on modifications and does not allow the investigation of temporal modification placement. Here, we present the principles of nucleic acid isotope labeling coupled with mass spectrometry, termed NAIL-MS, which overcomes these limitations by stable isotope labeling in human cell culture and gives detailed instructions on how to label cells and process samples in order to get reliable results. For absolute quantification in the context of NAIL-MS, we explain the production of internal standards in detail. Furthermore, we outline the requirements for stable isotope labeling in cell culture and all subsequent steps to receive nucleoside mixtures of native RNA for NAIL-MS analysis. In the final section of this chapter, we describe the distinctive features of NAIL-MS data analysis with a special focus toward absolute quantification of modified nucleosides.

Insomnia Associated with Tinnitus and Gender Differences.

Chronic tinnitus causes a decrease in well-being and can negatively affect sleep quality. It has further been indicated that there are clinically relevant gender differences, which may also have an impact on sleep quality. By conducting a retrospective and explorative data analysis for differences in patients with tinnitus and patients diagnosed with tinnitus and insomnia, hypothesized differences were explored in the summed test scores and on item-level of the validated psychometric instruments. A cross-sectional study was conducted collecting data from a sample of tinnitus patients (n = 76). Insomnia was diagnosed in 49 patients. Gender differences were found on aggregated test scores of the MADRS and BDI with men scoring higher than women, indicating higher depressive symptoms in men. Women stated to suffer more from headaches (p < 0.003), neck pain (p < 0.006) and nervousness as well as restlessness (p < 0.02). Women also reported an increase in tinnitus loudness in response to stress compared to men (p < 0.03). Male individuals with tinnitus and insomnia have higher depression scores and more clinically relevant depressive symptoms than women, who suffer more from psychosomatic symptoms. The results indicate a need for a targeted therapy of depressive symptoms in male patients and targeted treatment of psychosomatic symptoms, stress-related worsening of insomnia and tinnitus in women.

m5U54 tRNA Hypomodification by Lack of TRMT2A Drives the Generation of tRNA-Derived Small RNAs.

Transfer RNA (tRNA) molecules contain various post-transcriptional modifications that are crucial for tRNA stability, translation efficiency, and fidelity. Besides their canonical roles in translation, tRNAs also originate tRNA-derived small RNAs (tsRNAs), a class of small non-coding RNAs with regulatory functions ranging from translation regulation to gene expression control and cellular stress response. Recent evidence indicates that tsRNAs are also modified, however, the impact of tRNA epitranscriptome deregulation on tsRNAs generation is only now beginning to be uncovered. The 5-methyluridine (m5U) modification at position 54 of cytosolic tRNAs is one of the most common and conserved tRNA modifications among species. The tRNA methyltransferase TRMT2A catalyzes this modification, but its biological role remains mostly unexplored. Here, we show that TRMT2A knockdown in human cells induces m5U54 tRNA hypomodification and tsRNA formation. More specifically, m5U54 hypomodification is followed by overexpression of the ribonuclease angiogenin (ANG) that cleaves tRNAs near the anticodon, resulting in accumulation of 5'tRNA-derived stress-induced RNAs (5'tiRNAs), namely 5'tiRNA-GlyGCC and 5'tiRNA-GluCTC, among others. Additionally, transcriptomic analysis confirms that down-regulation of TRMT2A and consequently m5U54 hypomodification impacts the cellular stress response and RNA stability, which is often correlated with tiRNA generation. Accordingly, exposure to oxidative stress conditions induces TRMT2A down-regulation and tiRNA formation in mammalian cells. These results establish a link between tRNA hypomethylation and ANG-dependent tsRNAs formation and unravel m5U54 as a tRNA cleavage protective mark.

Instrumental analysis of RNA modifications.

Organisms from all domains of life invest a substantial amount of energy for the introduction of RNA modifications into nearly all transcripts studied to date. Instrumental analysis of RNA can focus on the modified residues and reveal the function of these epitranscriptomic marks. Here, we will review recent advances and breakthroughs achieved by NMR spectroscopy, sequencing, and mass spectrometry of the epitranscriptome.

Cell culture NAIL-MS allows insight into human tRNA and rRNA modification dynamics in vivo.

Recently, studies about RNA modification dynamics in human RNAs are among the most controversially discussed. As a main reason, we identified the unavailability of a technique which allows the investigation of the temporal processing of RNA transcripts. Here, we present nucleic acid isotope labeling coupled mass spectrometry (NAIL-MS) for efficient, monoisotopic stable isotope labeling in both RNA and DNA in standard cell culture. We design pulse chase experiments and study the temporal placement of modified nucleosides in tRNAPhe and 18S rRNA. In existing RNAs, we observe a time-dependent constant loss of modified nucleosides which is masked by post-transcriptional methylation mechanisms and thus undetectable without NAIL-MS. During alkylation stress, NAIL-MS reveals an adaptation of tRNA modifications in new transcripts but not existing ones. Overall, we present a fast and reliable stable isotope labeling strategy which allows in-depth study of RNA modification dynamics in human cell culture.

Chemical Amination/Imination of Carbonothiolated Nucleosides During RNA Hydrolysis.

Liquid chromatography-tandem mass spectrometry (LC-MS/MS) has become the gold-standard technique to study RNA and its various modifications. While most research on RNA nucleosides has been focused on their biological roles, discovery of new modifications remains of interest. With state-of-the-art technology, the presence of artifacts can confound the identification of new modifications. Here, we report the characterization of a non-natural mcm5 isoC ribonucleoside in S. cerevisiae total tRNA hydrolysate by higher-energy collisional dissociation (HCD)-based fingerprints and isotope labeling of RNA. Its discovery revealed a class of amino/imino ribonucleoside artifacts that are generated during RNA hydrolysis under ammonium-buffered mild basic conditions. We then identified digestion conditions that can reduce or eliminate their formation. These finding and method enhancements will improve the accurate detection of new RNA modifications.

Sequence- and structure-specific cytosine-5 mRNA methylation by NSUN6.

The highly abundant N6-methyladenosine (m6A) RNA modification affects most aspects of mRNA function, yet the precise function of the rarer 5-methylcytidine (m5C) remains largely unknown. Here, we map m5C in the human transcriptome using methylation-dependent individual-nucleotide resolution cross-linking and immunoprecipitation (miCLIP) combined with RNA bisulfite sequencing. We identify NSUN6 as a methyltransferase with strong substrate specificity towards mRNA. NSUN6 primarily targeted three prime untranslated regions (3'UTR) at the consensus sequence motif CTCCA, located in loops of hairpin structures. Knockout and rescue experiments revealed enhanced mRNA and translation levels when NSUN6-targeted mRNAs were methylated. Ribosome profiling further demonstrated that NSUN6-specific methylation correlated with translation termination. While NSUN6 was dispensable for mouse embryonic development, it was down-regulated in human tumours and high expression of NSUN6 indicated better patient outcome of certain cancer types. In summary, our study identifies NSUN6 as a methyltransferase targeting mRNA, potentially as part of a quality control mechanism involved in translation termination fidelity.

tRNA-modifying enzyme mutations induce codon-specific mistranslation and protein aggregation in yeast.

Protein synthesis rate and accuracy are tightly controlled by the cell and are essential for proteome homoeostasis (proteostasis); however, the full picture of how mRNA translational factors maintain protein synthesis accuracy and co-translational protein folding are far from being fully understood. To address this question, we evaluated the role of 70 yeast tRNA-modifying enzyme genes on protein aggregation and used mass spectrometry to identify the aggregated proteins. We show that modification of uridine at anticodon position 34 (U34) by the tRNA-modifying enzymes Elp1, Elp3, Sml3 and Trm9 is critical for proteostasis, the mitochondrial tRNA-modifying enzyme Slm3 plays a fundamental role in general proteostasis and that stress response proteins whose genes are enriched in codons decoded by tRNAs lacking mcm5U34, mcm5s2U34, ncm5U34, ncm5Um34, modifications are overrepresented in protein aggregates of the ELP1, SLM3 and TRM9 KO strains. Increased rates of amino acid misincorporation were also detected in these strains at protein sites that specifically mapped to the codons sites that are decoded by the hypomodified tRNAs, demonstrating that U34 tRNA modifications safeguard the proteome from translational errors, protein misfolding and proteotoxic stress.

METTL6 is a tRNA m3C methyltransferase that regulates pluripotency and tumor cell growth.

Recently, covalent modifications of RNA, such as methylation, have emerged as key regulators of all aspects of RNA biology and have been implicated in numerous diseases, for instance, cancer. Here, we undertook a combination of in vitro and in vivo screens to test 78 potential methyltransferases for their roles in hepatocellular carcinoma (HCC) cell proliferation. We identified methyltransferase-like protein 6 (METTL6) as a crucial regulator of tumor cell growth. We show that METTL6 is a bona fide transfer RNA (tRNA) methyltransferase, catalyzing the formation of 3-methylcytidine at C32 of specific serine tRNA isoacceptors. Deletion of Mettl6 in mouse stem cells results in changes in ribosome occupancy and RNA levels, as well as impaired pluripotency. In mice, Mettl6 knockout results in reduced energy expenditure. We reveal a previously unknown pathway in the maintenance of translation efficiency with a role in maintaining stem cell self-renewal, as well as impacting tumor cell growth profoundly.

Identification and rescue of a tRNA wobble inosine deficiency causing intellectual disability disorder.

The deamination of adenosine to inosine at the wobble position of tRNA is an essential post-transcriptional RNA modification required for wobble decoding in bacteria and eukaryotes. In humans, the wobble inosine modification is catalyzed by the heterodimeric ADAT2/3 complex. Here, we describe novel pathogenic ADAT3 variants impairing adenosine deaminase activity through a distinct mechanism that can be corrected through expression of the heterodimeric ADAT2 subunit. The variants were identified in a family in which all three siblings exhibit intellectual disability linked to biallelic variants in the ADAT3 locus. The biallelic ADAT3 variants result in a missense variant converting alanine to valine at a conserved residue or the introduction of a premature stop codon in the deaminase domain. Fibroblast cells derived from two ID-affected individuals exhibit a reduction in tRNA wobble inosine levels and severely diminished adenosine tRNA deaminase activity. Notably, the ADAT3 variants exhibit impaired interaction with the ADAT2 subunit and alterations in ADAT2-dependent nuclear localization. Based upon these findings, we find that tRNA adenosine deaminase activity and wobble inosine modification can be rescued in patient cells by overexpression of the ADAT2 catalytic subunit. These results uncover a key role for the inactive ADAT3 deaminase domain in proper assembly with ADAT2 and demonstrate that ADAT2/3 nuclear import is required for maintaining proper levels of the wobble inosine modification in tRNA.

Eukaryotic life without tQCUG: the role of Elongator-dependent tRNA modifications in Dictyostelium discoideum.

In the Elongator-dependent modification pathway, chemical modifications are introduced at the wobble uridines at position 34 in transfer RNAs (tRNAs), which serve to optimize codon translation rates. Here, we show that this three-step modification pathway exists in Dictyostelium discoideum, model of the evolutionary superfamily Amoebozoa. Not only are previously established modifications observable by mass spectrometry in strains with the most conserved genes of each step deleted, but also additional modifications are detected, indicating a certain plasticity of the pathway in the amoeba. Unlike described for yeast, D. discoideum allows for an unconditional deletion of the single tQCUG gene, as long as the Elongator-dependent modification pathway is intact. In gene deletion strains of the modification pathway, protein amounts are significantly reduced as shown by flow cytometry and Western blotting, using strains expressing different glutamine leader constructs fused to GFP. Most dramatic are these effects, when the tQCUG gene is deleted, or Elp3, the catalytic component of the Elongator complex is missing. In addition, Elp3 is the most strongly conserved protein of the modification pathway, as our phylogenetic analysis reveals. The implications of this observation are discussed with respect to the evolutionary age of the components acting in the Elongator-dependent modification pathway.

Sleep disorders in migrants and refugees: a systematic review with implications for personalized medical approach.

BACKGROUND: Sleep disorders are very common in migrants and refugees, often as a comorbid disorder to different somatic or psychiatric diagnoses and psychological disturbances such as metabolic syndrome, post-traumatic stress disorder, depression, and anxiety disorders. OBJECTIVES: To review published prevalence rates as well as possible predictors for sleep disturbances in these vulnerable groups, including pre-migration stress, acculturation, and trauma before, during, and after migration, integration, and lifestyle in the host country with implications for predictive, preventive, and personalized medical approach (3PM). DATA SOURCES: Electronic databases PubMed, PsycInfo, and Web of Knowledge were searched using (combined) search terms "migrant," "asylum seeker," "refugee," "sleep disturbances," "sleep disorder," "insomnia," and "sleep wake disorder." STUDY ELIGIBILITY CRITERIA: Peer-reviewed studies from 2000 to 2018 reporting data on prevalence and/or predictors of any measure of sleep disturbance were included. PARTICIPANTS: Studies on international migrants and refugees, as well as internally displaced populations, were included. METHODS: We conducted a systematic review on the topic of sleep disorders in migrant and refugee populations. Only published articles and reviews in peer-reviewed journals were included. RESULTS: We analyzed five studies on sleep disorders in migrants, five studies on adult refugees, and three on refugee children and adolescents. Prevalence of sleep disorders in migrants and refugees ranges between 39 and 99%. In migrant workers, stress related to integration and adaptation to the host society is connected to higher risks of snoring, metabolic diseases, and insomnia. Sleep disturbances in refugees are predicted by past war experience. Sleep difficulties in adult and child refugees are strongly correlated to trauma. Torture of parents and grandparents can predict sleep disorders in refugee children, while being accompanied by parents to the host country has a protective effect on children's sleep. CONCLUSIONS AND IMPLICATIONS: Considering the differences in risk factors, vulnerability, and traumatic life events for different migrant populations, origins of sleep difficulties vary, depending on the migrant populations. Effects on sleep disturbances and sleep quality may be a result of integration in the host country, including changes of lifestyle, such as diet and working hours with implication for OSAS (obstructive sleep apnea) and insomnia. Compared with migrant populations, sleep disturbances in refugee populations are more correlated with mental health symptoms and disorders, especially PTSD (post-traumatic stress disorder), than with psychosocial problems. In juvenile refugee populations, psychological problems and disturbed sleep are associated with traumatic experiences during their journey to the host country. Findings highlight the need for expert recommendations for development of 3P approach stratified in the following: (1) prediction, including structured exploration of predisposing and precipitating factors that may trigger acute insomnia, screening of the according sleep disorders by validated translated questionnaires and sleep diaries, and a face-to-face or virtual setting and screening of OSAS; (2) target prevention by sleep health education for female and male refugees and migrant workers, including shift workers; and (3) personalized medical approach, including translated cognitive behavioral treatment for insomnia (CBT-I) and imagery rehearsal therapy for refugees and telehealth programs for improved CPAP adherence in migrants, with the goal to enable better sleep health quality and improved health economy.

Synthesis and Metabolic Fate of 4-Methylthiouridine in Bacterial tRNA.

Ribonucleic acid (RNA) is central to many life processes and, to fulfill its function, it has a substantial chemical variety in its building blocks. Enzymatic thiolation of uridine introduces 4-thiouridine (s4 U) into many bacterial transfer RNAs (tRNAs), which is used as a sensor for UV radiation. A similar modified nucleoside, 2-thiocytidine, was recently found to be sulfur-methylated especially in bacteria exposed to antibiotics and simple methylating reagents. Herein, we report the synthesis of 4-methylthiouridine (ms4 U) and confirm its presence and additional formation under stress in Escherichia coli. We used the synthetic ms4 U for isotope dilution mass spectrometry and compared its abundance to other reported tRNA damage products. In addition, we applied sophisticated stable-isotope pulse chase studies (NAIL-MS) and showed its AlkB-independent removal in vivo. Our findings reveal the complex nature of bacterial RNA damage repair.