Transcription Process Research Articles

Inhibitory effects of berberine on fungal growth, biofilm formation, virulence, and drug resistance as an antifungal drug and adjuvant with prospects for future applications.

Berberine (BBR), an isoquinoline alkaloid found in medicinal plants such as Coptidis rhizoma, Berberis sp., and Hydrastis canadensis, is a distinctive compound known for its dual ability to exhibit broad-spectrum antifungal activity while offering beneficial effects to the host. These attributes make it a highly valuable candidate for antifungal therapy and as an antibiotic adjuvant. This review provides a comprehensive evaluation of BBR's antifungal properties, focusing on its in vitro and in vivo activity, underlying mechanisms, and its influence on fungal pathogenicity, including virulence, biofilm formation, and resistance. Additionally, the antifungal potential of BBR extracts, derivatives, and nanoformulations is examined in detail. BBR demonstrates fungicidal effects through multiple mechanisms. It targets critical fungal components such as mitochondria, cell membranes, and cell walls, while also inhibiting enzymatic activity and transcription processes. Furthermore, it suppresses the expression of virulence factors, effectively diminishing fungal pathogenicity. Beyond its direct antifungal activity, BBR exerts beneficial effects on the host by modulating gut microbiota, thereby bolstering host defenses against fungal infections and reducing potential adverse effects. BBR's interaction with conventional antifungal drugs presents a unique complexity, particularly in the context of resistance mechanisms. When used in combination therapies, conventional antifungal drugs enhance the intracellular accumulation of BBR, thereby amplifying its antifungal potency as the primary active agent. These synergistic effects position BBR as a promising candidate for combination strategies, especially in addressing drug-resistant fungal infections and persistent biofilms. As antifungal resistance and biofilm-associated infections continue to rise, the multifaceted properties of BBR and its advanced formulations highlight their significant therapeutic potential. However, the scarcity of robust in vivo and clinical studies limits a full understanding of its efficacy and safety profile. To bridge this gap, future investigations should prioritize well-designed in vivo and clinical trials to thoroughly evaluate the therapeutic effectiveness and safety of BBR in diverse clinical settings. This approach could pave the way for its broader application in combating fungal infections.

The 24-kDa subunit of mitochondrial complex I regulates growth, microsclerotia development, stress tolerance, and virulence in Verticillium dahliae

BackgroundThe complete mitochondrial respiratory chain is a precondition for maintaining cellular energy supply, development, and metabolic balance. Due to the evolutionary differentiation of complexes and the semi-autonomy of mitochondria, respiratory chain subunits have become critical targets for crop improvement and fungal control. In fungi, mitochondrial complex I mediates growth and metabolism. However, the role of this complex in the pathogenesis of phytopathogenic fungi is largely unknown.ResultsIn this study, we identified the NADH: ubiquinone oxidoreductase 24-kDa subunit (VdNuo1) of complex in vascular wilt pathogen, Verticillium dahliae, and examined its functional conservation in phytopathogenic fungi. Based on the treatments with respiratory chain inhibitors, the mitochondria-localized VdNuo1 was confirmed to regulate mitochondrial morphogenesis and homeostasis. VdNuo1 was induced during the different developmental stages in V. dahliae, including hyphal growth, conidiation, and melanized microsclerotia development. The VdNuo1 mutants displayed variable sensitivity to stress factors and decreased pathogenicity in multiple hosts, indicating that VdNuo1 is necessary in stress tolerance and full virulence. Comparative transcriptome analysis demonstrated that VdNuo1 mediates global transcriptional effects, including oxidation and reduction processes, fatty acid, sugar, and energy metabolism. These defects are partly attributed to impairments of mitochondrial morphological integrity, complex assembly, and related functions. Its homologue (CgNuo1) functions in the vegetative growth, melanin biosynthesis, and pathogenicity of Colletotrichum gloeosporioides; however, CgNuo1 does not restore the VdNuo1 mutant to normal phenotypes.ConclusionsOur results revealed that VdNuo1 plays important roles in growth, metabolism, microsclerotia development, stress tolerance, and virulence of V. dahliae, sharing novel insight into the function of complex I and a potential fungicide target for pathogenic fungi.

Locking up G‐Quadruplexes with Light‐Triggered Staples Leads to Increased Topological, Thermodynamic, and Metabolic Stability

AbstractG‐quadruplexes (G4 s) are secondary, tetraplexed DNA structures abundant in non‐coding regions of the genome, implicated in gene transcription processes and currently firmly recognised as important potential therapeutic targets. Given their affinity for human proteins, G4 structures are investigated as potential decoys and aptamers. However, G4 s tend to adopt different conformations depending on the exact environmental conditions, and often only one displays the specifically desired biological activity. Their less intensively studied counterparts, the elusive tetraplexed intercalated‐motifs (IMs) are typically unstable at neutral pH, hampering the investigation of their potential involvement in a biological context. We herein report on a photochemical method for “stapling” such tetraplexed‐structures, to increase their stability, lock their topology and enhance their enzymatic resistance, while maintaining biological activity. The chemical structure and topology of the stapled Thrombin Binding Aptamer (TBA) was spectroscopically characterised and rationalised in silico. The method was then extended to other biologically relevant G4‐ and IM‐prone sequences, hinting towards potential application of such stapled structures in a therapeutic context.

Locking up G-Quadruplexes with Light-Triggered Staples Leads to Increased Topological, Thermodynamic, and Metabolic Stability.

G-quadruplexes (G4 s) are secondary, tetraplexed DNA structures abundant in non-coding regions of the genome, implicated in gene transcription processes and currently firmly recognised as important potential therapeutic targets. Given their affinity for human proteins, G4 structures are investigated as potential decoys and aptamers. However, G4 s tend to adopt different conformations depending on the exact environmental conditions, and often only one displays the specifically desired biological activity. Their less intensively studied counterparts, the elusive tetraplexed intercalated-motifs (IMs) are typically unstable at neutral pH, hampering the investigation of their potential involvement in a biological context. We herein report on a photochemical method for "stapling" such tetraplexed-structures, to increase their stability, lock their topology and enhance their enzymatic resistance, while maintaining biological activity. The chemical structure and topology of the stapled Thrombin Binding Aptamer (TBA) was spectroscopically characterised and rationalised in silico. The method was then extended to other biologically relevant G4- and IM-prone sequences, hinting towards potential application of such stapled structures in a therapeutic context.

Role of Protein Ubiquitination and HIF Signaling in the Evolution of Hypoxic Breast Cancer.

Alternations in protein ubiquitination along with hypoxia-inducible factor (HIF) signaling contribute to tumorigenesis and breast tumor advancement. Ubiquitination is an impulsive process, which is coordinately governed by E3 ligases and deubiquitinases (DUBs), that have come out as charismatic therapeutic targets. HIF expression, as well as the transcriptional process in malignancies, are frequently elevated, resulting in pitiable clinical outcomes. According to increasing research, multiple E3 ligases, in addition to UBDs work together to modulate HIF expression and activity, permitting breast cancer cells to make out a hypoxic milieu. On the other hand, hypoxia and HIF signaling regulate numerous E3 ligases as well as DUBs. Interpreting involved networks connecting E3 ligase, DUBS, and HIF will reveal profound mechanisms of physiological response to hypoxia and aid in the discovery of new molecular references for cancer management. The present state of knowledge about the entire kinship among E3 ligase, DUBs, and HIF signaling is reviewed here, emphasizing using E3 ligase or DUB inhibitors in breast cancer.

CHARTING THE UNSEEN: EXPLORING THE EFFECTS OF EPIGENETIC MODIFICATIONS ON SPERM AND THEIR ROLE IN MALE INFERTILITY

Objective: This study aims to review existing literature on the impact of epigenetic modifications, including DNA methylation, histone modifications, and non-coding RNA, on gene expression and their implications for human health and disease mechanisms. Methods: A literature review was conducted between January 2023 and October 2024, analyzing nine selected studies that explored the relationship between epigenetic mechanisms and gene expression in patients with various conditions, such as heart disease and cancer. Key methodologies discussed include RNA sequencing (RNA-Seq) for quantifying gene expression, chromatin immunoprecipitation sequencing (ChIP-Seq) for protein-DNA interaction analysis, and DNA microarray analysis for genome-wide gene expression profiling. Results: The findings highlight that DNA methylation suppresses gene expression by chemically modifying DNA, histone modifications alter chromatin structure to regulate gene accessibility, and non-coding RNAs influence transcription and translation processes. The interplay among these mechanisms was shown to regulate gene expression without requiring genetic mutations, resulting in diverse biological outcomes. The review also discusses the role of daily nutrition, potential complications, and therapeutic strategies targeting epigenetic modifications. Novelty: This study underscores the critical role of epigenetic changes in modulating gene expression and provides insights into their potential for advancing the understanding of disease mechanisms and developing novel approaches for diagnosis, treatment, and prevention.

Community Pharmacists’ Experiences Regarding the Treatment Management of People with Diabetes During Ramadan: A Phenomenological Study

Community pharmacists are often the first point of contact for patients seeking assistance in managing chronic diseases, including diabetes, owing to their accessibility. They are readily available to patients, especially in outpatient settings, and can play a vital role in ensuring safe medication use in patients with diabetes. However, published research on the role of community pharmacists in managing diabetes in patients fasting during Ramadan in Türkiye and worldwide is limited. The aim of this study was to investigate the perspectives and experiences of Turkish community pharmacists in providing treatment management for people with diabetes during Ramadan. To obtain in-depth information about community pharmacists' experiences and perspectives regarding the treatment management of patients with diabetes during Ramadan, a qualitative methodology was chosen for data collection, which could not be obtained using quantitative methods. To gather the views of community pharmacists, a carefully designed semi-structured interview guide was developed to conduct the qualitative interviews. Following the transcription and translation processes, interpretive phenomenological analysis was conducted as an integral part of the data analysis. A total of 21 pharmacists participated in this study. Three main themes, along with their corresponding subthemes, emerged from the dataset: Advice from pharmacists to patients, challenges, and recommendations. The study revealed the professional services pharmacists provided and the barriers they encountered, and documented the recommendations they put forward regarding diabetes treatment management. Pharmacists reported providing diabetes-related services during Ramadan, such as dietary advice, medication adjustments, and dietitian referrals. However they identified several significant challenges, including increased workload, patients' religious beliefs, and limited public awareness of their expertise. To address these, expanding pharmacists' roles, improving communication with patients and physicians, and incorporating Ramadan-specific diabetes management into pharmacy education are seen as key strategies for enhancing care.

KPNB1-ATF4 induces BNIP3-dependent mitophagy to drive odontoblastic differentiation in dental pulp stem cells

BackgroundDifferentiating dental pulp stem cells (DPSCs) into odontoblasts is a critical process for tooth self-repair and dentine‒pulp engineering strategies in the clinic. However, the mechanism underlying the regulation of DPSC odontoblastic differentiation remains largely unknown. Here, we demonstrated that BCL-2 interacting protein 3 (BNIP3)-dependent mitophagy is associated with importin subunit beta-1 (KPNB1)-activating transcription factor 4 (ATF4), which promotes DPSC odontoblastic differentiation.MethodsThe key genes involved in DPSC odontogenic differentiation were identified via bioinformatics. Stable silencing or overexpression of BNIP3 was performed to investigate its impact on DPSC differentiation in vitro (n ≥ 3). To explore the role of BNIP3 in vivo, tooth root fragments loaded with the hydrogel-transfected DPSC complex were implanted into nude mice (n ≥ 6). Dual-luciferase reporter assays and chromatin immunoprecipitation (ChIP) polymerase chain reaction (PCR) were conducted to explore the binding site of ATF4 to the BNIP3 promoter (n ≥ 3). Mitochondrial function experiments were performed to investigate the impact of ATF4-BNIP3 on mitochondria (n ≥ 3). Immunoprecipitation (IP) mass spectrometry (MS) was used to investigate the interaction between ATF4 and its binding protein, KPNB1. Plasmids containing wild-type (WT)/mutant (MUT)-nuclear localization signal (NLS) forms of ATF4 were constructed to determine the specific amino acid residues recognized by KPNB1 and their effects on DPSC odontoblastic differentiation (n ≥ 3).ResultsCompared with those in the control group, the levels of autophagy and mitophagy, especially BNIP3-dependent mitophagy, were greater in the DPSC odontoblastic differentiation group (P < 0.05). Genetic silencing or overexpression of BNIP3 demonstrated that BNIP3 expression was positively correlated with the transition of DPSCs into odontoblasts both in vitro and in vivo (P < 0.05). ATF4 regulates the expression of BNIP3 by directly binding to approximately −1292 to −1279 bp and approximately −1185 to −1172 bp within the BNIP3 promoter region, which is associated with mitophagy and mitochondrial reactive oxygen species (mtROS) levels (P < 0.05). Moreover, ATF4 increased mitophagy, mitochondrial function, and cell differentiation potential via BNIP3 (P < 0.05). Mechanistically, KPNB1 is a novel interacting protein of ATF4 that specifically recognizes amino acids (aa) 280–299 within ATF4 to control its translocation into the nucleus and subsequent transcription and differentiation processes (P < 0.05).ConclusionsWe reported that the critical role of KPNB1/ATF4/BNIP3 axis-dependent mitophagy could provide new cues for the regeneration of the dental pulp‒dentin complex in DPSCs.Graphical

Full-length transcriptome sequencing of the longissimus dorsi muscle of yak and cattle-yak using nanopore technology

Short-read RNA sequencing has been used to sequence the transcriptome of the skeletal muscle of yak and cattle-yak; however, full-length transcripts cannot be obtained and alternative splicing (AS) events cannot be inferred using this sequencing approach. Here, we used Oxford Nanopore Technologies (ONT) full-length sequencing to sequence the transcriptome of the longissimus dorsi of yak and cattle-yak. A total of 20,323 novel genes and 172,870 novel transcripts were identified, and 159,700 novel transcripts were successfully annotated. A total of 157,812 AS events, 58,073 simple sequence repeats, 57,468 complete open reading frames, 2296 transcription factors, and 20,404 lncRNAs were detected. Differentially expressed transcripts (DETs) in the longissimus dorsi muscle of yak and cattle-yak were involved in the MAPK and JAK-STAT signaling pathways related to muscle development and growth. Protein–protein interaction analysis of DETs suggested that TNNI2 might make a major contribution to differences in muscle growth and meat quality traits between yak and cattle-yak. The results have enriched the transcriptome data of dorsal muscles, providing new ideas for the study of transcriptional regulation processes, and also providing useful information for the production of higher yields of yak meat.

The function of the ZFP189 transcription factor in the nucleus accumbens facilitates cocaine-specific transcriptional and behavioral adaptations.

Distinguishing the brain mechanisms affected by distinct addictive drugs may inform targeted therapies against specific substance use disorders (SUDs). Here, we explore the function of a drug-associated, transcriptionally repressive transcription factor (TF), ZFP189, whose expression in the nucleus accumbens (NAc) facilitates cocaine-induced molecular and behavioral adaptations. To uncover the necessity of ZFP189-mediated transcriptional control in driving cocaine-induced behaviors, we created synthetic ZFP189 TFs of distinct transcriptional function, including ZFP189VPR, which activates the expression of target genes and exerts opposite transcriptional control to the endogenously repressive ZFP189. By virally delivering synthetic ZFP189 TFs to the NAc of mice, we discover that the transcriptional control exerted by synthetic or endogenous ZFP189 solely alters behavioral adaptations to cocaine but not morphine, saline, or sucrose. Further, these synthetic ZFP189 TFs are only capable of producing gene-expression changes in rodents exposed to cocaine, but not morphine or saline. In these cocaine exposed mice, the gene-expression profile produced by ZFP189VPR is inversely related to the cocaine-induced transcriptional response, as characterized by Upstream Regulator Analysis in Ingenuity Pathway Analysis. Lastly, we demonstrate that NAc ZFP189WT increases vulnerability to cocaine reinforcement through selective sensitization to the reinforcing effects of small cocaine doses. In contrast, ZFP189VPR treated mice do not experience changes in cocaine sensitivity and had lower rates of cocaine self-administration. Collectively, this research describes the brain mechanisms by which a TF specifically coordinates the molecular adaptations that produce increased cocaine addiction-like behaviors. The use of synthetic ZFP189VPR uncovers novel strategies for therapeutic interventions to potentially halt these cocaine-induced transcriptional processes.

The interaction of BDNF with estrogen in the development of hypertension and obesity, particularly during menopause.

The expression of BDNF in both neuronal and non-neuronal cells is influenced by various stimuli, including prenatal developmental factors and postnatal conditions such as estrogens, dietary habits, and lifestyle factors like obesity, blood pressure, and aging. Central BDNF plays a crucial role in modulating how target tissues respond to these stimuli, influencing the pathogenesis of hypertension, mitigating obesity, and protecting neurons from aging. Thus, BDNF serves as a dynamic mediator of environmental influences, reflecting an individual's unique history of exposure. Estrogens, on the other hand, regulate various processes to maintain overall physiological well-being. Through nuclear estrogen receptors (ERα, ERβ) and the membrane estrogen receptor (GPER1), estrogens modulate transcriptional processes and signaling events that regulate the expression of target genes, such as ERα, components of the renin-angiotensin system (RAS), and hormone-sensitive lipase. Estrogens are instrumental in maintaining the set point for blood pressure and energy balance. BDNF and estrogens work cooperatively to prevent obesity by favoring lipolysis, and counteractively regulate blood pressure to adapt to the environment. Estrogen deficiency leads to menopause in women with low central BDNF level. This review delves into the complex mechanisms involving BDNF and estrogen, especially in the context of hypertension and obesity, particularly among postmenopausal women. The insights gained aim to inform the development of comprehensive therapeutic strategies for these prevalent syndromes affecting approximately 68% of adults.

Clues to transcription/replication collision-induced DNA damage: it was RNAP, in the chromosome, with the fork.

DNA replication and RNA transcription processes compete for the same DNA template and, thus, frequently collide. These transcription-replication collisions are thought to lead to genomic instability, which places a selective pressure on organisms to avoid them. Here, we review the predisposing causes, molecular mechanisms, and downstream consequences of transcription-replication collisions (TRCs) with a strong emphasis on prokaryotic model systems, before contrasting prokaryotic findings with cases in eukaryotic systems. Current research points to genomic structure as the primary determinant of steady-state TRC levels and RNA polymerase regulation as the primary inducer of excess TRCs. We review the proposed mechanisms of TRC-induced DNA damage, attempting to clarify their mechanistic requirements. Finally, we discuss what drives genomes to select against TRCs.

RNA-Binding Protein Hnrnpa1 Triggers Daughter Cardiomyocyte Formation by Promoting Cardiomyocyte Dedifferentiation and Cell Cycle Activity in a Post-Transcriptional Manner.

Stimulating cardiomyocyte (CM) dedifferentiation and cell cycle activity (DACCA) is essential for triggering daughter CM formation. In addition to transcriptional processes, RNA-binding proteins (RBPs) are emerging as crucial post-transcriptional players in regulating CM DACCA. However, whether post-transcriptional regulation of CM DACCA by RBPs could effectively trigger daughter CM formation remains unknown. By performing integrated bioinformatic analysis of snRNA-seq data from neonatal and adult hearts, this study identified Hnrnpa1 as a potential RBP regulating CM DACCA. Hnrnpa1 expression decreased significantly during postnatal heart development. With the use of α-MHC-H2B-mCh/CAG-eGFP-anillin transgenic mice, Hnrnpa1 overexpression promoted CM DACCA, thereby triggering daughter CM formation and enhancing cardiac repair after myocardial infarction (MI). In contrast, CRISPR/Cas9 technology is used to generate CM-specific Hnrnpa1 knockout mice. Hnrnpa1 knockout inhibited cardiac regeneration and worsened cardiac function in the neonatal MI model. Nanopore RNA sequencing, RIP assay, IP-MS, MeRIP-qPCR, PAR-CLIP and luciferase reporter experiments showed that Hnrnpa1 induced Mettl3 post-transcriptional splicing to inhibit m6A-dependent Pbx1 and E2F1 degradation, thereby increasing Runx1, Ccne1, Cdk2 and Ccnb2 expression to promote CM DACCA. In conclusion, Hnrnpa1 triggered daughter CM formation by promoting CM DACCA in a post-transcriptional manner, indicating that Hnrnpa1 might serve as a promising target in cardiac repair post-MI.

Cyclin‐dependent kinase 13 is indispensable for normal mouse heart development

AbstractCongenital heart disease (CHD) has an incidence of approximately 1%. Over the last decade, sequencing studies including large cohorts of individuals with CHD have begun to unravel the genetic mechanisms underpinning CHD. This includes the identification of variants in cyclin‐dependent kinase 13 (CDK13), in individuals with syndromic CHD. CDK13 encodes a serine/threonine protein kinase. The cyclin partner of CDK13 is cyclin K; this complex is thought to be important in transcription and RNA processing. Pathogenic variants in CDK13 cause CDK13‐related disorder in humans, characterised by intellectual disability and developmental delay, recognisable facial features, feeding difficulties and structural brain defects, with 35% of individuals having CHD. To obtain a greater understanding for the role that this essential protein kinase plays in embryonic heart development, we have analysed a presumed loss of function Cdk13 transgenic mouse model (Cdk13tm1b). The homozygous mutants were embryonically lethal in most cases by E15.5. X‐gal staining showed Cdk13 expression localised to developing facial regions, heart and surrounding areas at E10.5, whereas at E12.5, it was more widely present. In the E15.5 heart, staining was seen throughout. RT‐qPCR showed significant reduction in Cdk13 transcript expression in homozygous compared with WT and heterozygous hearts at E10.5 and E12.5. Detailed morphological 3D analysis of embryonic and postnatal hearts was performed using high‐resolution episcopic microscopy, which affords a more detailed analysis of structures such as cardiac valve leaflets and endocardial cushions, compared with more traditional histological techniques. We show that both the homozygous and heterozygous Cdk13tm1b mutants exhibit a range of CHD, including ventricular septal defects, bicuspid aortic valve, double outlet right ventricle and atrioventricular septal defects. 100% (n = 4) of homozygous hearts displayed CHD. Differential expression was seen in Cdk13tm1b homozygous mutants for two genes known to be necessary for normal heart development. The types of defects, and the presence of CHD in heterozygous mice (17.02%, n = 8/47), are consistent with the CDK13‐related disorder phenotype in humans. This study provides important insights into the effects of reduced function of CDK13 in the mouse heart and contributes to our understanding of the mechanism behind this disorder as a cause of CHD.

Novel CRITR-seq approach reveals influenza transcription is modulated by NELF and is a key event precipitating an interferon response.

Transcription of interferons upon viral infection is critical for cell-intrinsic innate immunity. This process is influenced by many host and viral factors. To identify host factors that modulate interferon induction within cells infected by influenza A virus, we developed CRISPR with Transcriptional Readout (CRITR-seq). CRITR-seq is a method linking CRISPR guide sequence to activity at a promoter of interest. Employing this method, we find that depletion of the Negative Elongation Factor complex increases both flu transcription and interferon expression. We find that the process of flu transcription, both in the presence and absence of viral replication, is a key contributor to interferon induction. Taken together, our findings highlight innate immune ligand concentration as a limiting factor in triggering an interferon response, identify NELF as an important interface with the flu life cycle, and validate CRITR-seq as a tool for genome-wide screens for phenotypes of gene expression.

Kinetic Features of Degradation of R-Loops by RNase H1 from Escherichia coli.

R-loops can act as replication fork barriers, creating transcription-replication collisions and inducing replication stress by arresting DNA synthesis, thereby possibly causing aberrant processing and the formation of DNA strand breaks. RNase H1 (RH1) is one of the enzymes that participates in R-loop degradation by cleaving the RNA strand within a hybrid RNA-DNA duplex. In this study, the kinetic features of the interaction of RH1 from Escherichia coli with R-loops of various structures were investigated. It was found that the values of the dissociation constants Kd were minimal for complexes of RH1 with model R-loops containing a 10-11-nt RNA-DNA hybrid part, indicating effective binding. Analysis of the kinetics of RNA degradation in the R-loops by RH1 revealed that the rate-limiting step of the process was catalytic-complex formation. In the presence of RNA polymerase, the R-loops containing a ≤16-nt RNA-DNA hybrid part were efficiently protected from cleavage by RH1. In contrast, R-loops containing longer RNA-DNA hybrid parts, as a model of an abnormal transcription process, were not protected by RNA polymerase and were effectively digested by RH1.

Novel insights into RNA polymerase II transcription regulation: transcription factors, phase separation, and their roles in cardiovascular diseases.

Transcription factors (TFs) are specialized proteins that bind DNA in a sequence-specific manner and modulate RNA polymerase II (Pol II) in multiple steps of the transcription process. Phase separation is a spontaneous or driven process that can form membrane-less organelles called condensates. By creating different liquid phases at active transcription sites, the formation of transcription condensates can reduce the water content of the condensate and lower the dielectric constant in biological systems, which in turn alters the structure and function of proteins and nucleic acids in the condensate. In RNA Pol II transcription, phase separation formation shortens the time at which TFs bind to target DNA sites and promotes transcriptional bursting. RNA Pol II transcription is engaged in developing several diseases, such as cardiovascular disease, by regulating different TFs and mediating the occurrence of phase separation. This review aims to summarize the advances in the molecular mechanisms of RNA Pol II transcriptional regulation, in particular the effect of TFs and phase separation. The role of RNA Pol II transcriptional regulation in cardiovascular disease will be elucidated, providing potential therapeutic targets for the management and treatment of cardiovascular disease.

Role and Interplay of Different Signaling Pathways Involved in Sciatic Nerve Regeneration.

Regeneration of the sciatic nerve is a sophisticated process that involves the interplay of several signaling pathways that orchestrate the cellular responses critical to regeneration. Among the key pathways are the mitogen-activated protein kinase (MAPK), phosphatidylinositol 3-kinase (PI3K)/AKT, cyclic adenosine monophosphate (cAMP), and Janus kinase/signal transducers and transcription activators (JAK/STAT) pathways. In particular, the cAMP pathway modulates neuronal survival and axonal regrowth. It influences various cellular behaviors and gene expression that are essential for nerve regeneration. MAPK is indispensable for Schwann cell differentiation and myelination, whereas PI3K/AKT is integral to the transcription, translation, and cell survival processes that are vital for nerve regeneration. Furthermore, GTP-binding proteins, including those of the Ras homolog gene family (Rho), regulate neural cell adhesion, migration, and survival. Notch signaling also appears to be effective in the early stages of nerve regeneration and in preventing skeletal muscle fibrosis after injury. Understanding the intricate mechanisms and interactions of these pathways is vital for the development of effective therapeutic strategies for sciatic nerve injuries. This review underscores the need for further research to fill existing knowledge gaps and improve therapeutic outcomes.

Exploring lncRNA expression in follicular fluid exosomes of patients with obesity and polycystic ovary syndrome based on high-throughput sequencing technology

BackgroundInfertility is a reproductive health problem that attracts worldwide attention. Polycystic ovary syndrome (PCOS) is a major cause of female infertility and patients with obesity and PCOS are particularly common in clinical practice. Long non-coding RNA (lncRNAs) are a functional core in cells that regulate gene expression, transcription, and chromatin modification processes, and participate in epigenetics, cell cycle, and cell differentiation. LncRNAs are assumed to play a role in the occurrence and development of PCOS; however, their specific mechanism of action remains to be elucidated.MethodsHigh-throughput sequencing technology has been used to sequence and analyze lncRNAs in exosomes from the follicular fluid of patients with obesity and PCOS and those who underwent assisted reproductive therapy owing to male factors. Specific expression profiles of patients with obesity and PCOS were obtained and functional information analysis combined with a literature review were performed to screen for differentially expressed lncRNAs, which were validated using real-time reverse transcription quantitative polymerase chain reaction (RT-qPCR).ResultsHigh-throughput sequencing analysis revealed that compared to normal patients with male infertility, patients with obesity and PCOS had a total of 20 lncRNAs with significant expression differences in follicular fluid exosomes. Among them, 17 lncRNAs were upregulated and three were downregulated. Functional analysis showed that differentially expressed genes were mainly enriched in “cell metabolism,” “cell adhesion,” and other aspects: related gene pathways mainly involved Huntington’s disease, Parkinson’s disease, spliceosomes, non-alcoholic fatty liver disease, and ribosomes. Verification of differentially expressed lncRNAs revealed that the expression of lncRNAs TPT1-AS1, PTOV1-AS1, PTPRG-AS1, and SNHG14 in follicular fluid exosomes was consistent with the sequencing results.ConclusionA preliminary differential expression profile of lncRNAs in exosomes of patients with obesity and PCOS was established by transcriptomic analysis of these individuals. Our bioinformatics analysis results may be applicable to further study of the impact mechanism involving obesity and PCOS. These differentially expressed lncRNAs maybe served as potential biomarkers for in-depth studies of the occurrence, development on Follicle quality and function for patients with PCOS in the future.

TMIC-50. IMPACT OF ZOTIRACICLIB ON GLIOMA TUMOR MICROENVIRONMENT

Abstract BACKGROUND Zotiraciclib, an orally administered multi-kinase inhibitor, has shown anti-glioma effect via suppression of transcriptional process and mitochondrial function. Here, we investigate the impact of zotiraciclib on the glioma immune microenvironment. METHODS Mouse glioma was generated by intracranial injection of mouse syngeneic cells (IDH1-wildtype NPA cells or IDH1R132H-mutant NPAI cells) in C57BL/6 mice. Zotiraciclib was administered (30 mg/kg, intraperitoneal injection, twice per week) starting seven days after tumor implantation. NPA and NPAI tumors were dissected after 10 and 22 days’ treatment, respectively. Immune cells in the tumor tissue were isolated and examined using flow cytometry. Primary human CD8+ cells and CD14+ monocytes isolated from the blood of a healthy donor were used to further investigate the effect of zotiraciclib (15 nM and 50 nM) on human cytotoxic T cell activation and immunosuppressive M2 macrophage polarization, respectively, using flow cytometry. RESULTS No significant difference in the abundance of T cells, myeloid cells, NK cells, NKT cells, DCs, and B cells between zotiraciclib-treated and vehicle-treated groups was detected in neither NPA nor NPAI tumors. The treatment-induced change in the abundance of cytotoxic T cells, helper T cells, macrophages, monocytes, neutrophils, M-MDSCs, PMN-MDSCs, Tregs, and M2 macrophages wasn’t observed. Zotiraciclib reduced the expansion of human CD8+ cytotoxic T cells, but did not affect the expression of CD25, HLA-DR, TNFα and IFNγ after activation in vitro. The viability of human M2 macrophages during differentiation and polarization was not affected, while the expression of CD163 and CD206 was reduced by zotiraciclib. CONCLUSIONS The immunophenotyping performed for this study demonstrates that zotiraciclib does not alter the mouse glioma immune microenvironment. Complementary studies using human immune cells shows that zotiraciclib does not suppress the activation of cytotoxic T cells but reduces the polarization of immunosuppressive M2 macrophages, supporting combining zotiraciclib and immune stimulatory approach in glioma treatment.