The m6A reader IMP2 directs autoimmune inflammation through an IL-17– and TNFα-dependent C/EBP transcription factor axis
Abstract: Excessive cytokine activity underlies many autoimmune conditions, particularly through the interleukin-17 (IL-17) and tumor necrosis factor-α (TNFα) signaling axis. Both cytokines activate nuclear factor κB, but appropriate induction of downstream effector genes requires coordinated activation of other transcription factors, notably, CCAAT/enhancer binding proteins (C/EBPs). Here, we demonstrate the unexpected involvement of a posttranscriptional "epitranscriptomic" mRNA modification [N6-methyladenosine (m6A)] in regulating C/EBPβ and C/EBPδ in response to IL-17A, as well as IL-17F and TNFα. Prompted by the observation that C/EBPβ/δ-encoding transcripts contain m6A consensus sites, we show that Cebpd and Cebpb mRNAs are subject to m6A modification. Induction of C/EBPs is enhanced by an m6A methylase "writer" and suppressed by a demethylase "eraser." The only m6A "reader" found to be involved in this pathway was IGF2BP2 (IMP2), and IMP2 occupancy of Cebpd and Cebpb mRNA was enhanced by m6A modification. IMP2 facilitated IL-17-mediated Cebpd mRNA stabilization and promoted translation of C/EBPβ/δ in response to IL-17A, IL-17F, and TNFα. RNA sequencing revealed transcriptome-wide IL-17-induced transcripts that are IMP2 influenced, and RNA immunoprecipitation sequencing identified the subset of mRNAs that are directly occupied by IMP2, which included Cebpb and Cebpd Lipocalin-2 (Lcn2), a hallmark of autoimmune kidney injury, was strongly dependent on IL-17, IMP2, and C/EBPβ/δ. Imp2-/- mice were resistant to autoantibody-induced glomerulonephritis (AGN), showing impaired renal expression of C/EBPs and Lcn2 Moreover, IMP2 deletion initiated only after AGN onset ameliorated disease. Thus, posttranscriptional regulation of C/EBPs through m6A/IMP2 represents a previously unidentified paradigm of cytokine-driven autoimmune inflammation.
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TL;DR: The authors showed that m6A modifications by METTL3 are required for germinal center (GC) reaction through the differential functions of m6As readers, and showed that two independent gene networks that support critical GC functions are modulated by m6a through distinct mRNA binders.
Abstract: Long-lasting immunity depends on the generation of protective antibodies through the germinal center (GC) reaction. N6-methyladenosine (m6A) modification of mRNAs by METTL3 activity modulates transcript lifetime primarily through the function of m6A readers; however, the physiological role of this molecular machinery in the GC remains unknown. Here, we show that m6A modifications by METTL3 are required for GC maintenance through the differential functions of m6A readers. Mettl3-deficient GC B cells exhibited reduced cell-cycle progression and decreased expression of proliferation- and oxidative phosphorylation-related genes. The m6A binder, IGF2BP3, was required for stabilization of Myc mRNA and expression of its target genes, whereas the m6A reader, YTHDF2, indirectly regulated the expression of the oxidative phosphorylation gene program. Our findings demonstrate how two independent gene networks that support critical GC functions are modulated by m6A through distinct mRNA binders.
20 citations
TL;DR: The up-to-date knowledges of IGF2BPs (IGF2BP1/2/3) as m6A readers in an m6a-modified manner in cancer progression are reviewed, and the oncogenic role of IGF 2BPs in cancer is discussed.
Abstract: RNA can be modified by over 170 types of distinct chemical modifications, and the most abundant internal modification of mRNA in eukaryotes is N6-methyladenosine (m6A). The m6A modification accelerates mRNA process, including mRNA splicing, translation, transcript stability, export and decay. m6A RNA modification is installed by methyltransferase-like proteins (writers), and potentially removed by demethylases (erasers), and this process is recognized by m6A-binding proteins (readers). Notably, alterations of m6A-modified proteins (writers, erasers and readers) are involved in the tumorigenesis, progression and metastasis. Importantly, the fate of m6A-methylated mRNA is mediated mostly through m6A readers, and among these readers, insulin-like growth factor 2 mRNA-binding proteins (IGF2BPs) are unique RNA-binding proteins (RBPs) that stabilize their targets mRNA via m6A modification. In this review, we update the writers, erasers and readers, and their cross-talks in m6A modification, and briefly discuss the oncogenic role of IGF2BPs in cancer. Most importantly, we mainly review the up-to-date knowledges of IGF2BPs (IGF2BP1/2/3) as m6A readers in an m6A-modified manner in cancer progression.
12 citations
TL;DR: In this article , a total of 65,362 single cells from single-cell RNA-seq data derived from 33 CRC tumor samples were analyzed by nonnegative matrix factorization (NMF) for 23 m 6 A RNA methylation regulators.
Abstract: Abstract Background N6-methyladenosine (m 6 A) RNA methylation plays a critical role in key genetic events for various cancers; yet, how m 6 A functions within the tumor microenvironment (TME) remains to be elucidated. Methods A total of 65,362 single cells from single-cell RNA-seq data derived from 33 CRC tumor samples were analyzed by nonnegative matrix factorization (NMF) for 23 m 6 A RNA methylation regulators. CRC and Immunotherapy cohorts from public repository were used to determine the prognosis and immune response of TME clusters. Results The fibroblasts, macrophages, T and B cells were respectively grouped into 4 to 5 subclusters and then classified according to various biological processes and different marker genes. Furthermore, it revealed that the m 6 A RNA methylation regulators might be significantly related to the clinical and biological features of CRC, as well as the pseudotime trajectories of main TME cell types. Bulk-seq analysis suggested that these m 6 A-mediated TME cell subclusters had significant prognostic value for CRC patients and distinguished immune response for patients who underwent ICB therapy, especially for the CAFs and macrophages. Notably, CellChat analysis revealed that RNA m 6 A methylation-associated cell subtypes of TME cells manifested diverse and extensive interaction with tumor epithelial cells. Further analysis showed that ligand-receptor pairs, including MIF − (CD74 + CXCR4), MIF − (CD74 + CD44), MDK–NCL and LGALS9 − CD45, etc. mediated the communication between m 6 A associated subtypes of TME cells and tumor epithelial cells. Conclusions Taken together, our study firstly revealed the m 6 A methylation mediated intercellular communication of the tumor microenvironment in the regulation of tumor growth and antitumor immunomodulatory processes.
10 citations
TL;DR: In this paper , a total of 65,362 single cells from single-cell RNA-seq data derived from 33 CRC tumor samples were analyzed by nonnegative matrix factorization (NMF) for 23 m 6 A RNA methylation regulators.
Abstract: Abstract Background N6-methyladenosine (m 6 A) RNA methylation plays a critical role in key genetic events for various cancers; yet, how m 6 A functions within the tumor microenvironment (TME) remains to be elucidated. Methods A total of 65,362 single cells from single-cell RNA-seq data derived from 33 CRC tumor samples were analyzed by nonnegative matrix factorization (NMF) for 23 m 6 A RNA methylation regulators. CRC and Immunotherapy cohorts from public repository were used to determine the prognosis and immune response of TME clusters. Results The fibroblasts, macrophages, T and B cells were respectively grouped into 4 to 5 subclusters and then classified according to various biological processes and different marker genes. Furthermore, it revealed that the m 6 A RNA methylation regulators might be significantly related to the clinical and biological features of CRC, as well as the pseudotime trajectories of main TME cell types. Bulk-seq analysis suggested that these m 6 A-mediated TME cell subclusters had significant prognostic value for CRC patients and distinguished immune response for patients who underwent ICB therapy, especially for the CAFs and macrophages. Notably, CellChat analysis revealed that RNA m 6 A methylation-associated cell subtypes of TME cells manifested diverse and extensive interaction with tumor epithelial cells. Further analysis showed that ligand-receptor pairs, including MIF − (CD74 + CXCR4), MIF − (CD74 + CD44), MDK–NCL and LGALS9 − CD45, etc. mediated the communication between m 6 A associated subtypes of TME cells and tumor epithelial cells. Conclusions Taken together, our study firstly revealed the m 6 A methylation mediated intercellular communication of the tumor microenvironment in the regulation of tumor growth and antitumor immunomodulatory processes.
10 citations
TL;DR: Wang et al. as discussed by the authors summarized current evidences on m6A mediates the pathophysiological process in cardiac development and cardiomyopathy to emphasize the importance of RNA methylation in maintain heart biogenesis and homeostasis.
Abstract: Sepsis is a life-threatening organ dysfunction caused by a host's dysfunctional response to infection. As is known to all, septic heart disease occurs because pathogens invading the blood stimulate the activation of endothelial cells, causing a large number of white blood cells to accumulate and trigger an immune response. However, in severe sepsis, the hematopoietic system is inhibited, and there will also be a decline in white blood cells, at which time the autoimmune system will also be suppressed. During the immune response, a large number of inflammatory factors are released into cells to participate in the inflammatory process, which ultimately damages cardiac myocytes and leads to impaired cardiac function. N6-methyladenosine (m6A) is a common RNA modification in mRNA and non-coding RNA that affects RNA splicing, translation, stability, and epigenetic effects of some non-coding RNAs. A large number of emerging evidences demonstrated m6A modification had been involved in multiple biological process. Especially for sepsis and immune disorder diseases. Unfortunately, there is limited results provided to analysis the association between m6A modification and SICD. In this review, we firstly summarized current evidences on m6A mediates the pathophysiological process in cardiac development and cardiomyopathy to emphasize the importance of RNA methylation in maintain heart biogenesis and homeostasis. Then we clarified the participants of m6A modification in extended inflammation responses and immune system activation, which is the dominant and initial changes secondary to sepsis attack. After that we deeply analyzed the top causes of SICD, and identified activation of inflammatory cytokines, endothelial cells dysfunction and mitochondrial failure. Thus, the highlights of this review is that we systematically collected all the related potential mechanisms between m6A modification and SICD causes. Although there is lacking of direct evidences on SICD, but indirect evidences had been demonstrated case by case on every particular molecular mechanisms and signaling transduction, which requires the further explorations into the potential links among the listed mechanisms. it provides novel insights into the understanding of SICD.
10 citations
References
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TL;DR: This work presents DESeq2, a method for differential analysis of count data, using shrinkage estimation for dispersions and fold changes to improve stability and interpretability of estimates, which enables a more quantitative analysis focused on the strength rather than the mere presence of differential expression.
Abstract: In comparative high-throughput sequencing assays, a fundamental task is the analysis of count data, such as read counts per gene in RNA-seq, for evidence of systematic changes across experimental conditions. Small replicate numbers, discreteness, large dynamic range and the presence of outliers require a suitable statistical approach. We present DESeq2, a method for differential analysis of count data, using shrinkage estimation for dispersions and fold changes to improve stability and interpretability of estimates. This enables a more quantitative analysis focused on the strength rather than the mere presence of differential expression. The DESeq2 package is available at http://www.bioconductor.org/packages/release/bioc/html/DESeq2.html
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47,038 citations
TL;DR: The Spliced Transcripts Alignment to a Reference (STAR) software based on a previously undescribed RNA-seq alignment algorithm that uses sequential maximum mappable seed search in uncompressed suffix arrays followed by seed clustering and stitching procedure outperforms other aligners by a factor of >50 in mapping speed.
Abstract: Motivation Accurate alignment of high-throughput RNA-seq data is a challenging and yet unsolved problem because of the non-contiguous transcript structure, relatively short read lengths and constantly increasing throughput of the sequencing technologies. Currently available RNA-seq aligners suffer from high mapping error rates, low mapping speed, read length limitation and mapping biases. Results To align our large (>80 billon reads) ENCODE Transcriptome RNA-seq dataset, we developed the Spliced Transcripts Alignment to a Reference (STAR) software based on a previously undescribed RNA-seq alignment algorithm that uses sequential maximum mappable seed search in uncompressed suffix arrays followed by seed clustering and stitching procedure. STAR outperforms other aligners by a factor of >50 in mapping speed, aligning to the human genome 550 million 2 × 76 bp paired-end reads per hour on a modest 12-core server, while at the same time improving alignment sensitivity and precision. In addition to unbiased de novo detection of canonical junctions, STAR can discover non-canonical splices and chimeric (fusion) transcripts, and is also capable of mapping full-length RNA sequences. Using Roche 454 sequencing of reverse transcription polymerase chain reaction amplicons, we experimentally validated 1960 novel intergenic splice junctions with an 80-90% success rate, corroborating the high precision of the STAR mapping strategy. Availability and implementation STAR is implemented as a standalone C++ code. STAR is free open source software distributed under GPLv3 license and can be downloaded from http://code.google.com/p/rna-star/.
30,684 citations
TL;DR: This report provides the best available prevalence estimates for the US for osteoarthritis, polymyalgia rheumatica, gout, fibromyalgia, and carpal tunnel syndrome as well as the symptoms of neck and back pain.
Abstract: Objective
To provide a single source for the best available estimates of the US prevalence of and number of individuals affected by osteoarthritis, polymyalgia rheumatica and giant cell arteritis, gout, fibromyalgia, and carpal tunnel syndrome, as well as the symptoms of neck and back pain. A companion article (part I) addresses additional conditions.
4,813 citations
TL;DR: This study developed a cell-based crosslinking approach to determine at high resolution and transcriptome-wide the binding sites of cellular RBPs and miRNPs and revealed that these factors bind thousands of sites containing defined sequence motifs and have distinct preferences for exonic versus intronic or coding versus untranslated transcript regions.
2,730 citations
University of Cincinnati Academic Health Center1, City of Hope National Medical Center2, Sun Yat-sen University3, Howard Hughes Medical Institute4, University of Chicago5, China Medical University (PRC)6, Cincinnati Children's Hospital Medical Center7, Martin Luther University of Halle-Wittenberg8, Zhejiang University9
TL;DR: This work reports the insulin-like growth factor 2 mRNA-binding proteins as a distinct family of m6A readers that target thousands of mRNA transcripts through recognizing the consensus GG(m6A)C sequence, and identifies IGF2BPs as an additional class of N6-methyladenosine (m 6A) reader proteins.
Abstract: N6-methyladenosine (m6A) is the most prevalent modification in eukaryotic messenger RNAs (mRNAs) and is interpreted by its readers, such as YTH domain-containing proteins, to regulate mRNA fate. Here, we report the insulin-like growth factor 2 mRNA-binding proteins (IGF2BPs; including IGF2BP1/2/3) as a distinct family of m6A readers that target thousands of mRNA transcripts through recognizing the consensus GG(m6A)C sequence. In contrast to the mRNA-decay-promoting function of YTH domain-containing family protein 2, IGF2BPs promote the stability and storage of their target mRNAs (for example, MYC) in an m6A-dependent manner under normal and stress conditions and therefore affect gene expression output. Moreover, the K homology domains of IGF2BPs are required for their recognition of m6A and are critical for their oncogenic functions. Thus, our work reveals a different facet of the m6A-reading process that promotes mRNA stability and translation, and highlights the functional importance of IGF2BPs as m6A readers in post-transcriptional gene regulation and cancer biology.
1,373 citations