Nuclear retention coupled with sequential polyadenylation dictates post-transcriptional m6A modification in the nucleus.

N6-methyladenosine (m6A) modification is deemed to be co-transcriptionally installed on pre-mRNAs, thereby influencing various downstream RNA metabolism events. However, the causal relationship between m6A modification and RNA processing is often unclear, resulting in premature or even misleading generalizations on the function of m6A modification. Here, we develop 4sU-coupled m6A-level and isoform-characterization sequencing (4sU-m6A-LAIC-seq) and 4sU-GLORI to quantify the m6A levels for both newly synthesized and steady-state RNAs at transcript and single-base-resolution levels, respectively, which enable dissecting the relationship between m6A modification and alternative RNA polyadenylation. Unexpectedly, our results show that many m6A addition events occur post-transcriptionally, especially on transcripts with high m6A levels. Importantly, we find higher m6A levels on shorter 3' UTR isoforms, which likely result from sequential polyadenylation of longer 3' UTR isoforms with prolonged nuclear dwelling time. Therefore, m6A modification can also take place post-transcriptionally to intimately couple with other key RNA metabolism processes to establish and dynamically regulate epi-transcriptomics in mammalian cells.

The polyA tail facilitates splicing of last introns with weak 3' splice sites via PABPN1.

The polyA tail of mRNAs is important for many aspects of RNA metabolism. However, whether and how it regulates pre-mRNA splicing is still unknown. Here, we report that the polyA tail acts as a splicing enhancer for the last intron via the nuclear polyA binding protein PABPN1 in HeLa cells. PABPN1-depletion induces the retention of a group of introns with a weaker 3' splice site, and they show a strong 3'-end bias and mainly locate in nuclear speckles. The polyA tail is essential for PABPN1-enhanced last intron splicing and functions in a length-dependent manner. Tethering PABPN1 to nonpolyadenylated transcripts also promotes splicing, suggesting a direct role for PABPN1 in splicing regulation. Using TurboID-MS, we construct the PABPN1 interactome, including many spliceosomal and RNA-binding proteins. Specifically, PABPN1 can recruit RBM26&27 to promote splicing by interacting with the coiled-coil and RRM domain of RBM27. PABPN1-regulated terminal intron splicing is conserved in mice. Together, our study establishes a novel mode of post-transcriptional splicing regulation via the polyA tail and PABPN1.

Atlas of interactions between SARS-CoV-2 macromolecules and host proteins.

The proteins and RNAs of viruses extensively interact with host proteins after infection. We collected and reanalyzed all available datasets of protein-protein and RNA-protein interactions related to SARS-CoV-2. We investigated the reproducibility of those interactions and made strict filters to identify highly confident interactions. We systematically analyzed the interaction network and identified preferred subcellular localizations of viral proteins, some of which such as ORF8 in ER and ORF7A/B in ER membrane were validated using dual fluorescence imaging. Moreover, we showed that viral proteins frequently interact with host machinery related to protein processing in ER and vesicle-associated processes. Integrating the protein- and RNA-interactomes, we found that SARS-CoV-2 RNA and its N protein closely interacted with stress granules including 40 core factors, of which we specifically validated G3BP1, IGF2BP1, and MOV10 using RIP and Co-IP assays. Combining CRISPR screening results, we further identified 86 antiviral and 62 proviral factors and associated drugs. Using network diffusion, we found additional 44 interacting proteins including two proviral factors previously validated. Furthermore, we showed that this atlas could be applied to identify the complications associated with COVID-19. All data are available in the AIMaP database (https://mvip.whu.edu.cn/aimap/) for users to easily explore the interaction map.

Tetrahydrocurcumin (THC) enhanced the clearance of Cryptococcus deneoformans during infection in vivo.

Cryptococcal species often cause lung infections and are the main cause of fungal meningitis. Claudin-4 appears to be a major structural component that maintains a tight alveolar barrier and prevents fluid and electrolyte leakage into the alveolar space. We aimed to determine whether S7-tetrahydrocurcumin (THC) could clearance of C. deneoformans and regulate claudin-4 expression during C. deneoformans infection. We investigated the effect of THC on C. deneoformans infection and its possible mechanism in vivo. Transmission electron microscopy was used to observe the ultrastructure of the lung tissue and the invasion of Cryptococcus. To clarify the effect of THC, we examined claudin-4, c-Jun, and Smad2 expression. We also measured claudin-4 expression in pulmonary specimens from clinical patients. THC reduced cryptococcal cell invasion in the lungs, improved alveolar exudation, and reduced inflammation. Pretreatment with THC suppressed c-Jun and Smad2 expression, resulting in significantly increased claudin-4 levels. In contrast, the expression of claudin-4 in clinical specimens from patients with cryptococcal infection was higher than that in normal specimens. THC enhanced the clearance of C. deneoformans during infection in vivo. We investigated the expression of claudin-4 and the possible mechanism of THC against C. deneoformans infection.

Recent advances in RNA structurome.

RNA structures are essential to support RNA functions and regulation in various biological processes. Recently, a range of novel technologies have been developed to decode genome-wide RNA structures and novel modes of functionality across a wide range of species. In this review, we summarize key strategies for probing the RNA structurome and discuss the pros and cons of representative technologies. In particular, these new technologies have been applied to dissect the structural landscape of the SARS-CoV-2 RNA genome. We also summarize the functionalities of RNA structures discovered in different regulatory layers-including RNA processing, transport, localization, and mRNA translation-across viruses, bacteria, animals, and plants. We review many versatile RNA structural elements in the context of different physiological and pathological processes (e.g., cell differentiation, stress response, and viral replication). Finally, we discuss future prospects for RNA structural studies to map the RNA structurome at higher resolution and at the single-molecule and single-cell level, and to decipher novel modes of RNA structures and functions for innovative applications.

Alternative polyadenylation by sequential activation of distal and proximal PolyA sites.

Analogous to alternative splicing, alternative polyadenylation (APA) has long been thought to occur independently at proximal and distal polyA sites. Using fractionation-seq, we unexpectedly identified several hundred APA genes in human cells whose distal polyA isoforms are retained in chromatin/nuclear matrix and whose proximal polyA isoforms are released into the cytoplasm. Global metabolic PAS-seq and Nanopore long-read RNA-sequencing provide further evidence that the strong distal polyA sites are processed first and the resulting transcripts are subsequently anchored in chromatin/nuclear matrix to serve as precursors for further processing at proximal polyA sites. Inserting an autocleavable ribozyme between the proximal and distal polyA sites, coupled with a Cleave-seq approach that we describe here, confirms that the distal polyA isoform is indeed the precursor to the proximal polyA isoform. Therefore, unlike alternative splicing, APA sites are recognized independently, and in many cases, in a sequential manner. This provides a versatile strategy to regulate gene expression in mammalian cells.

Circ_0093887 upregulates CCND2 and SUCNR1 to inhibit the ox-LDL-induced endothelial dysfunction in atherosclerosis by functioning as a miR-876-3p sponge.

Circular RNAs (circRNAs) are widely expressed in mammals and act as regulatory targets in the atherogenesis. The objective of this study was to research the biological role and molecular mechanism of circ_0093887 in oxidized low-density lipoprotein (ox-LDL)-induced atherosclerosis (AS) of human aortic endothelial cells (HAECs). Cell viability detection was performed by CCK-8 assay. Inflammatory molecules were examined using ELISA. Flow cytometry was used to measure cell-cycle progression and cell apoptotic rate. Caspase 3 activity was determined using caspase 3 activity assay. The expression levels of circ_0093887, miR-876-3p, CCND2 and SUCNR1 were assayed by quantitative real-time polymerase chain reaction (qRT-PCR). Dual-luciferase reporter, RNA immunoprecipitation (RIP) and RNA pull-down assays were used for the target analysis. EdU assay, wound healing assay/transwell assay and tube formation assay were, respectively, used to assess the effects of circ_0093887/miR-876-3p axis on cell proliferation, migration and tube formation. Oxidized low-density lipoprotein inhibited cell viability and cell-cycle progression but induced the inflammatory response and cell apoptosis. Circ_0093887 was downregulated and miR-876-3p was upregulated in AS patients and ox-LDL-treated HAECs. Functionally, the overexpression of circ_0093887 abrogated the cell injury of HAEC exposed to ox-LDL. For the functional mechanism, we found that circ_0093887 was a sponge for miR-876-3p and miR-876 targeted CCND2 or SUCNR1. The reverted experiment indicated that the function of circ_0093887 was achieved by sponging miR-876-3p. Meanwhile, miR-876-3p inhibitor relieved the inhibitory regulation of circ_0093887 knockdown in cell proliferation, migration and tube formation. Downregulation of miR-876-3p also alleviated the ox-LDL-induced cell injury by upregulating the expression of CCND2 or SUCNR1. Furthermore, circ_0093887 was validated to regulate the levels of CCND2 and SUCNR1 via the sponge effect on miR-876-3p. The protective effects of circ_0093887 on HAECs from ox-LDL were also ​alleviated by repressing the CCND2 and SUCNR1 levels. These findings suggested that circ_0093887 protected HAEC against the ox-LDL-induced inflammatory and apoptotic damages by targeting the miR-876-3p/CCND2 or miR-876/SUCNRA axis. Circ_0093887 could act as a potential therapeutic biomarker for AS patients.

The SARS-CoV-2 subgenome landscape and its novel regulatory features.

Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is currently a global pandemic. CoVs are known to generate negative subgenomes (subgenomic RNAs [sgRNAs]) through transcription-regulating sequence (TRS)-dependent template switching, but the global dynamic landscapes of coronaviral subgenomes and regulatory rules remain unclear. Here, using next-generation sequencing (NGS) short-read and Nanopore long-read poly(A) RNA sequencing in two cell types at multiple time points after infection with SARS-CoV-2, we identified hundreds of template switches and constructed the dynamic landscapes of SARS-CoV-2 subgenomes. Interestingly, template switching could occur in a bidirectional manner, with diverse SARS-CoV-2 subgenomes generated from successive template-switching events. The majority of template switches result from RNA-RNA interactions, including seed and compensatory modes, with terminal pairing status as a key determinant. Two TRS-independent template switch modes are also responsible for subgenome biogenesis. Our findings reveal the subgenome landscape of SARS-CoV-2 and its regulatory features, providing a molecular basis for understanding subgenome biogenesis and developing novel anti-viral strategies.

Downregulation of Uncoupling Protein 2(UCP2) Mediated by MicroRNA-762 Confers Cardioprotection and Participates in the Regulation of Dynamic Mitochondrial Homeostasis of Dynamin Related Protein1 (DRP1) After Myocardial Infarction in Mice.

Acute myocardial infarction (MI) is one of the leading causes of death in the world, and its pathophysiological mechanisms have not been fully elucidated. The purpose of this study was to investigate the role and mechanism of uncoupling protein 2 (UCP2) after MI in mouse heart. Here, we examined the expression and role of UCP2 in mouse heart 4 weeks after MI. The expression of UCP2 was detected by RT-PCR and western blotting. Cardiac function, myocardial fibrosis, and cardiomyocyte apoptosis were assessed by echocardiography and immunohistochemistry. Phosphatase dynamin-related protein1 (P-DRP1) and myocardial fibrosis-related proteins were measured. Cardiomyocytes were exposed to hypoxia for 6 h to mimic the model of MI. Mdivi, an inhibitor of P-DRP1, was used to inhibit DRP1-dependent mitochondrial fission. Mitochondrial superoxide, membrane potential, oxygen consumption rate, and cardiomyocyte apoptosis were detected after hypoxia. It is shown mitochondrial superoxide, membrane potential, oxygen consumption rate, and cardiomyocyte apoptosis were dependent on the level of P-DRP1. UCP2 overexpression reduced cardiomyocyte apoptosis (fibrosis), improved cardiac function and inhibit the phosphorylation of DRP1 and the ratio of P-DRP1/DRP1. However, inhibition of DRP1 by mdivi did not further reduce cell apoptosis rate and cardiac function in UCP2 overexpression group. In addition, bioinformatics analysis, luciferase activity, and western blot assay proved UCP2 was a direct target gene of microRNA-762, a up-regulated microRNA after MI. In conclusion, UCP2 plays a protective role after MI and the mechanism is involved in microRNA-762 upstream and DRP1-dependent mitochondrial fission downstream.

Tranilast prevents doxorubicin-induced myocardial hypertrophy and angiotensin II synthesis in rats.

An increase in oxidative stress is an important pathological mechanism of heart injury induced by doxorubicin (DOX). Tranilast is an anti-allergy drug that has been shown to possess good antioxidant activity in previous studies. The overexpression and secretion of chymase by mast cells (MCs) increase the pathological overexpression of angiotensin II (Ang II), which plays a crucial role in myocardial hypertrophy and the deterioration of heart disease. The MC stabilizer tranilast (N-(3,4-dimethoxycinnamoyl) anthranilic acid; tran) prevents mast cells from degranulating, which may reduce DOX-induced Ang II synthesis. Therefore, in the present study, we hypothesized that tranilast will protect rats from DOX-induced myocardial damage via its antioxidant activity, thereby inhibiting Ang II expression. Thirty male Wistar rats were divided into three groups (n = 10 in each group) that received DOX, a combination of DOX and tranilast or saline (the control group) to test this hypothesis. Tranilast suppressed chymase expression, reduced Ang II levels and prevented the myocardial hypertrophy and the deterioration of heart function induced by DOX. Based on the findings of the present study, the suppression of chymase-dependent Ang-II production and the direct effect of tranilast on the inhibition of apoptosis and fibrosis because of its antioxidant stress capacity may contribute to the protective effect of tranilast against DOX-induced myocardial hypertrophy.

Rotenone and 3-bromopyruvate toxicity impacts electrical and structural cardiac remodeling in rats.

3-Bromopyruvate (3-BrPA) is a promising agent that has been widely studied in the treatment of cancer and pulmonary hypertension. Rotenone is a pesticide commonly used on farms and was shown to have anti-cancer activity and delay fibrosis progression in chronic kidney disease in a recent study. However, there are few studies showing the toxicity of rotenone and 3-BrPA in the myocardium. To support further medical exploration, it is necessary to clarify the side effects of these compounds on the heart. This study was designed to examine the cardiotoxicity of 3-BrPA and rotenone by investigating electrical and structural cardiac remodeling in rats. Forty male rats were divided into 4 groups (n = 10 in each group) and injected intraperitoneally with 3-BrPA, rotenone or a combination of 3-BrPA and rotenone. The ventricular effective refractory period (VERP), corrected QT interval (QTc), and ventricular tachycardia/ventricular fibrillation (VT/VF) inducibility were measured. The expression of Cx43, Kir2.1, Kir6.2, DHPRα1, KCNH2, caspase3, caspase9, Bax, Bcl2, and P53 was detected. Masson's trichrome, TUNEL, HE, and PAS staining and transmission electron microscopy were used to detect pathological and ultrastructural changes. Our results showed that rotenone alone and rotenone combined with 3-BrPA significantly increased the risk of ventricular arrhythmias. Rotenone combined with 3-BrPA caused myocardial apoptosis, and rotenone alone and rotenone combined with 3-BrPA caused electrical and structural cardiac remodeling in rats.

Chronic B-Type Natriuretic Peptide Therapy Prevents Atrial Electrical Remodeling in a Rabbit Model of Atrial Fibrillation.

BACKGROUND: Atrial fibrillation (AF) is an important and growing clinical problem. Current pharmacological treatments are unsatisfactory. Electrical remodeling has been identified as one of the principal pathophysiological mechanisms that promote AF, but there are no effective therapies to prevent or correct electrical remodeling in patients with AF. In AF, cardiac production and circulating levels of B-type natriuretic peptide (BNP) are increased. However, its functional significance in AF remains to be determined. We assessed the hypotheses that chronic BNP treatment may prevent the altered electrophysiology in AF, and preventing AF-induced activation of Ca2+/calmodulin-dependent protein kinase II (CaMKII) may play a role. METHODS AND RESULTS: Forty-four rabbits were randomly divided into sham, rapid atrial pacing (RAP at 600 beats/min for 3 weeks), RAP/BNP, and sham/BNP groups. Rabbits in the RAP/BNP and sham/BNP groups received subcutaneous BNP (20 µg/kg twice daily) during the 3-week study period. HL-1 cells were subjected to rapid field stimulation for 24 hours in the presence or absence of BNP, KN-93 (a CaMKII inhibitor), or KN-92 (a nonactive analog of KN-93). We compared atrial electrical remodeling-related alterations in the ion channel/function/expression of these animals. We found that only in the RAP group, AF inducibility was significantly increased, atrial effective refractory periods and action potential duration were reduced, and the density of ICa, L and Ito decreased, while IK1 increased. The changes in the expressions of Cav1.2, Kv4.3, and Kir2.1 and currents showed a similar trend. In addition, in the RAP group, the activation of CaMKIIδ and phosphorylation of ryanodine receptor 2 and phospholamban significantly increased. Importantly, these changes were prevented in the RAP/BNP group, which were further validated by in vitro studies. CONCLUSIONS: Chronic BNP therapy prevents atrial electrical remodeling in AF. Inhibition of CaMKII activation plays an important role to its anti-AF efficacy in this model.

Radiofrequency ablation for lung squamous cell carcinoma in a single-lung patient: A case report and literature review.

RATIONALE: High morbidity and high mortality are the main features of non-small cell lung cancer (NSCLC). Radiofrequency ablation, which produces a large amount of heat to kill tumor cells, is one effective way to treat this disease. PATIENT CONCERNS: We report the case of a 74-year-old man who presented with a 1-month history of right chest pain. His left lung was removed 12 years prior. Chest computed tomography (CT) revealed a mass in the right lower lobe. DIAGNOSES: An excision biopsy of the mass showed lung squamous cell carcinoma. INTERVENTIONS: We performed radiofrequency ablation. OUTCOMES: The patient underwent 3.5 and 10 months of follow-up, with a partial response and complete remission, respectively. LESSONS: CT-guided radiofrequency ablation is a safe and an effective minimally invasive treatment option. Radiofrequency appears to be a valuable alternative to surgery for inoperable patients presenting with a single-lung NSCLC.

Assessment of aging characteristics of female condylar trabecular structure by cone-beam computed tomography.

OBJECTIVES: This study was performed to analyze the aging-related changes of the female condylar bone mineral density (BMD) and trabecular structure by cone-beam computed tomography (CBCT), and determine whether the condylar structure shows obvious changes after menopause. METHODS: The CBCT images of 160 female patients who met the inclusion criteria for the study were collected and divided into four groups by age (20-29 years, 30-39 years, premenopausal, and postmenopausal groups). Computer processing software CT-Analyser (Version 1.15.2.2+; SkyScan, Antwerp, Belgium) was used to measure the condylar BMD and related indexes, namely the bone volume/tissue volume ratio (BV/TV), trabecular number (Tb.N), trabecular thickness (Tb.Th), trabecular separation (Tb.Sp), trabecular structure model index (SMI), and bone surface area/volume ratio (BS/BV). SPSS 12.0 (SPSS Inc., Chicago, IL, USA) was used to analyze the radiographic findings and statistical differences. RESULTS: No significant differences were found between the bilateral condyles in each group (P > 0.05). BV/TV, Tb.N, and Tb.Th of the condyle decreased with age, and the postmenopausal group showed significantly different values for each index compared with the other groups (P < 0.01). Tb.Sp, SMI, and BS/BV of the condyle increased with age, and the postmenopausal group showed significantly different values for each index compared with the other groups (P < 0.01). CONCLUSIONS: With increasing age, the female condylar bone volume decreases, the Tb.N and Tb.Th decrease, the gap between the trabecular bone increases, and plate-like trabecular bone gradually transforms into a rod-like form. These changes are much more obvious in postmenopausal women.

Caspase-Independent Pathway is Related to Nilotinib Cytotoxicity in Cultured Cardiomyocytes.

BACKGROUND/AIMS: Cardiotoxicity is a predominant side-effect of nilotinib during chronic myeloid leukemia treatment. The underlying molecular mechanism remains unclear. The role of autophagy and mitochondrial signaling was investigated in nilotinib-treated cardiac H9C2 cells. METHODS: Cytotoxicity was assessed using Cell Death Detection kit. Immunoblot and immunofluorescence staining was performed, and cathepsin B and caspase3 activity was assessed in nilotinib-treated H9C2 cells with or without distinct pathway inhibitor or specific siRNA. RESULTS: Nilotinib time- and dose-dependently induced H9C2 apoptosis, which was not completely prevented by the pan caspase inhibitor z-VAD-fmk. Following nilotinib treatment, mitochondrial membrane potential decreased significantly accompanied with remarkable morphological changes. Nuclear translocation of mitochondrial apoptosis inducing factor (AIF) and increased p53 was detected in nilotinib-treated cells. AIF knockdown prevented nilotinib-induced increase of p53 and apoptosis. Additionally, increased cathepsin B activity was detected, and inhibition of cathepsin B by CA-074Me prevented nilotinib-induced apoptosis and nuclear translocation of AIF. Moreover, increased Atg5 and transition of LC3-I to LC3-II was revealed following nilotinib treatment. Increased cathepsin B activity and apoptosis by nilotinib was significantly prohibited by specific autophagy inhibitor bafilomycin A and Atg5 knockdown. CONCLUSION: Our findings demonstrate that nilotinib increases autophagy and cathepsin B activity, leading to mitochondrial AIF release and nuclear translocation, which is responsible for p53 and apoptosis induction in H9C2 cells.

Schisandrin B displays a protective role against primary pulmonary hypertension by targeting transforming growth factor ß1.

Pulmonary arterial smooth muscle cells (PASMCs) in the medial layer of the vessel wall are involved in vessel homeostasis, but also for pathologic vascular remodeling in diverse diseases, such as pulmonary arterial hypertension (PAH). Pulmonary vascular remodeling in PAH results in vascular disorders, but its underlying molecular mechanisms are still not to be fully disclosed. In this study, we investigated the expression and function of the transforming growth factor (TGF)-ß1 in human PASMC cultured under the condition of hypoxia and elucidated the effect of schisandra chinensis and its active ingredients on proliferation, migration, and apoptosis in human PASMCs. We demonstrated that schisandrin B (Sch.B) alleviated the severity of PAH in PASMCs cultured under the condition of hypoxia. Significant upregulation of TGF-ß1 was observed in hypoxia-induced human PASMCs. Interestingly, administration of Sch.B substantially attenuated TGF-ß1 level in these PASMCs. In order to elucidate Sch.B function, the hypoxia-induced human PASMC was stimulated with Sch.B or cotreatment with TGF-ß1 in vitro. In agreement with its TGF-ß1-reducing effect, Sch B relieved human PASMCs migration and promoted the apoptosis of human PASMCs, by activation of TGF-ß1 downstream signal pathways in PASMCs. In contrast, co-treatment with TGF-ß1 promoted human PASMC proliferation and migration and inhibited the apoptosis of human PASMC, which can attenuate the protective role of Sch.B in human PASMC. Taken collectively, these findings suggest that the vascular relaxation evoked by Sch.B was mediated by direct effect on vascular smooth muscle cell via TGF-ß1 downstream signal pathways.

Resveratrol Ameliorates Cardiac Hypertrophy by Down-regulation of miR-155 Through Activation of Breast Cancer Type 1 Susceptibility Protein.

BACKGROUND: The polyphenol resveratrol (Rev) has been reported to exhibit cardioprotective effects, such as inhibition of TAC (transverse aortic constriction) or isoprenaline (ISO)-induced hypertrophy. MicroRNA-155 (miR-155) was found to be decreased in hypertrophic myocardium, which could be further reduced by pretreatment of Rev. The study was designed to investigate the molecular effects of miR-155 on cardiac hypertrophy, focusing on the role of breast cancer type 1 susceptibility protein (BRCA1). METHODS AND RESULTS: We demonstrated that Rev alleviated severity of hypertrophic myocardium in a mice model of cardiac hypertrophy by TAC treatment. Down-regulation of miR-155 was observed in pressure overload- or ISO-induced hypertrophic cardiomyoctyes. Interestingly, administration of Rev substantially attenuated miR-155 level in cardiomyocytes. In agreement with its miR-155 reducing effect, Rev relieved cardiac hypertrophy and restored cardiac function by activation of BRCA1 in cardiomyoctyes. Our results further revealed that forkhead box O3a (FoxO3a) was a miR-155 target in the heart. And miR-155 directly repressed FoxO3a, whose expression was mitigated in miR-155 agomir and mimic treatment in vivo and in vitro. CONCLUSIONS: We conclude that BRCA1 inactivation can increase expression of miR-155, contributing to cardiac hypertrophy. And Rev produces their beneficial effects partially by down-regulating miR-155 expression, which might be a novel strategy for treatment of cardiac hypertrophy.

Long non coding RNA-UCA1 contributes to cardiomyocyte apoptosis by suppression of p27 expression.

BACKGROUND/AIMS: Urothelial carcinoma-associated 1 (UCA1) is a recently identified long non coding RNA (lncRNA). However, few studies have explored its role in cardiomyocytes after focal cardiac ischemia reperfusion injury (CIR). METHODS: Rat CIR models were established using ligation of the Lower Anterior Descending artery (LAD). Cell apoptosis and reactive oxygen species (ROS) production in cardiac tissues were explored using immunohistochemistry and DHE staining. lncRNA expression patterns were detected using microarray and validated by qPCR. Cell viability and apoptosis were examined using MTT assay and flow cytometry. RESULTS: CIR significantly induced cell apoptosis and ROS production in the rat model. The results of microarray demonstrated the reduced expression of UCA1, which was validated by qPCR. Follow-up experiments showed that UCA1 was involved in H2O2-induced cell apoptosis. We further showed that UCA1 negatively correlated with the expression of p27. Moreover, overexpression of p27 could induce primary cardiomyocyte apoptosis. CONCLUSIONS: Reduction of UCA1 levels plays a pro-apoptotic role in primary cardiomyocytes partially through stimulation of p27 protein expression. These results are in agreement with the observed levels of UCA1, p27 and apoptosis after cardiac I/R injury, suggesting that UCA1 might have an important role during I/R injury.

Heterogeneously expressed fezf2 patterns gradient Notch activity in balancing the quiescence, proliferation, and differentiation of adult neural stem cells.

Balancing quiescence, self-renewal, and differentiation in adult stem cells is critical for tissue homeostasis. The underlying mechanisms, however, remain incompletely understood. Here we identify Fezf2 as a novel regulator of fate balance in adult zebrafish dorsal telencephalic neural stem cells (NSCs). Transgenic reporters show intermingled fezf2-GFP(hi) quiescent and fezf2-GFP(lo) proliferative NSCs. Constitutive or conditional impairment of fezf2 activity demonstrates its requirement for maintaining quiescence. Analyses of genetic chimeras reveal a dose-dependent role of fezf2 in NSC activation, suggesting that the difference in fezf2 levels directionally biases fate. Single NSC profiling coupled with genetic analysis further uncovers a fezf2-dependent gradient Notch activity that is high in quiescent and low in proliferative NSCs. Finally, fezf2-GFP(hi) quiescent and fezf2-GFP(lo) proliferative NSCs are observed in postnatal mouse hippocampus, suggesting possible evolutionary conservation. Our results support a model in which fezf2 heterogeneity patterns gradient Notch activity among neighbors that is critical to balance NSC fate.

The development of hippocampal cellular assemblies.

The proper assembly of a cohort of distinct cell types is a prerequisite for building a functional hippocampus. In this review, we describe the major molecular events of the developmental program leading to the cellular construction of the hippocampus. Data from rodent studies are used here to elaborate on our understanding of these processes.