Nuclear Export Research Articles

Allosteric degraders induce CRL5 ASB8 mediated degradation of XPO1.

The nuclear export receptor Exportin 1 (XPO1/CRM1) is often overexpressed in cancer cells resulting in aberrant localization of many cancer-related protein cargoes. The XPO1 inhibitor and cancer drug selinexor (KPT-330), and its analog KPT-185, block XPO1-cargo binding thereby restoring cargo localization. Selinexor binding induces cullin-RING E3 ubiquitin ligase (CRL) substrate receptor ASB8-mediated XPO1 degradation. Here we reveal the mechanism of inhibitor-XPO1 engagement by CRL5ASB8. Cryogenic electron microscopy (cryo-EM) structures show ASB8 binding to a large surface of selinexor/KPT-185-XPO1 that includes a three-dimensional degron unique to the drug-bound exportin. The structure explains weak XPO1-ASB8 binding in the absence of selinexor/KPT-185 that is unproductive for proteasomal degradation, and the substantial affinity increase upon selinexor/KPT-185 conjugation, which results in CRL5 ASB8 -mediated XPO1 ubiquitination. In contrast to previously characterized small molecule degraders, which all act as molecular glues, selinexor/KPT-185 binds extensively to XPO1 but hardly contacts ASB8. Instead, selinexor/KPT-185 binds XPO1 and stabilizes a unique conformation of the NES/inhibitor-binding groove that binds ASB8. Selinexor/KPT-185 is an allosteric degrader. We have explained how drug-induced protein degradation is mediated by a CRL5 system through an allosteric rather than a molecular glue mechanism, expanding the modes of targeted protein degradation beyond the well-known molecular glues of CRL4.

Modeling of mRNA deadenylation rates reveal a complex relationship between mRNA deadenylation and decay.

Complete cytoplasmic polyadenosine tail (polyA-tail) deadenylation is thought to be essential for initiating mRNA decapping and subsequent degradation. To investigate this prevalent model, we conducted direct RNA sequencing of S. cerevisiae mRNAs derived from chase experiments under steady-state and stress condition. Subsequently, we developed a numerical model based on a modified gamma distribution function, which estimated the transcriptomic deadenylation rate at 10 A/min. A simplified independent method, based on the delineation of quantile polyA-tail values, showed a correlation between the decay and deadenylation rates of individual mRNAs, which appeared consistent within functional transcript groups and associated with codon optimality. Notably, these rates varied during the stress response. Detailed analysis of ribosomal protein-coding mRNAs (RPG mRNAs), constituting 40% of the transcriptome, singled out this transcript group. While deadenylation and decay of RPG mRNAs accelerated under heat stress, their degradation could proceed even when deadenylation was blocked, depending entirely on ongoing nuclear export. Our findings support the general primary function of deadenylation in dictating the onset of decapping, while also demonstrating complex relations between these processes.

Cellular NS1-BP protein interacts with the mRNA export receptor NXF1 to mediate nuclear export of influenza virus M mRNAs

Influenza A viruses have eight genomic RNAs that are transcribed in the host cell nucleus. Two of the viral mRNAs undergo alternative splicing. The M1 mRNA encodes the matrix protein 1 (M1) and is also spliced into M2 mRNA, which encodes the proton channel matrix protein 2 (M2). Our previous studies have shown that the cellular NS1-binding protein (NS1-BP) interacts with the viral non-structural protein 1 (NS1) and M1 mRNA to promote M1 to M2 splicing. Another pool of NS1 protein binds the mRNA export receptor NXF1 (nuclear RNA export factor-1), leading to nuclear retention of cellular mRNAs. Here we show a series of biochemical and cell biological findings that suggest a model for nuclear export of M1 and M2 mRNAs despite the mRNA nuclear export inhibition imposed by the viral NS1 protein. NS1-BP competes with NS1 for NXF1 binding, allowing the recruitment of NXF1 to the M mRNAs after splicing. NXF1 then binds GANP (Germinal-center Associated Nuclear Protein), a member of the TRanscription and EXport complex (TREX)-2. Although both NS1 and NS1-BP remain in complex with GANP-NXF1, they dissociate once this complex docks at the nuclear pore complex (NPC), and the M mRNAs are translocated to the cytoplasm. Since this mRNA nuclear export pathway is key for expression of M1 and M2 proteins that function in viral intracellular trafficking and budding, these viral-host interactions are critical for influenza virus replication.

Siwu tablet attenuates high fructose-induced glomerular podocyte senescence in rats through increasing Nup155 to promote INO80 mRNA nuclear export

Ethnopharmacological relevanceSiwu tablet (SWT), derived from a traditional Chinese medicinal formula named Siwu decoction, is widely used for blood deficiency syndrome. Siwu decoction and its derived formulas have been proven to improve renal anemia and prevent senescence. Whether SWT prevents glomerular podocyte senescence and the underlying molecular mechanism remains unknow. Aim of the studyTo elucidate the protective effect and possible mechanism of SWT on glomerular podocyte senescence. Material and methodsLiquid chromatography-tandem mass spectrometry (LC-MS/MS) was performed to characterize components of SWT. Male Sprague-Dawley rats were given 10% fructose drinking water for 16 weeks. SWT (810 and 1620 mg/kg) was administered orally for the last 8 weeks. The assays of senescence-associated beta-galactosidase (SA-β-gal) staining, immunohistochemistry, quantitative real-time polymerase chain reaction (qRT-PCR), and Western blot as well as enzyme linked immunosorbent assay were performed to evaluate rat glomerular podocyte senescence. The mRNA and protein levels of nucleoporin 155 (Nup155) and inositol requiring mutant 80 (INO80) in rat glomeruli were detected by qRT-PCR, Western blot and immunofluorescence. Foot processes and nuclear pore complexes (NPCs) of rat glomerular podocytes were visualized by transmission electron microscopy. ResultsOne hundred and fifty-nine components were preliminarily identified in SWT. The results of animal experiments showed that SWT decreased the activity of SA-β-gal, protein levels of p16, p21, p53 and phosphorylated histone H2AX (γ-H2AX), and mRNA levels of interleukin-1β (IL-1β), IL-6 and tumor necrosis factor-α (TNF-α) in glomeruli of high fructose-fed rats. As expected, SWT increased renal cortex erythropoietin mRNA expression and serum erythropoietin concentration in this animal model. SWT reduced urine albumin-to-creatinine ratio and serum levels of uric acid, creatinine and blood urea nitrogen, and recovered glomerular structure injury in high fructose-fed rats. It up-regulated mRNA and protein levels of Nup155 and the number of podocyte NPCs, and subsequently reinforced mRNA nuclear export and protein expression of INO80 in rat glomeruli under high fructose stimulation. ConclusionsSWT ameliorates glomerular podocyte senescence in high fructose-fed rats possibly by increasing Nup155 to promote INO80 mRNA nuclear export.

RNA surveillance by the RNA helicase MTR4 determines volume of mouse oocytes.

Oocytes are the largest cell type in multicellular animals. Here, we show that mRNA transporter 4 (MTR4) is indispensable for oocyte growth and functions as part of the RNA surveillance mechanism, which is responsible for nuclear waste RNA clearance. MTR4 ensures the normal post-transcriptional processing of maternal RNAs, their nuclear export to the cytoplasm, and the accumulation of properly processed transcripts. Oocytes with Mtr4 knockout fail to accumulate sufficient and normal transcripts in the cytoplasm and cannot grow to normal sizes. MTR4-dependent RNA surveillance has a previously unrecognized function in maintaining a stable nuclear environment for the establishment of non-canonical histone H3 lysine-4 trimethylation and chromatin reorganization, which is necessary to form a nucleolus-like structure in oocytes. In conclusion, MTR4-dependent RNA surveillance activity is a checkpoint that allows oocytes to grow to a normal size, undergo nuclear and cytoplasmic maturation, and acquire developmental competence.

HMGB2-induced calreticulin translocation required for immunogenic cell death and ferroptosis of cancer cells are controlled by the nuclear exporter XPO1

Cisplatin and oxaliplatin cause the secretion of high mobility group box 1 (HMGB1) protein from cancer cells, which is necessary for initiation of immunogenic cell death (ICD). Calreticulin (CRT) translocation from the endoplasmic reticulum to the plasma membrane is also required; oxaliplatin induces this translocation but cisplatin does not. We have discovered that oxaliplatin causes the secretion of both HMGB1 and HMGB2 from the cell nucleus into the extracellular milieu. We previously showed that cisplatin-mediated secretion of HMGB1 is controlled by the nuclear exporter XPO1 (chromosomal maintenance 1; CRM1). We now find that XPO1 regulates oxaliplatin-mediated secretion of both HMGB1 and HMGB2. XPO1 inhibition causes nuclear accumulation of both proteins, inhibition of oxaliplatin-mediated ferroptosis of colon cancer cells, and inhibition of CRT translocation to the plasma membrane of lung and colon cancer cells. Incubation of cancer cells with cell targeted (CT)-HMGB2 confirmed that HMGB2 is required for the CRT translocation. Furthermore, CT-HMGB2 is three orders of magnitude more potent at inducing CRT translocation than oxaliplatin.

RNA 5-Methylcytosine Modification: Regulatory Molecules, Biological Functions, and Human Diseases.

RNA methylation modifications influence gene expression, and disruptions of these processes are often associated with various human diseases. The common RNA methylation modification 5-methylcytosine (m5C), which is dynamically regulated by writers, erasers, and readers, widely occurs in transfer RNAs (tRNAs), messenger RNAs (mRNAs), ribosomal RNAs (rRNAs), enhancer RNAs (eRNAs), and other non-coding RNAs (ncRNAs). RNA m5C modification regulates metabolism, stability, nuclear export, and translation of RNA molecules. An increasing number of studies have revealed the critical roles of the m5C RNA modification and its regulators in the development, diagnosis, prognosis, and treatment of various human diseases. In this review, we summarized the recent studies on RNA m5C modification and discussed the advances in its detection methodologies, distribution, and regulators. Furthermore, we addressed the significance of RNAs modified with m5C marks in essential biological processes as well as in the development of various human disorders, from neurological diseases to cancers. This review provides a new perspective on the diagnosis, treatment, and monitoring of human diseases by elucidating the complex regulatory network of the epigenetic m5C modification.

NSUN4-mediated m5C modification of circERI3 promotes lung cancer development by altering mitochondrial energy metabolism

As a highly important methylation modification, the 5-methyladenosine (m5C) modification can profoundly affect RNAs by regulating their transcription, structure and stability. With the continuous development of high-throughput technology, differentially expressed circular RNAs (circRNAs) have been increasingly discovered, and circRNAs play unique roles in tumorigenesis and development. However, the regulatory mechanism of the m5C modification of circRNAs has not yet been revealed. In this study, circERI3, which is highly expressed in lung cancer tissue and significantly correlated with the clinical progression of lung cancer, was initially identified through differential expression profiling of circRNAs. A combined m5C microarray analysis revealed that circERI3 contains the m5C modification and that the NSUN4-mediated m5C modification of circERI3 can increase its nuclear export. The important function of circERI3 in promoting lung cancer progression in vitro and in vivo was clarified. Moreover, we elucidated the novel mechanism by which circERI3 targets DNA binding protein 1 (DDB1), regulates its ubiquitination, enhances its stability, and in turn promotes the transcription of peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α) through DDB1 to affect mitochondrial function and energy metabolism, which ultimately promotes the development of lung cancer. This study not only revealed the reasons for the abnormal distribution of circERI3 in lung cancer tissues from the perspective of methylation and clarified the important role of circERI3 in lung cancer progression but also described a novel mechanism by which circERI3 promotes lung cancer development through mitochondrial energy metabolism, providing new insights for the study of circRNAs in lung cancer.

Translocon Subunits of the COP9 Signalosome Complex Are a Central Hub for Regulating Multiple Photoresponsive Processes and Autophagic Flux in Magnaporthe oryzae.

Photodependent processes, including circadian rhythm, autophagy, ubiquitination, neddylation/deneddylation, and metabolite biosynthesis, profoundly influence microbial pathogenesis. Although a photomorphogenesis signalosome (COP9/CSN) has been identified, the mechanism by which this large complex contributes to the pathophysiological processes in filamentous fungi remains unclear. Here, we identified eight CSN complex subunits in the rice blast fungus Magnaporthe oryzae and functionally characterized the translocon subunits containing a nuclear export or localization signal (NES/NLS). Targeted gene replacement of these CSN subunits, including MoCSN3, MoCSN5, MoCSN6, MoCSN7, and MoCSN12, attenuated vegetative growth and conidiation and rendered the deletion strains nonpathogenic. MoCSN7 deletion significantly suppressed arachidonic acid catabolism, and compromised cell wall integrity in M. oryzae. Surprisingly, we also discovered that MoCSN subunits, particularly MoCsn7, are required for the cAMP-dependent regulation of autophagic flux. Therefore, MoCSN significantly contributes to morphological, physiological, and pathogenic differentiation in M. oryzae by fostering cross-talk between multiple pathways.

ITSN1 binds the E2-conjugating enzyme UBC9

Aim. Scaffolding protein of the intersectin 1 (ITSN1) associated with malignant cell transformation. A short isoform of ITSN1 (ITSN1-S) can localize to the nucleus and inhibit breast cancer cell proliferation but the exact mechanisms of ITSN1 nuclear export have not been fully elucidated. SUMOylation of ITSN1, or its interaction with components of SUMO modification, may be one of the regulatory mechanisms contributing to the nuclear-cytoplasmic shuffle of ITSN1 in the cell. Methods. Full-length human UBC9 sequence was subcloned in pGEX4T2 vector for in vitro GST-binding assays with overexpressed Omni-ITSN1-S in 293 cell line. Lysates of 293 cells with overexpressed FLAG-UBC9 were used for co-immunoprecipitation with endogenous proteins of ITSN1 and ITSN2. Results. Endogenous ITSN1-S form complexes with full-length overexpressed UBC9 in 293 in vivo. Further analysis revealed that GST-UBC9 binds human full-length short isoform ITSN1-S in vitro. Conclusions. E2-conjugating enzyme of the SUMOylation, UBC9, is confirmed as a novel protein partner for ITSN1 both in vitro and in vivo. Considering the tumor suppressor role of a nuclear ITSN1-S in breast cancer and the unique role UBC9 plays in SUMO-modification of proteins, we suggest a possibility of UBC9 and ITSN1 interaction association with malignant transformation, which can be the ground for the further studies.

The HIV-1 Nuclear Export Complex Reveals the Role of RNA in Crm1 Cargo Recognition.

Crm1 is a highly conserved nuclear exportin that transports >1000 human proteins including ribonucleoprotein (RNP) complexes. The interface between Crm1 and RNP cargos is unknown. The HIV regulatory protein, Rev, was one of the first identified cargos for Crm1 and contains a prototypic nuclear export sequence (NES). We present the cryo-electron microscopy structure of the HIV-1 nuclear export complex (Crm1/Ran-GTP and the Rev/RRE RNP). Rev binds at a previously unseen protein-protein binding site that stabilizes a unique Crm1 dimer and positions two NESs within the Crm1 dimer. The orientation of Rev binding positions the RRE within a charged pocket on the inside of the Crm1 toroid, mediating direct RNA-Ran-GTP contacts, highlighting the significant role of the RRE in the interaction. Structure based mutations, combined with cell-based assays, show that Crm1 has multiple distinct cargo recognition sites and explains how Crm1 can recognize a diverse range of protein and RNP cargos.

Enhanced Extracellular Vesicle Cargo Loading via microRNA Biogenesis Pathway Modulation.

Extracellular vesicles (EVs) are physiological vectors for the intercellular transport of a variety of molecules. Among these, small RNAs, and especially microRNAs (miRNAs), have been identified as prevalent components, and there has thus been a robust investigation of EVs for therapeutic miRNAs delivery. However, intrinsic levels of EV-associated miRNAs are generally too low to enable efficient and effective therapeutic outcomes. We hypothesized that miRNA localization to EVs could be improved by limiting competing interactions that occur throughout the miRNA biogenesis process. Using miR-146a-5p as a model, modulation of transcription, nuclear export, and enzymatic cleavage steps of miRNA biogenesis were tested for impact on EV miRNA loading. Working in HEK293T cells, various alterations in the EV biogenesis pathway were shown to impact miRNA localization to EVs. The system was then applied in induced pluripotent stem cells (iPSCs), a more promising substrate for therapeutic EV production, and EVs were separated and assessed for anti-inflammatory efficacy in vitro and in a murine colitis model, where the preservation of function was validated. Overall, the results highlight necessary considerations when designing a cell culture system for the devoted production of miRNA-loaded EVs.

Fluorescent tagging of endogenous IRS2 with an auxin-dependent degron to assess dynamic intracellular localization and function

Insulin Receptor Substrate 2 (IRS2) is a signaling adaptor protein for the insulin (IR) and Insulin-like Growth Factor-1 (IGF-1R) receptors. In breast cancer, IRS2 contributes to both initiation of primary tumor growth and establishment of secondary metastases through regulation of cancer stem cell (CSC) function and invasion. However, how IRS2 mediates its diverse functions is not well understood. We used CRISPR/Cas9-mediated gene editing to modify endogenous IRS2 to study the expression, localization, and function of this adaptor protein. A cassette containing an auxin inducible degradation (AID) sequence, 3X-FLAG tag and mNeon-green was introduced at the N-terminus of the IRS2 gene to provide rapid and reversible control of IRS2 protein degradation and analysis of endogenous IRS2 expression and localization. Live fluorescence imaging of these cells revealed that IRS2 shuttles between the cytoplasm and nucleus in response to growth regulatory signals in a PI3K-dependent manner. Inhibition of nuclear export or deletion of a putative nuclear export sequence in the C-terminal tail promotes nuclear retention of IRS2, implicating nuclear export in the mechanism by which IRS2 intracellular localization is regulated. Moreover, the acute induction of IRS2 degradation reduces tumor cell invasion, demonstrating the potential for therapeutic targeting of this adaptor protein. Our data highlight the value of our model of endogenously tagged IRS2 as a tool to study IRS2 localization and function.

What makes Candida auris pan-drug resistant? Integrative insights from genomic, transcriptomic, and phenomic analysis of clinical strains resistant to all four major classes of antifungal drugs.

The global epidemic of drug-resistant Candida auris continues unabated. The initial report on pan-drug resistant (PDR) C. auris strains in a hospitalized patient in New York was unprecedented. PDR C. auris showed both known and unique mutations in the prominent gene targets of azoles, amphotericin B, echinocandins, and flucytosine. However, the factors that allow C. auris to acquire pan-drug resistance are not known. Therefore, we conducted a genomic, transcriptomic, and phenomic analysis to better understand PDR C. auris. Among 1,570 genetic variants in drug-resistant C. auris, 299 were unique to PDR strains. The whole-genome sequencing results suggested perturbations in genes associated with nucleotide biosynthesis, mRNA processing, and nuclear export of mRNA. Whole transcriptome sequencing of PDR C. auris revealed two genes to be significantly differentially expressed-a DNA repair protein and DNA replication-dependent chromatin assembly factor 1. Of 59 novel transcripts, 12 transcripts had no known homology. We observed no fitness defects among multi-drug resistant (MDR) and PDR C. auris strains grown in nutrient-deficient or -enriched media at different temperatures. Phenotypic profiling revealed wider adaptability to nitrogenous nutrients and increased utilization of substrates critical in upper glycolysis and tricarboxylic acid cycle. Structural modeling of a 33-amino acid deletion in the gene for uracil phosphoribosyl transferase suggested an alternate route in C. auris to generate uracil monophosphate that does not accommodate 5-fluorouracil as a substrate. Overall, we find evidence of metabolic adaptations in MDR and PDR C. auris in response to antifungal drug lethality without deleterious fitness costs.

Cancer-associated SF3B1 Mutations Inhibit mRNA Nuclear Export by Disrupting SF3B1-THOC5 Interactions.

Mutations in SF3B1 are common in many types of cancer, which promotes cancer progression through aberrant RNA splicing. Recently, mRNA nuclear export has been reported to be defective in cells with SF3B1 K700E mutation. However, the mechanism remains unclear. Our study reveals that the K700E mutation in SF3B1 attenuates its interaction with THOC5, an essential component of mRNA nuclear export complex THO. Furthermore, SF3B1 mutation caused reduced binding of THOC5 with some mRNA and inhibited the nuclear export of these mRNA. Interestingly, THOC5 overexpression restores the nuclear export of these mRNA in cells with SF3B1 K700E mutation. Importantly, other types of cancer-associated SF3B1 mutations also inhibited mRNA nuclear export similarly, suggesting that it is common for cancer-associated SF3B1 mutation to inhibit mRNA nuclear export. Our research highlights the critical role of the THOC5-SF3B1 interaction in the regulation of mRNA nuclear export and provides valuable insights into the impact of SF3B1 mutations on mRNA nuclear export.

Alu insertion-mediated dsRNA structure formation with pre-existing Alu elements as a disease-causing mechanism

We previously identified a homozygous Alu insertion variant (Alu_Ins) in the 3'-untranslated region (3'-UTR) of SPINK1 as the cause of severe infantile isolated exocrine pancreatic insufficiency. Although we established that Alu_Ins leads to the complete loss of SPINK1 mRNA expression, the precise mechanisms remained elusive. Here, we aimed to elucidate these mechanisms through a hypothesis-driven approach. Initially, we speculated that, owing to its particular location, Alu_Ins could independently disrupt mRNA 3' end formation and/or affect other post-transcriptional processes such as nuclear export and translation. However, employing a 3'-UTR luciferase reporter assay, Alu_Ins was found to result in only an ∼50% reduction in luciferase activity compared to wild type, which is insufficient to account for the severe pancreatic deficiency in the Alu_Ins homozygote. We then postulated that double-stranded RNA (dsRNA) structures formed between Alu elements, an upstream mechanism regulating gene expression, might be responsible. Using RepeatMasker, we identified two Alu elements within SPINK1's third intron, both oriented oppositely to Alu_Ins. Through RNAfold predictions and full-length gene expression assays, we investigated orientation-dependent interactions between these Alu repeats. We provide compelling evidence to link the detrimental effect of Alu_Ins to extensive dsRNA structures formed between Alu_Ins and pre-existing intronic Alu sequences, including the restoration of SPINK1 mRNA expression by aligning all three Alu elements in the same orientation. Given the widespread presence of Alu elements in the human genome and the potential for new Alu insertions at almost any locus, our findings have important implications for detecting and interpreting Alu insertions in disease genes.

The Ubiquitination of Arrestin3 within the Nucleus Triggers the Nuclear Export of Mdm2, Which, in Turn, Mediates the Ubiquitination of GRK2 in the Cytosol.

GRK2 and arrestin3, key players in the functional regulation of G protein-coupled receptors (GPCRs), are ubiquitinated by Mdm2, a nuclear protein. The agonist-induced increase in arrestin3 ubiquitination occurs in the nucleus, underscoring the crucial role of its nuclear translocation in this process. The ubiquitination of arrestin3 occurs in the nucleus, highlighting the pivotal role of its nuclear translocation in this process. In contrast, GRK2 cannot translocate into the nucleus; thus, facilitation of the cytosolic translocation of nuclear Mdm2 is required to ubiquitinate GRK2 in the cytosol. Among the explored cellular components and processes, arrestin, Gβγ, clathrin, and receptor phosphorylation were found to be required for the nuclear import of arrestin3, the ubiquitination of arrestin3 in the nucleus, nuclear export of Mdm2, and the ubiquitination of GRK2 in the cytosol. In conclusion, our findings demonstrate that agonist-induced ubiquitination of arrestin3 in the nucleus is interconnected with cytosolic GRK2 ubiquitination.

RCC1 depletion drives protein transport defects and rupture in micronuclei.

Micronuclei (MN) are a commonly used marker of chromosome instability that form when missegregated chromatin recruits its own nuclear envelope (NE) after mitosis. MN frequently rupture, which results in genome instability, upregulation of metastatic genes, and increased immune signaling. MN rupture is linked to NE defects, but the cause of these defects is poorly understood. Previous work from our lab found that chromosome identity correlates with rupture timing for small MN, i.e. MN containing a short chromosome, with more euchromatic chromosomes forming more stable MN with fewer nuclear lamina gaps. Here we demonstrate that histone methylation promotes rupture and nuclear lamina defects in small MN. This correlates with increased MN size, and we go on to find that all MN have a constitutive nuclear export defect that drives MN growth and nuclear lamina gap expansion, making the MN susceptible to rupture. We demonstrate that these export defects arise from decreased RCC1 levels in MN and that additional loss of RCC1 caused by low histone methylation in small euchromatic MN results in additional import defects that suppress nuclear lamina gaps and MN rupture. Through analysis of mutational signatures associated with early and late rupturing chromosomes in the Pan-Cancer Analysis of Whole Genomes (PCAWG) dataset, we identify an enrichment of APOBEC and DNA polymerase E hypermutation signatures in chromothripsis events on early and mid rupturing chromosomes, respectively, suggesting that MN rupture timing could determine the landscape of structural variation in chromothripsis. Our study defines a new model of MN rupture where increased MN growth, caused by defects in protein export, drives gaps in nuclear lamina organization that make the MN susceptible to membrane rupture with long-lasting effects on genome architecture.

M5C-Seq: Machine learning-enhanced profiling of RNA 5-methylcytosine modifications

Epigenetic modifications, particularly RNA methylation and histone alterations, play a crucial role in heredity, development, and disease. Among these, RNA 5-methylcytosine (m5C) is the most prevalent RNA modification in mammalian cells, essential for processes such as ribosome synthesis, translational fidelity, mRNA nuclear export, turnover, and translation. The increasing volume of nucleotide sequences has led to the development of machine learning-based predictors for m5C site prediction. However, these predictors often face challenges related to training data limitations and overfitting due to insufficient external validation. This study introduces m5C-Seq, an ensemble learning approach for RNA modification profiling, designed to address these issues. m5C-Seq employs a meta-classifier that integrates 15 probabilities generated from a novel, large dataset using systematic encoding methods to make final predictions. Demonstrating superior performance compared to existing predictors, m5C-Seq represents a significant advancement in accurate RNA modification profiling. The code and the newly established datasets are made available through GitHub at https://github.com/Z-Abbas/m5C-Seq.

The RNA binding protein Arid5a drives IL-17-dependent autoantibody-induced glomerulonephritis.

Autoantibody-mediated glomerulonephritis (AGN) arises from dysregulated renal inflammation, with urgent need for improved treatments. IL-17 is implicated in AGN and drives pathology in a kidney-intrinsic manner via renal tubular epithelial cells (RTECs). Nonetheless, downstream signaling mechanisms provoking kidney pathology are poorly understood. A noncanonical RNA binding protein (RBP), Arid5a, was upregulated in human and mouse AGN. Arid5a-/- mice were refractory to AGN, with attenuated myeloid infiltration and impaired expression of IL-17-dependent cytokines and transcription factors (C/EBPβ, C/EBPδ). Transcriptome-wide RIP-Seq revealed that Arid5a inducibly interacts with conventional IL-17 target mRNAs, including CEBPB and CEBPD. Unexpectedly, many Arid5a RNA targets corresponded to translational regulation and RNA processing pathways, including rRNAs. Indeed, global protein synthesis was repressed in Arid5a-deficient cells, and C/EBPs were controlled at the level of protein rather than RNA accumulation. IL-17 prompted Arid5a nuclear export and association with 18S rRNA, a 40S ribosome constituent. Accordingly, IL-17-dependent renal autoimmunity is driven by Arid5a at the level of ribosome interactions and translation.