N6-methyladenosine (m6A) is the most abundant internal modification of mRNAs and is implicated in all aspects of post-transcriptional RNA metabolism. However, little is known about m6A modifications ...to circular (circ) RNAs. We developed a computational pipeline (AutoCirc) that, together with depletion of ribosomal RNA and m6A immunoprecipitation, defined thousands of m6A circRNAs with cell-type-specific expression. The presence of m6A circRNAs is corroborated by interaction between circRNAs and YTHDF1/YTHDF2, proteins that read m6A sites in mRNAs, and by reduced m6A levels upon depletion of METTL3, the m6A writer. Despite sharing m6A readers and writers, m6A circRNAs are frequently derived from exons that are not methylated in mRNAs, whereas mRNAs that are methylated on the same exons that compose m6A circRNAs exhibit less stability in a process regulated by YTHDF2. These results expand our understanding of the breadth of m6A modifications and uncover regulation of circRNAs through m6A modification.
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•N6-adenosine methylation (m6A) is widespread in circRNAs•circRNAs exhibit patterns of m6A modifications that are distinct from those of mRNAs•m6A circRNAs are expressed in cell-type-specific patterns•m6A modifications are written and read by the same complexes in circRNAs and mRNAs
Zhou et al. find that N6-adenosine methylation (m6A) is widespread in circular (circ) RNAs and exhibits cell-type-specific patterns of expression. m6A modifications are written and read by the same protein complexes that interact with mRNAs, but many sites of m6A modifications in circRNAs are distinct from those in mRNAs.
Transforming growth factor beta (TGF-β) signaling, mediated through the transcription factors Smad2 and Smad3 (Smad2/3), directs different responses in different cell types. Here we report that Smad3 ...co-occupies the genome with cell-type-specific master transcription factors. Thus, Smad3 occupies the genome with Oct4 in embryonic stem cells (ESCs), Myod1 in myotubes, and PU.1 in pro-B cells. We find that these master transcription factors are required for Smad3 occupancy and that TGF-β signaling largely affects the genes bound by the master transcription factors. Furthermore, we show that induction of Myod1 in nonmuscle cells is sufficient to redirect Smad3 to Myod1 sites. We conclude that cell-type-specific master transcription factors determine the genes bound by Smad2/3 and are thus responsible for orchestrating the cell-type-specific effects of TGF-β signaling.
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► In ESCs, TGF-β effector Smad3 occupies genomic sites bound by master regulator Oct4 ► Oct4 is required for Smad3 occupancy of these genomic sites in ESCs ► Smad3 co-occupies the genome with master transcription factors in multiple cell types ► TGF-β signaling largely affects genes bound by the cell-specific master regulators
Fate-determining transcription factors drive Smad3 to its target genes to control tissue-specific responses to TGF-β signaling.
Many long noncoding RNA (lncRNA) species have been identified in mammalian cells, but the genomic origin and regulation of these molecules in individual cell types is poorly understood. We have ...generated catalogs of lncRNA species expressed in human and murine embryonic stem cells and mapped their genomic origin. A surprisingly large fraction of these transcripts (>60%) originate from divergent transcription at promoters of active protein-coding genes. The divergently transcribed lncRNA/mRNA gene pairs exhibit coordinated changes in transcription when embryonic stem cells are differentiated into endoderm. Our results reveal that transcription of most lncRNA genes is coordinated with transcription of protein-coding genes.
In this trial, patients with atrial fibrillation undergoing mitral-valve surgery were assigned to surgical ablation of AF or no ablation. At 6 and 12 months, more patients in the ablation group were ...free from AF, but more patients in that group required permanent pacemakers.
Atrial fibrillation, which is associated with reduced survival and increased risk of stroke, is present in 30 to 50% of patients presenting for mitral-valve surgery.
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The development of open surgical procedures for the ablation of atrial fibrillation has led to their widespread application during cardiac operations, but their effectiveness and safety have not been rigorously established. It is hypothesized that long-term outcomes can be improved by successful ablation in patients with preexisting persistent or long-standing persistent atrial fibrillation who are undergoing mitral-valve surgery.
The Cox maze III operation (sometimes called the “cut-and-sew” maze operation) is a complex surgical procedure . . .
Cooperation between DNA, RNA and protein regulates gene expression and controls differentiation through interactions that connect regions of nucleic acids and protein domains and through the assembly ...of biomolecular condensates. Here, we report that endoderm differentiation is regulated by the interaction between the long non-coding RNA (lncRNA) DIGIT and the bromodomain and extraterminal domain protein BRD3. BRD3 forms phase-separated condensates of which the formation is promoted by DIGIT, occupies enhancers of endoderm transcription factors and is required for endoderm differentiation. BRD3 binds to histone H3 acetylated at lysine 18 (H3K18ac) in vitro and co-occupies the genome with H3K18ac. DIGIT is also enriched in regions of H3K18ac, and the depletion of DIGIT results in decreased recruitment of BRD3 to these regions. Our findings show that cooperation between DIGIT and BRD3 at regions of H3K18ac regulates the transcription factors that drive endoderm differentiation and suggest that protein-lncRNA phase-separated condensates have a broader role as regulators of transcription.
Fusion of nascent myoblasts to pre-existing myofibres is critical for skeletal muscle growth and repair. The vast majority of molecules known to regulate myoblast fusion are necessary in this ...process. Here, we uncover, through high-throughput in vitro assays and in vivo studies in the chicken embryo, that TGFβ (SMAD2/3-dependent) signalling acts specifically and uniquely as a molecular brake on muscle fusion. While constitutive activation of the pathway arrests fusion, its inhibition leads to a striking over-fusion phenotype. This dynamic control of TGFβ signalling in the embryonic muscle relies on a receptor complementation mechanism, prompted by the merging of myoblasts with myofibres, each carrying one component of the heterodimer receptor complex. The competence of myofibres to fuse is likely restored through endocytic degradation of activated receptors. Altogether, this study shows that muscle fusion relies on TGFβ signalling to regulate its pace.
Recent research on public transport has seen increasing focus on issues like coordination, collaboration and steering in complex governance settings. One of the themes in this field of research is ...related to partnership approaches, as one way of stimulating functioning collaboration between formally independent private and public organisations. The aim of this paper is to explore the role and function of partnerships as a way of supporting well-functioning public transport networks and services in fragmented institutional settings. The empirical focus is on partnerships between operators and public (transport) authorities in two different legal settings: England and Sweden. The analysis is based on interviews with operators and public transport authorities in two metropolitan regions in each country where innovative partnership working has been developed to deal with various types of barriers to delivering better public transport. The results show the key qualities of these partnerships that are required for them to function. Although the regulatory contexts are very different, the partnership qualities are very similar in both cases.
How TGF-β signaling switches from enforcing pluripotency to promoting mesendodermal differentiation remains an open question. Recently in Cell Reports, Beyer et al. demonstrated that Hippo signaling ...components recruit the NuRD complex to repress expression of key genes targeted by TGF-β and thus determine whether TGF-β signaling will favor pluripotency or differentiation.
Osteocytes are terminally differentiated bone cells, derived from osteoblasts, which are vital for the regulation of bone formation and resorption. ECM stiffness and cell seeding density have been ...shown to regulate osteoblast differentiation, but the precise cues that initiate osteoblast–osteocyte differentiation are not yet understood. In this study, we cultured MC3T3-E1 cells on (A) substrates of different chemical compositions and stiffnesses, as well as, (B) substrates of identical chemical composition but different stiffnesses. The effect of cell separation was investigated by seeding cells at different densities on each substrate. Cells were evaluated for morphology, alkaline phosphatase (ALP), matrix mineralisation, osteoblast specific genes (Type 1 collagen, Osteoblast specific factor (OSF-2)), and osteocyte specific proteins (dentin matrix protein 1 (DMP-1), sclerostin (Sost)). We found that osteocyte differentiation (confirmed by dendritic morphology, mineralisation, reduced ALP, Col type 1 and OSF-2 and increased DMP-1 and Sost expression) was significantly increased on soft collagen based substrates, at low seeding densities compared to cells on stiffer substrates or those plated at high seeding density. We propose that the physical nature of the ECM and the necessity for cells to establish a communication network contribute substantially to a concerted shift toward an osteocyte-like phenotype by osteoblasts in vitro.
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•MC3T3-E1 cells were cultured on collagen based substrates of different stiffnesses.•Cells display a dendritic morphology on soft collagen based substrates at low seeding density.•Genes indicating osteocyte development (DMP-1, Sost) upregulated under the same conditions.•The physical nature of the ECM may contribute to osteocyte differentiation.•At low seeding densities, MC3T3-E1 cells extend processes to establish a communication network.
Load-induced fluid flow acts as an important biophysical signal for bone cell mechanotransduction in vivo, where the mechanical environment is thought to be monitored by integrin and primary cilia ...mechanoreceptors on the cell body. However, precisely how integrin- and primary cilia-based mechanosensors interact with the surrounding fluid flow stimulus and ultimately contribute to the biochemical response of bone cells within either the in vitro or in vivo environment remains poorly understood. In this study, we developed fluid–structure interaction models to characterise the deformation of integrin- and primary cilia-based mechanosensors in bone cells under fluid flow stimulation. Under in vitro fluid flow stimulation, these models predicted that integrin attachments on the cell–substrate interface were highly stimulated
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, while the presence of a primary cilium on the cell also resulted in significant strain amplifications, arising at the ciliary base. As such, these mechanosensors likely play a role in mediating bone mechanotransduction in vitro. Under in vivo fluid flow stimulation, integrin attachments along the canalicular wall were highly stimulated and likely play a role in mediating cellular responses in vivo. The role of the primary cilium as a flow sensor in vivo depended upon its configuration within the lacunar cavity. Specifically, our results showed that a short free-standing primary cilium could not effectively fulfil a flow sensing role in vivo. However, a primary cilium that discretely attaches the lacunar wall can be highly stimulated, due to hydrodynamic pressure in the lacunocanalicular system and, as such, could play a role in mediating bone mechanotransduction in vivo.
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