Premature termination codons (PTCs) can result in the production of truncated proteins or the degradation of messenger RNAs by nonsense-mediated mRNA decay (NMD). Which of these outcomes occurs can ...alter the effect of a mutation, with the engagement of NMD being dependent on a series of rules. Here, by applying these rules genome-wide to obtain a resource called NMDetective, we explore the impact of NMD on genetic disease and approaches to therapy. First, human genetic diseases differ in whether NMD typically aggravates or alleviates the effects of PTCs. Second, failure to trigger NMD is a cause of ineffective gene inactivation by CRISPR-Cas9 gene editing. Finally, NMD is a determinant of the efficacy of cancer immunotherapy, with only frameshifted transcripts that escape NMD predicting a response. These results demonstrate the importance of incorporating the rules of NMD into clinical decision-making. Moreover, they suggest that inhibiting NMD may be effective in enhancing cancer immunotherapy.
Premature termination codons (PTCs) cause a large proportion of inherited human genetic diseases. PTC-containing transcripts can be degraded by an mRNA surveillance pathway termed nonsense-mediated ...mRNA decay (NMD). However, the efficiency of NMD varies; it is inefficient when a PTC is located downstream of the last exon junction complex (EJC). We used matched exome and transcriptome data from 9,769 human tumors to systematically elucidate the rules of NMD targeting in human cells. An integrated model incorporating multiple rules beyond the canonical EJC model explains approximately three-fourths of the non-random variance in NMD efficiency across thousands of PTCs. We also show that dosage compensation may sometimes mask the effects of NMD. Applying the NMD model identifies signatures of both positive and negative selection on NMD-triggering mutations in human tumors and provides a classification for tumor-suppressor genes.
The nonsense-mediated mRNA decay (NMD) pathway degrades some but not all mRNAs bearing premature termination codons (PTCs). Decades of work have elucidated the molecular mechanisms of NMD. More ...recently, statistical analyses of large genomic datasets have allowed the importance of known and novel 'rules of NMD' to be tested and combined into methods that accurately predict whether PTC-containing mRNAs are degraded or not. We discuss these genomic approaches and how they can be applied to identify diseases and individuals that may benefit from inhibition or activation of NMD. We also discuss the importance of NMD for gene editing and tumor evolution, and how inhibiting NMD may be an effective strategy to increase the efficacy of cancer immunotherapy.
Genomic analyses have quantified the importance of known and novel 'rules of nonsense-mediated mRNA decay (NMD)', allowing accurate prediction of whether premature termination codon (PTC)-containing mRNAs are degraded or not.NMD can both aggravate and alleviate the effects of PTCs that cause genetic disease, and this varies across both diseases and individuals.Overall, NMD more frequently aggravates the effects of detrimental, disease-causing mutations.NMD frequently inactivates tumor-suppressor genes and silences the expression of neoantigens in cancer.NMD inhibition may be an effective strategy to enhance cancer immunotherapy and to treat a wide variety of genetic diseases.
Comprehensively characterizing the cellular composition and organization of tissues has been a long-term scientific challenge that has limited our ability to study fundamental and clinical aspects of ...human physiology. The Human Cell Atlas (HCA) is a global collaborative effort to create a reference map of all human cells as a basis for both understanding human health and diagnosing, monitoring, and treating disease. Many aspects of the HCA are analogous to the Human Genome Project (HGP), whose completion presents a major milestone in modern biology. To commemorate the HGP’s 20-year anniversary of completion, we discuss the launch of the HCA in light of the HGP, and highlight recent progress by the HCA consortium.
The Human Cell Atlas (HCA) consortium was founded as a collaborative and open effort to create a reference map of the cells in the human body.Organizing a large-scale project such as the HCA draws inspiration from the Human Genome Project (HGP) that was completed 20 years ago.Significant progress has been made by the HCA community, including profiling more than 39 million cells from 15 major organs to date.The expected impact of the HCA is illustrated by its use during the coronavirus disease 2019 (COVID-19) pandemic.
The deubiquitinating enzyme BAP1 is a tumor suppressor, among others involved in cholangiocarcinoma. BAP1 has many proposed molecular targets, while its Drosophila homolog is known to deubiquitinate ...histone H2AK119. We introduce BAP1 loss-of-function by CRISPR/Cas9 in normal human cholangiocyte organoids. We find that BAP1 controls the expression of junctional and cytoskeleton components by regulating chromatin accessibility. Consequently, we observe loss of multiple epithelial characteristics while motility increases. Importantly, restoring the catalytic activity of BAP1 in the nucleus rescues these cellular and molecular changes. We engineer human liver organoids to combine four common cholangiocarcinoma mutations (TP53, PTEN, SMAD4, and NF1). In this genetic background, BAP1 loss results in acquisition of malignant features upon xenotransplantation. Thus, control of epithelial identity through the regulation of chromatin accessibility appears to be a key aspect of BAP1’s tumor suppressor function. Organoid technology combined with CRISPR/Cas9 provides an experimental platform for mechanistic studies of cancer gene function in a human context.
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•BAP1 loss affects cell polarity and epithelial organization in human liver tissue•BAP1 controls chromatin accessibility of junctional and cytoskeletal genes•Rescuing BAP1 catalytic activity in the nucleus restores epithelial organization•In an engineered human cancer model, malignant features are induced upon BAP1 loss
Artegiani et al. show that BAP1 mutation in human liver organoids coincides with loss of multiple epithelial characteristics through impairment of chromatin accessibility and gene expression, and this is critical for the acquisition of malignant features in a human model of cholangiocarcinoma.
Assessment of single-cell gene expression (single-cell RNA sequencing) and adaptive immune receptor (AIR) sequencing (scVDJ-seq) has been invaluable in studying lymphocyte biology. Here we introduce ...Dandelion, a computational pipeline for scVDJ-seq analysis. It enables the application of standard V(D)J analysis workflows to single-cell datasets, delivering improved V(D)J contig annotation and the identification of nonproductive and partially spliced contigs. We devised a strategy to create an AIR feature space that can be used for both differential V(D)J usage analysis and pseudotime trajectory inference. The application of Dandelion improved the alignment of human thymic development trajectories of double-positive T cells to mature single-positive CD4/CD8 T cells, generating predictions of factors regulating lineage commitment. Dandelion analysis of other cell compartments provided insights into the origins of human B1 cells and ILC/NK cell development, illustrating the power of our approach. Dandelion is available at https://www.github.com/zktuong/dandelion .
Abstract Recent advances in single-cell immune profiling have enabled the simultaneous measurement of transcriptome and T cell receptor (TCR) sequences, offering great potential for studying immune ...responses at the cellular level. However, integrating these diverse modalities across datasets is challenging due to their unique data characteristics and technical variations. Here, to address this, we develop the multimodal generative model mvTCR to fuse modality-specific information across transcriptome and TCR into a shared representation. Our analysis demonstrates the added value of multimodal over unimodal approaches to capture antigen specificity. Notably, we use mvTCR to distinguish T cell subpopulations binding to SARS-CoV-2 antigens from bystander cells. Furthermore, when combined with reference mapping approaches, mvTCR can map newly generated datasets to extensive T cell references, facilitating knowledge transfer. In summary, we envision mvTCR to enable a scalable analysis of multimodal immune profiling data and advance our understanding of immune responses.
Nonsense-mediated decay (NMD) is a surveillance system that degrades mRNAs containing a premature termination codon (PTC) and plays important roles in protein homeostasis and disease. The efficiency ...of NMD is variable, impacting the clinical outcome of genetic mutations. However, limited resolution of bulk analyses has hampered the study of NMD efficiency. Here, we develop an assay to visualize NMD of individual mRNA molecules in real time. We find that NMD occurs with equal probability during each round of translation of an mRNA molecule. However, this probability is variable and depends on the exon sequence downstream of the PTC, the PTC-to-intron distance, and the number of introns both upstream and downstream of the PTC. Additionally, a subpopulation of mRNAs can escape NMD, further contributing to variation in NMD efficiency. Our study uncovers real-time dynamics of NMD, reveals key mechanisms that influence NMD efficiency, and provides a powerful method to study NMD.
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•Nonsense-mediated decay (NMD) of single mRNA molecules is visualized in live cells•Each terminating ribosome has an equal probability of inducing NMD•NMD efficiency is affected by the number and position of introns•Kinetics of XRN1-dependent decay of the 3′ mRNA cleavage fragment are uncovered
Hoek et al. develop a method to visualize nonsense-mediated mRNA decay (NMD) of individual mRNA molecules in live cells. Using this method, they uncover the precise timing of NMD, identify multiple parameters that affect NMD efficiency, and determine the kinetics of decay of the 3′ mRNA cleavage fragments by the exonuclease XRN1.
NuRD (nucleosome remodeling and histone deacetylase) is a versatile multi-protein complex with roles in transcription regulation and the DNA damage response. Here, we show that ZMYND8 bridges NuRD to ...a number of putative DNA-binding zinc finger proteins. The MYND domain of ZMYND8 directly interacts with PPPLΦ motifs in the NuRD subunit GATAD2A. Both GATAD2A and GATAD2B exclusively form homodimers and define mutually exclusive NuRD subcomplexes. ZMYND8 and NuRD share a large number of genome-wide binding sites, mostly active promoters and enhancers. Depletion of ZMYND8 does not affect NuRD occupancy genome-wide and only slightly affects expression of NuRD/ZMYND8 target genes. In contrast, the MYND domain in ZMYND8 facilitates the rapid, poly(ADP-ribose)-dependent recruitment of GATAD2A/NuRD to sites of DNA damage to promote repair by homologous recombination. Thus, these results show that a specific substoichiometric interaction with a NuRD subunit paralogue provides unique functionality to distinct NuRD subcomplexes.
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•ZMYND8 MYND domain interacts with PPPLФ motifs in NuRD subunit GATAD2A•GATAD2A and GATAD2B define mutually exclusive NuRD subcomplexes•ZMYND8 and NuRD co-localize on active (super)enhancers and promoters•ZMYND8 recruits GATAD2A/NuRD to DNA breaks in a poly(ADP-ribose)-dependent manner
Spruijt et al. show that the ZMYND8 MYND domain interacts with PPPLФ motifs in the NuRD subunit GATAD2A. Furthermore, GATAD2A and GATAD2B define mutually exclusive NuRD subcomplexes. The interaction between ZMYND8 and GATAD2A is required for poly(ADP-ribose)-dependent recruitment of NuRD to DNA double-strand breaks.
RNA modifications are integral to the regulation of RNA metabolism. One abundant mRNA modification is N
-methyladenosine (m
A), which affects various aspects of RNA metabolism, including splicing, ...translation and degradation. Current knowledge about the proteins recruited to m
A to carry out these molecular processes is still limited. Here we describe comprehensive and systematic mass-spectrometry-based screening of m
A interactors in various cell types and sequence contexts. Among the main findings, we identified G3BP1 as a protein that is repelled by m
A and positively regulates mRNA stability in an m
A-regulated manner. Furthermore, we identified FMR1 as a sequence-context-dependent m
A reader, thus revealing a connection between an mRNA modification and an autism spectrum disorder. Collectively, our data represent a rich resource and shed further light on the complex interplay among m
A, m
A interactors and mRNA homeostasis.
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