Over 150 types of RNA modifications are identified in RNA molecules. Transcriptome profiling is one of the key steps in decoding the epitranscriptomic panorama of these chemical modifications and ...their potential functions. N
-methylguanosine (m
G) is one of the most abundant modifications present in tRNA, rRNA and mRNA 5'cap, and has critical roles in regulating RNA processing, metabolism and function. Besides its presence at the cap position in mRNAs, m
G is also identified in internal mRNA regions. However, its transcriptome-wide distribution and dynamic regulation within internal mRNA regions remain unknown. Here, we have established m
G individual-nucleotide-resolution cross-linking and immunoprecipitation with sequencing (m
G miCLIP-seq) to specifically detect internal mRNA m
G modification. Using this approach, we revealed that m
G is enriched at the 5'UTR region and AG-rich contexts, a feature that is well-conserved across different human/mouse cell lines and mouse tissues. Strikingly, the internal m
G modification is dynamically regulated under both H
O
and heat shock treatments, with remarkable accumulations in the CDS and 3'UTR regions, and functions in promoting mRNA translation efficiency. Consistently, a PCNA 3'UTR minigene reporter harboring the native m
G modification site displays both enriched m
G modification and increased mRNA translation upon H
O
treatment compared to the m
G site-mutated minigene reporter (G to A). Taken together, our findings unravel the dynamic profiles of internal mRNA m
G methylome and highlight m
G as a novel epitranscriptomic marker with regulatory roles in translation.
CTCF plays key roles in gene regulation, chromatin insulation, imprinting, X chromosome inactivation and organizing the higher-order chromatin architecture of mammalian genomes. Previous studies have ...mainly focused on the roles of the canonical CTCF isoform. Here, we explore the functions of an alternatively spliced human CTCF isoform in which exons 3 and 4 are skipped, producing a shorter isoform (CTCF-s). Functionally, we find that CTCF-s competes with the genome binding of canonical CTCF and binds a similar DNA sequence. CTCF-s binding disrupts CTCF/cohesin binding, alters CTCF-mediated chromatin looping and promotes the activation of IFI6 that leads to apoptosis. This effect is caused by an abnormal long-range interaction at the IFI6 enhancer and promoter. Taken together, this study reveals a non-canonical function for CTCF-s that antagonizes the genomic binding of canonical CTCF and cohesin, and that modulates chromatin looping and causes apoptosis by stimulating IFI6 expression.
5-methylcytosine (m
C) is a post-transcriptional RNA modification identified in both stable and highly abundant tRNAs and rRNAs, and in mRNAs. However, its regulatory role in mRNA metabolism is still ...largely unknown. Here, we reveal that m
C modification is enriched in CG-rich regions and in regions immediately downstream of translation initiation sites and has conserved, tissue-specific and dynamic features across mammalian transcriptomes. Moreover, m
C formation in mRNAs is mainly catalyzed by the RNA methyltransferase NSUN2, and m
C is specifically recognized by the mRNA export adaptor ALYREF as shown by in vitro and in vivo studies. NSUN2 modulates ALYREF's nuclear-cytoplasmic shuttling, RNA-binding affinity and associated mRNA export. Dysregulation of ALYREF-mediated mRNA export upon NSUN2 depletion could be restored by reconstitution of wild-type but not methyltransferase-defective NSUN2. Our study provides comprehensive m
C profiles of mammalian transcriptomes and suggests an essential role for m
C modification in mRNA export and post-transcriptional regulation.
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•ACP was introduced to improve the surface hydrophilic of zein films.•Surface roughness and O- and N- groups were responsible for modification.•ACP improved the cytocompatibility ...C2C12 cells on zein films.
The cell adhesion and proliferation of zein-based biomaterials in cell culture application are limited by the strong hydrophobic surface with low surface energy. In this study, atmospheric cold plasma (ACP) was developed as a modification approach in enhancing the surface hydrophilic and cytocompatibility of zein films. The results indicated that water contact angles decreased from 72.85° (untreated) to 47.43° under the voltage of 100 V. The improvement of the surface free energy (SFE) was mainly attributed to the polar component rather than dispersive component. X-ray photoelectron spectroscopy (XPS) results indicated the change of surface physicochemical properties was mainly due to the partially transformation of long-chain aliphatic hydrocarbons (CH and/or CC) to oxygen- and nitrogen-containing bonds (CO, CO and CN). Furthermore, the content of secondary structure demonstrated β-turn and α-helix were transformed into β-sheet and random coil after ACP treatment. Combining with the cell experiment results, plasma treatment could significantly improve the adhesion rate and proliferation activity of C2C12 cells on zein films. With better cytocompatibility, the potentials of zein in tissue engineering scaffold could be readily exploited.
The role of Fat Mass and Obesity-associated protein （FTO） and its substrate N6-methyladenosine （m6A） in mRNA processing and adipogenesis remains largely unknown. We show that FTO expression and m6A ...levels are inversely correlated during adipogenesis. FTO depletion blocks differentiation and only catalytically active FTO restores adi- pogenesis. Transcriptome analyses in combination with m6A-seq revealed that gene expression and mRNA splicing of grouped genes are regulated by FTO. M6A is enriched in exonic regions flanking 5＇- and 3＇-splice sites, spatially over- lapping with mRNA splicing regulatory serine/arginine-rich （SR） protein exonic splicing enhancer binding regions. Enhanced levels of m6A in response to FTO depletion promotes the RNA binding ability of SRSF2 protein, leading to increased inclusion of target exons. FTO controls exonic splicing of adipogenie regulatory factor RUNX1T1 by regulating m6A levels around splice sites and thereby modulates differentiation. These findings provide compelling evidence that FTO-dependent m6A demethylation functions as a novel regulatory mechanism of RNA processing and plays a critical role in the regulation of adipogenesis.
The detection sensitivity and lift-off impaction of the eddy current sensing are the challenges in the field of electromagnetic nondestructive testing. Higher detection sensitivity, larger lift-off ...capability, and deeper defect detectability remain challenges. This paper presents a double-layer differential planar coil with the discussion of the most sensitive coupling gap. The upper layer contains two symmetrical excitation coils as it can generate symmetrical and opposite magnetic fields. The rectangular structure makes the induced current uniform and symmetrical. The lower layer constructs a differential receiving coil, which can reduce the interference caused by the lift-off impaction as well as the external magnetic field. Different types of surface defects of metallic materials were used for validation. Experimental results show that properly increasing the distance between the excitation and detection layers can effectively improve its sensitivity. Simulation and experimental results confirm that the performance of high sensitivity can be maintained under large lift-off affection.
The 2-hydroxyglutarate (2-HG) has been reported to result from mutations of isocitrate dehydrogenase 1 and 2 (IDH1 and IDH2) genes and to function as an “oncometabolite.” To evaluate the clinical ...significance of serum 2-HG levels in hematologic malignancies, acute myeloid leukemia (AML) in particular, we analyzed this metabolite in distinct types of human leukemia and lymphoma and established the range of serum 2-HG in appropriate normal control individuals by using gas chromatograph–time-of-flight mass spectrometry. Aberrant serum 2-HG pattern was detected in the multicenter group of AML, with 62 of 367 (17%) patients having 2-HG levels above the cutoff value (2.01, log ₂-transformed from 4.03 μg/mL). IDH1/2 mutations occurred in 27 of 31 (87%) AML cases with very high 2-HG, but were observed only in 9 of 31 (29%) patients with moderately high 2-HG, suggesting other genetic or biochemical events may exist in causing 2-HG elevation. Indeed, glutamine-related metabolites exhibited a pattern in favor of 2-HG synthesis in the high 2-HG group. In AML patients with cytogenetically normal AML (n = 234), high 2-HG represented a negative prognostic factor in both overall survival and event-free survival. Univariate and multivariate analyses confirmed high serum 2-HG as a strong prognostic predictor independent of other clinical and molecular features. We also demonstrated distinct gene-expression/DNA methylation profiles in AML blasts with high 2-HG compared with those with normal ones, supporting a role that 2-HG plays in leukemogenesis.
Nonsense‐mediated mRNA decay (NMD) is a post‐transcriptional mechanism that targets aberrant transcripts and regulates the cellular RNA reservoir. Genetic modulation in vertebrates suggests that NMD ...is critical for cellular and tissue homeostasis, although the underlying mechanism remains elusive. Here, we generate knockout mice lacking Smg6/Est1, a key nuclease in NMD and a telomerase cofactor. While the complete loss of Smg6 causes mouse lethality at the blastocyst stage, inducible deletion of Smg6 is compatible with embryonic stem cell (ESC) proliferation despite the absence of telomere maintenance and functional NMD. Differentiation of Smg6‐deficient ESCs is blocked due to sustained expression of pluripotency genes, normally repressed by NMD, and forced down‐regulation of one such target, c‐Myc, relieves the differentiation block. Smg6‐null embryonic fibroblasts are viable as well, but are refractory to cellular reprograming into induced pluripotent stem cells (iPSCs). Finally, depletion of all major NMD factors compromises ESC differentiation, thus identifying NMD as a licensing factor for the switch of cell identity in the process of stem cell differentiation and somatic cell reprograming.
Synopsis
New genetic data show that the NMD pathway is dispensable for growth of embryonic stem cells (ESCs) but essential for differentiation and reprogramming via repression of key pluripotency genes.
The RNA endonuclease Smg6 functions in telomere maintenance and nonsense‐mediated decay (NMD) in mouse ESCs and MEFs.
Loss of Smg6 does not compromise the viability of ESCs and MEFs.
Smg6 regulates ESC differentiation and somatic cellular reprograming via its NMD function, not telomere maintenance.
Smg6 modulates ESC differentiation by controlling mRNA stability of pluripotency genes, for example, c‐Myc.
The NMD pathway acts as a general RNA surveillance mechanism safeguarding the cell identity switch.
New genetic data show that the NMD pathway is dispensable for growth of embryonic stem cells (ESCs) but essential for differentiation and reprogramming via repression of key pluripotency genes.
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