Biology subjects

Landthaler, M.

Publications and source records attributed to Landthaler, M..

At least 37 records · Page 2Linked to original sources

bioRxiv2022

A live attenuated vaccine confers superior mucosal and systemic immunity to SARS-CoV-2 variants

Vaccines are a cornerstone in COVID-19 pandemic management. Here, we compare immune responses to and preclinical efficacy of the mRNA vaccine BNT162b2, an adenovirus-vectored spike vaccine, and the live-attenuated-virus vaccine candidate sCPD9 after single and double vaccination in Syrian hamsters. All regimens containing sCPD9 showed superior efficacy. The robust immunity elicited by sCPD9 was evident in a wide range of immune parameters after challenge with heterologous SARS-CoV-2 including rapid viral clearance, reduced tissue damage, fast differentiation of pre-plasmablasts, strong systemic and mucosal humoral responses, and rapid recall of memory T cells from lung tissue. Our results demonstrate that use of live-attenuated vaccines may offer advantages over available COVID-19 vaccines, specifically when applied as booster, and may provide a solution for containment of the COVID-19 pandemic.

immunology↗

bioRxiv2022

In vitro Kinase-to-Phosphosite database (iKiP-DB) predicts kinase activity in phosphoproteomic datasets

Phosphoproteomics routinely quantifies changes in the levels of thousands of phosphorylation sites, but functional analysis of such data remains a major challenge. While databases like PhosphoSitePlus contain information about many phosphorylation sites, the vast majority of known sites are not assigned to any protein kinase. Assigning changes in the phosphoproteome to the activity of individual kinases therefore remains a key challenge.. A recent large-scale study systematically identified in vitro substrates for most human protein kinases. Here, we reprocessed and filtered these data to generate an in vitro Kinase-to-Phosphosite database (iKiP-DB). We show that iKiP-DB can accurately predict changes in kinase activity in published phosphoproteomic datasets for both well-studied and poorly characterized kinases. We apply iKiP-DB to a newly generated phosphoproteomic analysis of SARS-CoV-2 infected human lung epithelial cells and provide evidence for coronavirus-induced changes in host cell kinase activity. In summary, we show that iKiP-DB is widely applicable to facilitate the functional analysis of phosphoproteomic datasets.

bioinformatics↗

bioRxiv2021

In contrast to TH2-biased approaches, TH1 COVID-19 vaccines protect Syrian hamsters from severe disease in the absence of dexamethasone-treatable vaccine-associated enhanced respiratory pathology

Since December 2019, the novel human coronavirus SARS-CoV-2 has spread globally, causing millions of deaths. Unprecedented efforts have enabled development and authorization of a range of vaccines, which reduce transmission rates and confer protection against the associated disease COVID-19. These vaccines are conceptually diverse, including e.g. classical adjuvanted whole-inactivated virus, viral vectors, and mRNA vaccines. We have analysed two prototypic model vaccines, the strongly TH1-biased measles vaccine-derived candidate MeVvac2-SARS2-S(H) and a TH2-biased Alum-adjuvanted, non-stabilized Spike (S) protein side-by-side, for their ability to protect Syrian hamsters upon challenge with a low-passage SARS-CoV-2 patient isolate. As expected, the MeVvac2-SARS2-S(H) vaccine protected the hamsters safely from severe disease. In contrast, the protein vaccine induced vaccine-associated enhanced respiratory disease (VAERD) with massive infiltration of eosinophils into the lungs. Global RNA-Seq analysis of hamster lungs revealed reduced viral RNA and less host dysregulation in MeVvac2-SARS2-S(H) vaccinated animals, while S protein vaccination triggered enhanced host gene dysregulation compared to unvaccinated control animals. Of note, mRNAs encoding the major eosinophil attractant CCL-11, the TH2 response-driving cytokine IL-19, as well as TH2-cytokines IL-4, IL-5, and IL-13 were exclusively up-regulated in the lungs of S protein vaccinated animals, consistent with previously described VAERD induced by RSV vaccine candidates. IL-4, IL-5, and IL-13 were also up-regulated in S-specific splenocytes after protein vaccination. Using scRNA-Seq, T cells and innate lymphoid cells were identified as the source of these cytokines, while Ccl11 and Il19 mRNAs were expressed in lung macrophages displaying an activated phenotype. Interestingly, the amount of viral reads in this macrophage population correlated with the abundance of Fc-receptor reads. These findings suggest that VAERD is triggered by induction of TH2-type helper cells secreting IL-4, IL-5, and IL-13, together with stimulation of macrophage subsets dependent on non-neutralizing antibodies. Via this mechanism, uncontrolled eosinophil recruitment to the infected tissue occurs, a hallmark of VAERD immunopathogenesis. These effects could effectively be treated using dexamethasone and were not observed in animals vaccinated with MeVvac2-SARS2-S(H). Taken together, our data validate the potential of TH2-biased COVID-19 vaccines and identify the transcriptional mediators that underlie VAERD, but confirm safety of TH1-biased vaccine concepts such as vector-based or mRNA vaccines. Dexamethasone, which is already in use for treatment of severe COVID-19, may alleviate such VAERD, but in-depth scrutiny of any next-generation protein-based vaccine candidates is required, prior and after their regulatory approval.

immunology↗

bioRxiv2021

Insights into standards of care: dexamethasone and antibodies against COVID-19 in hamster models

RationaleIn face of the ongoing SARS-CoV-2 pandemic, effective and well-understood treatment options are still scarce. While vaccines have proven instrumental in fighting SARS-CoV-2, their efficacy is challenged by vaccine hesitancy, novel variants and short-lasting immunity. Therefore, understanding and optimization of therapeutic options remains essential. ObjectivesWe aimed at generating a deeper understanding on how currently used drugs, specifically dexamethasone and anti-SARS-CoV-2 antibodies, affect SARS-CoV-2 infection and host responses. Possible synergistic effects of both substances are investigated to evaluate combinatorial treatments. MethodsBy using two COVID-19 hamster models, pulmonary immune responses were analyzed to characterize effects of treatment with either dexamethasone, anti-SARS-CoV-2 spike monoclonal antibody or a combination of both. scRNA sequencing was employed to reveal transcriptional response to treatment on a single cell level. Measurements and main resultsDexamethasone treatment resulted in similar or increased viral loads compared to controls. Anti-SARS-CoV-2 antibody treatment alone or combined with dexamethasone successfully reduced pulmonary viral burden. Dexamethasone exhibited strong anti-inflammatory effects and prevented fulminant disease in a severe COVID-19-like disease model. Combination therapy showed additive benefits with both anti-viral and anti-inflammatory potency. Bulk and single-cell transcriptomic analyses confirmed dampened inflammatory cell recruitment into lungs upon dexamethasone treatment and identified a candidate subpopulation of neutrophils specifically responsive to dexamethasone. ConclusionsOur analyses i) confirm the anti-inflammatory properties and indicate possible modes of action for dexamethasone, ii) validate anti-viral effects of anti-SARS-CoV-2 antibody treatment, and iii) reveal synergistic effects of a combination therapy and can thus inform more effective COVID-19 therapies.

molecular biology↗

bioRxiv2021

Transcriptomic responses of the human kidney to acute injury at single cell resolution

BackgroundAcute kidney injury (AKI) occurs frequently in critically ill patients and is associated with adverse outcomes. Cellular mechanisms underlying AKI and kidney cell responses to injury remain incompletely understood. MethodsWe performed single-nuclei transcriptomics, bulk transcriptomics, molecular imaging studies, and conventional histology on kidney tissues from 8 individuals with severe AKI (stage 2 or 3 according to Kidney Disease: Improving Global Outcomes (KDIGO) criteria). Specimens were obtained within 1-2 hours after individuals had succumbed to critical illness associated with respiratory infections, with 4 of 8 individuals diagnosed with COVID-19. Control kidney tissues were obtained post-mortem or after nephrectomy from individuals without AKI. ResultsHigh-depth single cell-resolved gene expression data of human kidneys affected by AKI revealed enrichment of novel injury-associated cell states within the major cell types of the tubular epithelium, in particular in proximal tubules, thick ascending limbs and distal convoluted tubules. Four distinct, hierarchically interconnected injured cell states were distinguishable and characterized by transcriptome patterns associated with oxidative stress, hypoxia, interferon response, and epithelial-to-mesenchymal transition, respectively. Transcriptome differences between individuals with AKI were driven primarily by the cell type-specific abundance of these four injury subtypes rather than by private molecular responses. AKI-associated changes in gene expression between individuals with and without COVID-19 were similar. ConclusionThe study provides an extensive resource of the cell type-specific transcriptomic responses associated with critical illness-associated AKI in humans, highlighting recurrent disease-associated signatures and inter-individual heterogeneity. Personalized molecular disease assessment in human AKI may foster the development of tailored therapies.

molecular biology↗

bioRxiv2021

Global analysis of human-to-mouse intercellular RNA transfer in cell culture

Full-length mRNAs can transfer between adjacent mammalian cells via direct cell-to-cell connections called tunneling nanotubes (TNTs). However, the extent of mRNA transfer at the transcriptome-wide level (the transferome) is unknown. Here, we analyzed whole transcriptome mRNA and lncRNA transfer between heterogeneous human-mouse cell populations in in vitro co-culture using RNA-sequencing. Our data indicate that mRNA transfer is non-selective, prevalent across the human transcriptome, and that the amount of transfer to mouse embryonic fibroblasts (MEFs) strongly correlates with the endogenous level of gene expression in donor human breast cancer cells (MCF7). These results were validated by both quantitative RT-PCR and in situ hybridization, and analysis shows that typically <1% of endogenous mRNAs and lncRNAs undergo transfer. Non-selective expression-dependent RNA transfer was further validated using synthetic RNA reporters. Notably, significant differential changes in the native MEF transcriptome were observed in response to co-culture, including the upregulation of multiple cancer and cancer-associated fibroblast-related genes and pathways. Together, these results lead us to suggest that TNT-mediated RNA transfer could be a phenomenon of physiological importance under both normal and pathogenic conditions. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=183 SRC="FIGDIR/small/470233v2_ufig1.gif" ALT="Figure 1"> View larger version (44K): org.highwire.dtl.DTLVardef@13a7176org.highwire.dtl.DTLVardef@1ef1c75org.highwire.dtl.DTLVardef@1355eeforg.highwire.dtl.DTLVardef@ebc275_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2021

De Novo Whole Genome Assembly of the Roborovski Dwarf Hamster (Phodopus roborovskii) Genome, an Animal Model for Severe/Critical COVID-19

The Roborovski dwarf hamster Phodopus roborovskii belongs to the Phodopus genus, one of seven within Cricetinae subfamily. Like other rodents such as mice, rats or ferrets, hamsters can be important animal models for a range of diseases. Whereas the Syrian hamster from the genus Mesocricetus is now widely used as a model for mild to moderate COVID-19, Roborovski dwarf hamster show a severe to lethal course of disease upon infection with the novel human coronavirus SARS-CoV-2.

genomics↗

bioRxiv2021

ENGINEERING, DECODING AND SYSTEMS-LEVEL CHARACTERIZATION OF CHIMPANZEE CYTOMEGALOVIRUS

The chimpanzee cytomegalovirus (CCMV) is the closest relative of human CMV (HCMV). Because of the high conservation between these two species and the ability of human cells to fully support CCMV replication, CCMV holds great potential as a model system for HCMV. To make the CCMV genome available for precise and rapid gene manipulation techniques, we captured the genomic DNA of CCMV strain Heberling as a bacterial artificial chromosome (BAC). Selected BAC clones were reconstituted to infectious viruses, growing to similar high titers as parental CCMV. DNA sequencing confirmed the integrity of our clones and led to the identification of two polymorphic loci within the CCMV genome. To re-evaluate the CCMV coding potential, we analyzed the transcriptome and proteome of infected cells and identified several novel ORFs, splice variants, and regulatory RNAs. We further characterized the dynamics of CCMV gene expression and found that viral proteins cluster into five distinct temporal classes. In addition, our datasets revealed that the host response to CCMV infection and the de-regulation of cellular pathways are in line with known hallmarks of HCMV infection. In a first functional experiment, we investigated a proposed frameshift mutation in UL128 that was suspected to restrict CCMVs cell tropism. In fact, repair of this frameshift re-established productive CCMV infection in endothelial and epithelial cells, expanding the options of CCMV as an infection model. Thus, BAC-cloned CCMV can serve as a powerful tool for systematic approaches in comparative functional genomics, exploiting the close phylogenetic relationship between CCMV and HCMV.

microbiology↗

bioRxiv2021

Timed global reorganization of protein synthesis during neocortex neurogenesis at codon resolution

Translation modulates the timing and amplification of gene expression after transcription. Brain development requires uniquely complex gene expression patterns, but large-scale measurements of translation directly in the prenatal brain are lacking. We measure the reactants, synthesis, and products of translation spanning mouse neocortex neurogenesis, and discover a transient window of dynamic regulation at mid-gestation. Timed translation upregulation of chromatin binding proteins like Satb2, which is essential for neuronal subtype differentiation, restricts protein expression in neuronal lineages despite broad transcriptional priming in progenitors. In contrast, translation downregulation of ribosomal proteins sharply decreases ribosome number, coinciding with a major shift in protein synthesis dynamics at mid-gestation. Changing levels of eIF4EBP1, a direct inhibitor of ribosomal protein translation, are concurrent with ribosome downregulation and controls Satb2 fate acquisition during neuronal differentiation. Thus, the refinement of transcriptional programs by translation is central to the molecular logic of brain development. Modeling of the developmental neocortex translatome is provided as an open-source searchable resource: https://shiny.mdc-berlin.de/cortexomics/.

neuroscience↗

bioRxiv2021

Herpesviral induction of germline transcription factor DUX4 is critical for viral gene expression

DUX4 is a transcription factor and a master regulator of embryonic genome activation (EGA). During early embryogenesis, EGA is crucial for maternal to zygotic transition at the 8-cell stage in order to overcome silencing of genes and enable transcription from the zygotic genome. In adult somatic cells, DUX4 expression is largely silenced. Activation is likely pathogenic, and in adult muscle cells causes genetic disorder Facioscapulohumeral Muscular Dystrophy (FSHD). We identified activation of DUX4 expression upon lytic replication of the herpesviruses HSV-1, HCMV, EBV and KSHV, but not of adenoviruses, negative strand RNA viruses or positive strand RNA viruses. We demonstrate by RNA-Seq analysis that DUX4 expression upon herpesviral replication leads to the induction of hundreds of DUX4 target genes including germline-specific retroelements as well as several members of the TRIM, PRAMEF and ZSCAN protein families. Moreover, we show that DUX4 expression is a direct consequence of herpesviral infection. DUX4 can be stimulated by overexpression of HSV-1 immediate early proteins, indicating active induction of EGA genes by herpesviral infection. We further show that DUX4 expression is critical for driving HSV-1 gene expression. Our results show that viruses from alpha-, beta- and gamma-herpesvirus subfamilies induce DUX4 expression and downstream germline-specific genes and retroelements. We hypothesize that herpesviruses induce DUX4 expression in order to induce an early embryonic-like transcriptional program that prevents epigenetic silencing of the viral genome and facilitates herpesviral gene expression.

microbiology↗

bioRxiv2021

Neurodegeneration in human brain organoids infected with herpes simplex virus type 1

Herpes simplex virus type 1 (HSV-1) infection of the nervous system may lead to brain damage, including neurodegeneration. However, lack of suitable experimental models hinders understanding molecular mechanisms and cell-type-specific responses triggered by HSV-1. Here, we infected human brain organoids with HSV-1. Known features of HSV-1 infection such as alteration of neuronal electrophysiology and induction of antisense transcription were confirmed. Full-length mRNA-sequencing revealed aberrant 3 end formation and poly(A)-tail lengthening. Single-cell RNA-seq and spatial transcriptomics uncovered changes in the cellular composition of the infected organoids caused by viral replication and dysregulation of molecular pathways in cell-type specific manner. Furthermore, hallmarks of early neurodegeneration were observed, namely extracellular matrix disruption, STMN2 and TARDBP/TDP43 downregulation, and upregulation of the AD-related non-coding RNA BC200/BCYRN1. These hallmarks were weaker/absent when infecting with a mutant HSV-1 control. Together, our data indicate that brain organoids serve as a powerful model to study mechanisms of HSV-1-driven neurodegeneration.

neuroscience↗

bioRxiv2020

Longitudinal omics in Syrian hamsters integrated with human data unravel complexity of moderate immune responses to SARS-CoV-2

In COVID-19, the immune response largely determines disease severity and is key to therapeutic strategies. Cellular mechanisms contributing to inflammatory lung injury and tissue repair in SARS-CoV-2 infection, particularly endothelial cell involvement, remain ill-defined. We performed detailed spatiotemporal analyses of cellular and molecular processes in SARS-CoV-2 infected Syrian hamsters. Comparison of hamster single-cell sequencing and proteomics with data sets from COVID-19 patients demonstrated inter-species concordance of cellular and molecular host-pathogen interactions. In depth vascular and pulmonary compartment analyses (i) supported the hypothesis that monocyte-derived macrophages dominate inflammation, (ii) revealed endothelial inflammation status and T-cell attraction, and (iii) showed that CD4+ and CD8+ cytotoxic T-cell responses precede viral elimination. Using the Syrian hamster model of self-limited moderate COVID-19, we defined the specific roles of endothelial and epithelial cells, among other myeloid and non-myeloid lung cell subtypes, for determining the disease course.

immunology↗

bioRxiv2020

Spatio-temporal mRNA dynamics in the early zebrafish embryo

Early stages of embryogenesis depend heavily on subcellular localization and transport of maternally deposited mRNA. However, systematic analysis of these processes is currently hindered by a lack of spatio-temporal information in single-cell RNA sequencing. Here, we combined spatially-resolved transcriptomics and single-cell RNA labeling to study the spatio-temporal dynamics of the transcriptome during the first few hours of zebrafish development. We measured spatial localization of mRNA molecules with sub-single-cell resolution at the one-cell stage, which allowed us to identify a class of mRNAs that are specifically localized at an extraembryonic position, the vegetal pole. Furthermore, we established a method for high-throughput single-cell RNA labeling in early zebrafish embryos, which enabled us to follow the fate of individual maternal transcripts until gastrulation. This approach revealed that many localized transcripts are specifically transported to the primordial germ cells. Finally, we acquired spatial transcriptomes of two xenopus species, and we compared evolutionary conservation of localized genes as well as enriched sequence motifs. In summary, we established sub-single-cell spatial transcriptomics and single-cell RNA labeling to reveal principles of mRNA localization in early vertebrate development.

systems biology↗

bioRxiv2020

The zinc finger antiviral protein ZAP destabilises viral transcripts and restricts cytomegalovirus

Interferon-stimulated gene products (ISGs) play a crucial role in early infection control. The ISG zinc finger CCCH-type antiviral protein 1 (ZAP/ZC3HAV1) antagonises several RNA viruses by binding to CG-rich RNA sequences, whereas its effect on DNA viruses is largely unknown. Here, we decipher the role of ZAP in the context of human cytomegalovirus (HCMV) infection, a {beta}-herpesvirus that is associated with high morbidity in immunosuppressed individuals and newborns. We show that expression of the two major isoforms of ZAP, the long (ZAP-L) and short (ZAP-S), is induced during HCMV infection and that both negatively affect HCMV replication. Transcriptome and proteome analyses demonstrated that the expression of ZAP decelerates the progression of HCMV infection. SLAM-sequencing revealed that ZAP restricts HCMV at early stages of infection by destabilising a distinct subset of viral transcripts with low CG content. In summary, this report provides evidence of an important antiviral role for ZAP in host defense against HCMV infection and highlights its differentiated function during DNA virus infection.

molecular biology↗

bioRxiv2020

Single Cell RNA-Sequencing-based Analysis of CD4+ T-Cell Subset-Specific Susceptibility to Transcriptional Modulation by HIV-1 Latency-Reversing Agents

Shock-and-kill is one of the most advanced, yet unrealized, concepts towards establishment of HIV-1 cure. Treatment with latency-reversing agents (LRAs), including histone deacetylase inhibitors (HDACis) exerting chromatin remodelling and gene expression reprogramming, combined with anti-retroviral therapy reactivates HIV-1 transcription in vitro, ex vivo and in vivo. However, HDACi treatment fails to significantly reduce the size of the viral reservoir in people living with HIV-1 (PLHIV). Here, by combining scRNA-seq and functional approaches, we characterised the HDACi treatment-imposed remodulation of CD4+ T-cells state and its consequences for HIV-1 latency reversal and the apparent resistance of HIV-1-reactivating cells to immune-mediated elimination. Exposure of CD4+ T-cells from three aviremic PLHIV with clinically applicable concentrations of Panobinostat markedly reduced the expression of genes mediating T-cell activation and IFN-driven antiviral immunity in a largely CD4+ T-cell subset-nonspecific manner, with exception of an PLHIV-specific exhausted CD4+ T-cell subpopulation. Altered transcriptomic profiles were accompanied by large refractoriness to peptide and IL-2/PHA stimulation, and to exogenous type I interferon, that would otherwise induce T-cell activation and expression of a plethora of antiviral genes, respectively. Type I interferon, when added to Panobinostat during HIV-1 reactivation, was unable to counteract HDACi-mediated inhibition of IFN signalling and failed to interfere with HIV-1 reactivation per se. However, it imposed a pre-budding block and boosted surface levels of HIV-1 Env on reactivating cells. Co-treatment with type I IFNs, most prominently IFN-{beta} and -14, sensitised HIV-1-reactivating cells for killing by NK cells through antibody-dependent cytotoxicity. Together, our study provides proof-of-concept of the benefit of combining a potent LRA with immunostimulatory molecules, such as type I IFNs, to reduce the resistance of HIV-1-reactivating T-cells to immune-mediated elimination to improve current shock-and-kill strategies.

microbiology↗

bioRxiv2020

Bulk and single-cell gene expression profiling of SARS-CoV-2 infected human cell lines identifies molecular targets for therapeutic intervention

The coronavirus disease 2019 (COVID-19) pandemic, caused by the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is an ongoing global health threat with more than two million infected people since its emergence in late 2019. Detailed knowledge of the molecular biology of the infection is indispensable for understanding of the viral replication, host responses, and disease progression. We provide gene expression profiles of SARS-CoV and SARS-CoV-2 infections in three human cell lines (H1299, Caco-2 and Calu-3 cells), using bulk and single-cell transcriptomics. Small RNA profiling showed strong expression of the immunity and inflammation-associated microRNA miRNA-155 upon infection with both viruses. SARS-CoV-2 elicited approximately two-fold higher stimulation of the interferon response compared to SARS-CoV in the permissive human epithelial cell line Calu-3, and induction of cytokines such as CXCL10 or IL6. Single cell RNA sequencing data showed that canonical interferon stimulated genes such as IFIT2 or OAS2 were broadly induced, whereas interferon beta (IFNB1) and lambda (IFNL1-4) were expressed only in a subset of infected cells. In addition, temporal resolution of transcriptional responses suggested interferon regulatory factors (IRFs) activities precede that of nuclear factor-{kappa}B (NF-{kappa}B). Lastly, we identified heat shock protein 90 (HSP90) as a protein relevant for the infection. Inhibition of the HSP90 charperone activity by Tanespimycin/17-N-allylamino-17-demethoxygeldanamycin (17-AAG) resulted in a reduction of viral replication, and of TNF and IL1B mRNA levels. In summary, our study established in vitro cell culture models to study SARS-CoV-2 infection and identified HSP90 protein as potential drug target for therapeutic intervention of SARS-CoV-2 infection.

systems biology↗

bioRxiv2020

The architecture of protein synthesis in the developing neocortex at near-atomic resolution reveals Ebp1-mediated neuronal proteostasis at the 60S tunnel exit

Protein synthesis must be finely tuned in the nervous system, as it represents an essential feature of neurodevelopmental gene expression, and dominant pathology in neurological disease. However, the architecture of ribosomal complexes in the developing mammalian brain has not been analyzed at high resolution. This study investigates the architecture of ribosomes ex vivo from the embryonic and perinatal mouse neocortex, revealing Ebp1 as a 60S peptide tunnel exit binding factor at near-atomic resolution by multiparticle cryo-electron microscopy. The impact of Ebp1 on the neuronal proteome was analyzed by pSILAC and BONCAT coupled mass spectrometry, implicating Ebp1 in neurite outgrowth proteostasis, with in vivo embryonic Ebp1 knockdown resulting in dysregulation of neurite outgrowth. Our findings reveal Ebp1 as a central component of neocortical protein synthesis, and the 60S peptide tunnel exit as a focal point of gene expression control in the molecular specification of neuronal morphology. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/939488v1_ufig1.gif" ALT="Figure 1"> View larger version (59K): org.highwire.dtl.DTLVardef@1231351org.highwire.dtl.DTLVardef@1c940f7org.highwire.dtl.DTLVardef@1c81e2corg.highwire.dtl.DTLVardef@135008b_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2020

CRNKL1 is a highly selective regulator of intron-retaining HIV-1 and cellular mRNAs

The HIV-1 Rev protein is a nuclear export factor for unspliced and incompletely-spliced HIV-1 RNAs. Without Rev, these intron-retaining RNAs are trapped in the nucleus. A genome-wide screen identified nine proteins of the spliceosome which all enhanced expression from the HIV-1 unspliced RNA after CRISPR/Cas knock-down. Depletion of DHX38, WDR70 and four proteins of the Prp19-associated complex (ISY1, BUD31, XAB2, CRNKL1) resulted in a more than 20-fold enhancement of unspliced HIV-1 RNA levels in the cytoplasm. Targeting of CRNKL1, DHX38, and BUD31 affected nuclear export efficiencies of the HIV-1 unspliced RNA to a much larger extent than splicing. Transcriptomic analyses further revealed that CRNKL1 also suppresses cytoplasmic levels of cellular mRNAs with selectively retained introns. Thus, CRNKL1 dependent nuclear retention seems to be a novel mechanism for the regulation of cytoplasmic levels of intron-retaining cellular mRNAs that is harnessed by HIV-1 to direct its complex splicing pattern.

molecular biology↗