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Maternal diabetes mellitus is among the most frequent environmental contributors to congenital birth defects, including heart defects and craniofacial anomalies, yet the cell types affected and mechanisms of disruption are largely unknown. Using multi-modal single cell analyses, here we show that maternal diabetes affects the epigenomic landscape of specific subsets of cardiac and craniofacial progenitors during embryogenesis. A previously unrecognized cardiac progenitor subpopulation expressing the homeodomain-containing protein ALX3 showed prominent chromatin accessibility changes and acquired a more posterior identity. Similarly, a subpopulation of neural crest-derived cells in the second pharyngeal arch, which contributes to craniofacial structures, displayed abnormalities in the epigenetic landscape and axial patterning defects. Chromatin accessibility changes in both populations were associated with increased retinoic acid signaling, known to establish anterior-posterior identity. This work highlights how an environmental insult can have highly selective epigenomic consequences on discrete cell types leading to developmental patterning defects.

Traditionally, the group 1 intron of the T4 td gene is used to generate a foreign circular sequence. However, the T4 system has been shown to be fairly inefficient in expressing circular RNA (circRNA). Here, a new method is developed to express circular sequences with high circularization efficiency to strengthen the confidence for future circRNA functional studies. CircRNA expression plasmids, constructed with different lengths derived from the actin intron (15‐nt, 30‐nt, 60‐nt, 100‐nt, 180‐nt) and T4 intron, are introduced into human and mouse cell lines 293T and B16. Junction detection and sequencing are used to determine successful circularization of introns and their expression efficiencies. An actin intron with a medium length (60‐nt–100‐nt) shows significantly increased efficiency of circularization, whereas intron‐100‐nt shows the best efficiency in most conditions. RNA pull‐down assays are designed to precipitate the splicing factors that are bound to the introns and intron/exon junction. The precipitated proteins are analyzed by mass spectrometry (MS), aiming to identify the possible underlying mechanism behind the high circularization efficiency. This expression system has been validated using different circRNAs, and such method shows potential in contributing to the expanding field of circRNA studies. [ABSTRACT FROM AUTHOR]

Multiple cancers arise due to aberrations in the wingless integrated (Wnt) signaling pathway. Several miRNAs modulate the integral components of the Wnt signaling pathway. miR-3648 is a human-specific miRNA that is of particular interest due to its minimal off-targeting effect. In this study, we investigated the expression of miR-3648 and APC2 in breast cancer patients of Pakistan. Correlations of miR-3648 and APC2 expression with clinico-pathological features and breast cancer subtypes were observed in tissue samples by means of quantitative real time PCR. Our results showed that miR-3648 was relatively downregulated in Luminal A subtype, with corresponding upregulation of APC2 in these patients. Moreover, the transcript levels of both miR- 3648 and APC2 were found to be inversely regulated in breast cancer women presented with early disease onset, pre-menopause, low tumor grade, early clinical stage, absence of nodal invasion and metastasis, further suggesting the molecular interplay of these molecules in breast cancer development and progression. [ABSTRACT FROM AUTHOR]

Colitis is a chronic bowel disease characterized by damage to the lining of the large intestine, with its precise underlying causes remaining incompletely understood. In this study, we provide evidence that circular RNA circNlgn plays a pivotal role in promoting the development of colitis. Colitis patients produce significant higher levels of circNlgn. Transgenic mice expressing circNlgn exhibit heightened susceptibility to colitis development and progression, primarily attributed to the presence of the protein isoform Nlgn173 encoded by circNlgn. Nlgn173 undergoes translocation into cell nuclei, where it interacts with actin, impeding the binding of actin-related protein 2 and 3 (Arp2/3) complex to actin molecules. Consequently, this leads to a reduction in actin polymerization. Mechanistically, Nlgn173 enhances tyrosine-53 phosphorylation of nuclear actin, diminishing its capacity to interact with the Arp2/3 complex and causing a decrease in filamentous actin levels. These alterations in actin dynamics result in inhibited cell cycle progression, increased apoptosis, and decreased proliferation of colonic epithelial cells, thereby exacerbating colitis development and progression. In contrast, the silencing of circNlgn or the targeted inhibition of Nlgn173 translation and nuclear translocation leads to the promotion of nuclear actin polymerization, enhanced cell survival, and reduced apoptosis and ultimately improves the outcome of colitis in vivo. Interestingly, nuclear actin polymerization is highly related with expression of PIAS3, which modulates signal transducer and activator of transcription 3 and NF-κB activity in colitis. Strategies such as circNlgn knockdown and targeting nuclear actin polymerization of the colonic epithelium may explore a novel avenue for acute ulcerative colitis clinical intervention., Competing Interests: Competing interests: The authors declare that they have no competing interests., (Copyright © 2024 William W. Du et al.)

Recent studies have highlighted the pivotal roles of circular RNAs (circRNAs) in cardiovascular diseases. Through high-throughput circRNA sequencing of both normal myocardial tissues and hypertrophic patients, we unveiled 32,034 previously undiscovered circRNAs with distinct cardiac expression patterns. Notably, circITGa9, a circRNA derived from integrin-α9, exhibited substantial up-regulation in cardiac hypertrophy patients. This elevation was validated across extensive sample pools from cardiac patients and donors. In vivo experiments revealed heightened cardiac fibrosis in mice subjected to transverse aortic constriction (TAC) after circITGa9 injection. We identified circITGa9 binding proteins through circRNA precipitation followed by liquid chromatography tandem-mass spectrometry. Furthermore, circRNA pull-down/precipitation assays demonstrated that increased circITGa9 expression facilitated binding with tropomyosin 3 (TPM3). Specific binding sites between circITGa9 and TPM3 were identified through computational algorithms and further validated by site-directed mutagenesis. We further showed that circITGa9 induced actin polymerization, characteristic of tissue fibrosis. Finally, we developed approaches that improved cardiac function and decreased fibrosis by delivering small interfering RNA targeting circITGa9 or blocking oligo inhibiting the interaction of circITGa9 and TPM3 into TAC mice, which is amenable for further preclinical and translational development. We conclude that elevated circITGa9 levels drive cardiac remodeling and fibrosis. By pinpointing circITGa9 as a therapeutic target, we open doors to innovative interventions for mitigating cardiac remodeling and fibrosis., Competing Interests: Competing interests: The authors declare no conflict of interest for the study., (Copyright © 2024 Feiya Li et al.)