Transcriptional Target Research Articles

Expanded Gene Regulatory Network Reveals Potential Light-Responsive Transcription Factors and Target Genes in Cordyceps militaris.

Cordyceps militaris, a fungus widely used in traditional Chinese medicine and pharmacology, is recognized for its abundant bioactive compounds, including cordycepin and carotenoids. The growth, development, and metabolite production in various fungi are influenced by the complex interactions between regulatory cascades and light-signaling pathways. However, the mechanisms of gene regulation in response to light exposure in C. militaris remain largely unexplored. This study aimed to identify light-responsive genes and potential transcription factors (TFs) in C. militaris through an integrative transcriptome analysis. To achieve this, we reconstructed an expanded gene regulatory network (eGRN) comprising 507 TFs and 8662 regulated genes using both interolog-based and homolog-based methods to build the protein-protein interaction network. Aspergillus nidulans and Neurospora crassa were chosen as templates due to their relevance as fungal models and the extensive study of their light-responsive mechanisms. By utilizing the eGRN as a framework for comparing transcriptomic responses between light-exposure and dark conditions, we identified five key TFs-homeobox TF (CCM_07504), FlbC (CCM_04849), FlbB (CCM_01128), C6 zinc finger TF (CCM_05172), and mcrA (CCM_06477)-along with ten regulated genes within the light-responsive subnetwork. These TFs and regulated genes are likely crucial for the growth, development, and secondary metabolite production in C. militaris. Moreover, molecular docking analysis revealed that two novel TFs, CCM_05727 and CCM_06992, exhibit strong binding affinities and favorable docking scores with the primary light-responsive protein CmWC-1, suggesting their potential roles in light signaling pathways. This information provides an important functional interactive network for future studies on global transcriptional regulation in C. militaris and related fungi.

A bulge uridine in the HIF2α IRE allows IRP1 but not IRP2 to selectively regulate HIF2α expression and ensuing EPO levels

AbstractIron regulatory proteins (IRP1 and IRP2) play a pivotal role in maintaining cellular iron homeostasis by binding to iron-responsive elements (IREs) of target messenger RNAs and regulating the expression of these iron-related genes. Mice and humans who lack functional IRP1 develop erythrocytosis due to erythropoietin (EPO) overproduction, whereas those who lack IRP2 develop microcytic anemia, believed to result from iron deficiency of erythroblasts. Here, we discovered that IRP2 deficiency reduced the expression of hypoxia-inducible factor 2α (HIF2α) and its transcriptional target, EPO, thereby compromising the stress erythropoiesis response to generate red blood cells upon anemia. The distinct consequences of IRP2 and IRP1 on EPO result from the higher binding affinity of the HIF2α IRE for IRP1 than IRP2. This difference in binding affinity arises from a bulge uridine in the upper stem of HIF2α IRE that impairs the ability of IRP2 to bind the IRE. These results reveal that IRP1 and IRP2 play distinct roles in erythropoiesis and unveil an unsuspected IRE binding preference that contributes to the divergent phenotypes observed in IRP1- and IRP2-deficient mammals.

M6ATM: a deep learning framework for demystifying the m6A epitranscriptome with Nanopore long-read RNA-seq data.

N6-methyladenosine (m6A) is one of the most abundant and well-known modifications in messenger RNAs since its discovery in the 1970s. Recent studies have demonstrated that m6A is involved in various biological processes, such as alternative splicing and RNA degradation, playing an important role in a variety of diseases. To better understand the role of m6A, transcriptome-wide m6A profiling data are indispensable. In recent years, the Oxford Nanopore Technology Direct RNA Sequencing (DRS) platform has shown promise for RNA modification detection based on current disruptions measured in transcripts. However, decoding current intensity data into modification profiles remains a challenging task. Here, we introduce the m6A Transcriptome-wide Mapper (m6ATM), a novel Python-based computational pipeline that applies deep neural networks to predict m6A sites at a single-base resolution using DRS data. The m6ATM model architecture incorporates a WaveNet encoder and a dual-stream multiple-instance learning model to extract features from specific target sites and characterize the m6A epitranscriptome. For validation, m6ATM achieved an accuracy of 80% to 98% across in vitro transcription datasets containing varying m6A modification ratios and outperformed other tools in benchmarking with human cell line data. Moreover, we demonstrated the versatility of m6ATM in providing reliable stoichiometric information and used it to pinpoint PEG10 as a potential m6A target transcript in liver cancer cells. In conclusion, m6ATM is a high-performance m6A detection tool, and our results pave the way for future advancements in epitranscriptomic research.

Chemical modulation of Akt signaling enhances spinal cord regeneration in zebrafish

Central nervous system lesions often cause permanent motility defects in mammals since the injured neurons cannot regenerate. In contrast, lower vertebrates like zebrafish can regenerate lost neurons and restore motor function. This study investigates the efficacy of SC79, a pan-Akt activator, and A674563, a selective Akt1 inhibitor, as potential therapeutic agents for promoting spinal cord recovery post-injury. Spinal cord injury was induced in zebrafish larvae, and the effects of SC79 and A674563 on neuronal and glial regeneration were examined. SC79 promoted neuronal regeneration without affecting glial bridging, while A674563 induced glial bridging but reduced neuronal regeneration. The combination of SC79 and A674563 induced both glial bridging and neuronal regeneration. Optomotor response tests revealed improved motor function recovery with the combined treatment compared to individual treatments. Additionally, these chemical treatments altered the expression of 12 Akt downstream transcriptional target genes, affirming that the combination treatment preferentially regulates spinal cord regeneration through its action on Akt signaling. These findings highlight the complex interplay of Akt signaling pathways in spinal cord regeneration and suggest potential therapeutic strategies for enhancing functional recovery in spinal cord injury patients

Recruitment of the m6A/m6Am demethylase FTO to target RNAs by the telomeric zinc finger protein ZBTB48

BackgroundN6-methyladenosine (m6A), the most abundant internal modification on eukaryotic mRNA, and N6, 2′-O-dimethyladenosine (m6Am), are epitranscriptomic marks that function in multiple aspects of posttranscriptional regulation. Fat mass and obesity-associated protein (FTO) can remove both m6A and m6Am; however, little is known about how FTO achieves its substrate selectivity.ResultsHere, we demonstrate that ZBTB48, a C2H2-zinc finger protein that functions in telomere maintenance, associates with FTO and binds both mRNA and the telomere-associated regulatory RNA TERRA to regulate the functional interactions of FTO with target transcripts. Specifically, depletion of ZBTB48 affects targeting of FTO to sites of m6A/m6Am modification, changes cellular m6A/m6Am levels and, consequently, alters decay rates of target RNAs. ZBTB48 ablation also accelerates growth of HCT-116 colorectal cancer cells and modulates FTO-dependent regulation of Metastasis-associated protein 1 (MTA1) transcripts by controlling the binding to MTA1 mRNA of the m6A reader IGF2BP2.ConclusionsOur findings thus uncover a previously unknown mechanism of posttranscriptional regulation in which ZBTB48 co-ordinates RNA-binding of the m6A/m6Am demethylase FTO to control expression of its target RNAs.

Similarities and distinctions in the activation of the Candida glabrata Pdr1 regulatory pathway by azole and non-azole drugs.

Incidences of fluconazole (FLC) resistance among Candida glabrata clinical isolates is a growing issue in clinics. The pleiotropic drug response (PDR) network in C. glabrata confers azole resistance and is defined primarily by the Zn2Cys6 zinc cluster-containing transcription factor Pdr1 and target genes such as CDR1, that encodes an ATP-binding cassette transporter protein thought to act as a FLC efflux pump. Mutations in the PDR1 gene that render the transcription factor hyperactive are the most common cause of fluconazole resistance among clinical isolates. The phenothiazine class drug fluphenazine and a molecular derivative, CWHM-974, which both exhibit antifungal properties, have been shown to induce the expression of Cdr1 in Candida spp. We have used a firefly luciferase reporter gene driven by the CDR1 promoter to demonstrate two distinct patterns of CDR1 promoter activation kinetics: gradual promoter activation kinetics that occur in response to ergosterol limitations imposed by exposure to azole and polyene class antifungals and a robust and rapid CDR1 induction occurring in response to the stress imposed by fluphenazines. We can attribute these different patterns of CDR1 induction as proceeding through the promoter region of this gene since this is the only segment of the gene included in the luciferase reporter construct. Genetic analysis indicates that the signaling pathways responsible for phenothiazine and azole induction of CDR1 overlap but are not identical. The short time course of phenothiazine induction suggests that these compounds may act more directly on the Pdr1 protein to stimulate its activity.

Germline variant affecting p53β isoforms predisposes to familial cancer

Germline and somatic TP53 variants play a crucial role during tumorigenesis. However, genetic variations that solely affect the alternatively spliced p53 isoforms, p53β and p53γ, are not fully considered in the molecular diagnosis of Li-Fraumeni syndrome and cancer. In our search for additional cancer predisposing variants, we identify a heterozygous stop-lost variant affecting the p53β isoforms (p.*342Serext*17) in four families suspected of an autosomal dominant cancer syndrome with colorectal, breast and papillary thyroid cancers. The stop-lost variant leads to the 17 amino-acid extension of the p53β isoforms, which increases oligomerization to canonical p53α and dysregulates the expression of p53’s transcriptional targets. Our study reveals the capacity of p53β mutants to influence p53 signalling and contribute to the susceptibility of different cancer types. These findings underscore the significance of p53 isoforms and the necessity of comprehensive investigation into the entire TP53 gene in understanding cancer predisposition.

LncRNA-HMG incites colorectal cancer cells to chemoresistance via repressing p53-mediated ferroptosis

Upon chemotherapy, excessive reactive oxygen species (ROS) often lead to the production of massive lipid peroxides in cancer cells and induce cell death, namely ferroptosis. The elimination of ROS is pivotal for tumor cells to escape from ferroptosis and acquire drug resistance. Nevertheless, the precise functions of long non-coding RNAs (lncRNAs) in ROS metabolism and tumor drug-resistance remain elusive. In this study, we identify LncRNA-HMG as a chemoresistance-related lncRNA in colorectal cancer (CRC) by high-throughput screening. Abnormally high expression of LncRNA-HMG predicts poorer prognosis in CRC patients. Concurrently, we found that LncRNA-HMG protects CRC cells from ferroptosis upon chemotherapy, thus enhancing drug resistance of CRC cells. LncRNA-HMG binds to p53 and facilitates MDM2-mediated degradation of p53. Decreased p53 induces upregulation of SLC7A11 and VKORC1L1, which contribute to increase the supply of reducing agents and eliminate excessive ROS. Consequently, CRC cells escape from ferroptosis and acquire chemoresistance. Importantly, inhibition of LncRNA-HMG by anti-sense oligo (ASO) dramatically sensitizes CRC cells to chemotherapy in patient-derived xenograft (PDX) model. LncRNA-HMG is also a transcriptional target of β-catenin/TCF and activated Wnt signals trigger the marked upregulation of LncRNA-HMG. Collectively, these findings demonstrate that LncRNA-HMG promotes CRC chemoresistance and might be a prognostic or therapeutic target for CRC.

The RNA helicase DDX21 activates YAP to promote tumorigenesis and is transcriptionally upregulated by β-catenin in colorectal cancer

The RNA helicase DDX21 is vital for ribosome biogenesis and is upregulated in CRC, but the mechanism by which DDX21 is dysregulated and by which DDX21 promotes tumorigenesis in CRC remains poorly understood. Here, we showed that DDX21 is a direct transcriptional target gene of β-catenin and mediates the protumorigenic function of β-catenin in CRC. DDX21 expression is correlated with the expression and activity of β-catenin, and high DDX21 expression is associated with a poor prognosis in CRC patients. Loss of DDX21 leads to cytoplasmic translocation and decreased transcriptional activity of YAP and suppresses the proliferation and migration of CRC cells, which can be partially rescued by YAP reactivation. Importantly, by using translation elongation inhibitors and DNA intercalators, we showed that ribosomal stress upregulates DDX21 expression and induces the downregulation of LATS and the activation of YAP, probably through the ZAKα-MKK4/7-JNK axis. Overall, our study revealed the transcriptional activation mechanism of DDX21 in CRC and the activation of YAP in the ribosomal stress response, indicating the potential of combination therapy involving the induction of ribosomal stress and YAP inhibition.

The role of circular RNA targeting IGF2BPs in cancer-a potential target for cancer therapy.

Circular RNAs (circRNAs) are an interesting class of conserved single-stranded RNA molecules derived from exon or intron sequences produced by the reverse splicing of precursor mRNA. CircRNAs play important roles as microRNA sponges, gene splicing and transcriptional regulators, RNA-binding protein sponges, and protein/peptide translation factors. Abnormal functions of circRNAs and RBPs in tumor progression have been widely reported. Insulin-like growth factor-2 mRNA-binding proteins (IGF2BPs) are a highly conserved family of RBPs identified in humans that function as post-transcriptional fine-tuners of target transcripts. Emerging evidence suggests that IGF2BPs regulate the processing and metabolism of RNA, including its stability, translation, and localization, and participate in a variety of cellular functions and pathophysiology. In this review, we have summarized the roles and molecular mechanisms of circRNAs and IGF2BPs in cancer development and progression. In addition, we briefly introduce the role of other RNAs and IGF2BPs in cancer, discuss the current clinical applications and challenges faced by circRNAs and IGF2BPs, and propose future directions for this promising research field.

PRDM3/16 regulate chromatin accessibility required for NKX2-1 mediated alveolar epithelial differentiation and function

While the critical role of NKX2-1 and its transcriptional targets in lung morphogenesis and pulmonary epithelial cell differentiation is increasingly known, mechanisms by which chromatin accessibility alters the epigenetic landscape and how NKX2-1 interacts with other co-activators required for alveolar epithelial cell differentiation and function are not well understood. Combined deletion of the histone methyl transferases Prdm3 and Prdm16 in early lung endoderm causes perinatal lethality due to respiratory failure from loss of AT2 cells and the accumulation of partially differentiated AT1 cells. Combination of single-cell RNA-seq, bulk ATAC-seq, and CUT&RUN data demonstrate that PRDM3 and PRDM16 regulate chromatin accessibility at NKX2-1 transcriptional targets critical for perinatal AT2 cell differentiation and surfactant homeostasis. Lineage specific deletion of PRDM3/16 in AT2 cells leads to lineage infidelity, with PRDM3/16 null cells acquiring partial AT1 fate. Together, these data demonstrate that NKX2-1-dependent regulation of alveolar epithelial cell differentiation is mediated by epigenomic modulation via PRDM3/16.

NMDtxDB: data-driven identification and annotation of human NMD target transcripts.

The nonsense-mediated RNA decay (NMD) pathway is a crucial mechanism of mRNA quality control. Current annotations of NMD substrate RNAs are rarely data-driven, but use generally established rules. We present a data set with four cell lines and combinations for SMG5, SMG6, and SMG7 knockdowns or SMG7 knockout. Based on this data set, we implemented a workflow that combines Nanopore and Illumina sequencing to assemble a transcriptome, which is enriched for NMD target transcripts. Moreover, we use coding sequence information (CDS) from Ensembl, Gencode consensus Ribo-seq ORFs, and OpenProt to enhance the CDS annotation of novel transcript isoforms. In summary, 302,889 transcripts were obtained from the transcriptome assembly process, out of which 24% are absent from Ensembl database annotations, 48,213 contain a premature stop codon, and 6433 are significantly upregulated in three or more comparisons of NMD active versus deficient cell lines. We present an in-depth view of these results through the NMDtxDB database, which is available at https://shiny.dieterichlab.org/app/NMDtxDB, and supports the study of NMD-sensitive transcripts. We open sourced our implementation of the respective web-application and analysis workflow at https://github.com/dieterich-lab/NMDtxDB and https://github.com/dieterich-lab/nmd-wf.

Disrupted oxylipin biosynthesis mitigates pathogen infections and pest infestations in cotton (Gossypium hirsutum).

Cotton (Gossypium hirsutum) is the world's most important fiber crop, critical to global textile industries and agricultural economies. However, cotton yield and harvest quality are undermined by the challenges introduced from invading pathogens and pests. Plant-synthesized oxylipins, specifically 9-hydroxy fatty acids resulting from 9-lipoxygenase activity (9-LOX), enhance the growth and development of many microbes and pests. We hypothesized that targeted disruption of 9-LOX-encoding genes in cotton could bolster crop resilience against prominent agronomic threats. Fusarium oxysporum f. sp. vasinfectum (FOV), Aphis gossypii (cotton aphid), and Tobacco rattle virus induced the expression of 9-oxylipin biosynthesis genes, suggesting that the 9-LOX gene products were susceptibility factors to these stressors. Transiently disrupting the expression of the 9-LOX-encoding genes by virus-induced gene silencing significantly reduced target transcript accumulation, and this correlated with impaired progression of FOV infections and a significant decrease in the fecundity of cotton aphids. These findings emphasize that the cotton 9-LOX-derived oxylipins are leveraged by multiple pathogens and pests to enhance their virulence in cotton, and reducing the expression of 9-LOX-encoding genes can benefit cotton crop vitality.

RTP801 interacts with the tRNA ligase complex and dysregulates its RNA ligase activity in Alzheimer's disease.

RTP801/REDD1 is a stress-responsive protein overexpressed in neurodegenerative diseases such as Alzheimer's disease (AD) that contributes to cognitive deficits and neuroinflammation. Here, we found that RTP801 interacts with HSPC117, DDX1 and CGI-99, three members of the tRNA ligase complex (tRNA-LC), which ligates the excised exons of intron-containing tRNAs and the mRNA exons of the transcription factor XBP1 during the unfolded protein response (UPR). We also found that RTP801 modulates the mRNA ligase activity of the complex in vitro since RTP801 knockdown promoted XBP1 splicing and the expression of its transcriptional target, SEC24D. Conversely, RTP801 overexpression inhibited the splicing of XBP1. Similarly, in human AD postmortem hippocampal samples, where RTP801 is upregulated, we found that XBP1 splicing was dramatically decreased. In the 5xFAD mouse model of AD, silencing RTP801 expression in hippocampal neurons promoted Xbp1 splicing and prevented the accumulation of intron-containing pre-tRNAs. Finally, the tRNA-enriched fraction obtained from 5xFAD mice promoted abnormal dendritic arborization in cultured hippocampal neurons, and RTP801 silencing in the source neurons prevented this phenotype. Altogether, these results show that elevated RTP801 impairs RNA processing in vitro and in vivo in the context of AD and suggest that RTP801 inhibition could be a promising therapeutic approach.

Saroglitazar Enhances Memory Functions and Adult Neurogenesis via Up-Regulation of Wnt/β Catenin Signaling in the Rat Model of Dementia.

Peroxisome proliferator-activated receptors (PPARs) have emerged as a promising target for the treatment of various neurodegenerative disorders. Studies have shown that both PPAR α & γ individually modulate various pathophysiological events like neuroinflammation and insulin resistance, which are known to variedly affect neurogenesis. Our study aimed to evaluate the effect of saroglitazar (SGZR), a dual PPAR agonist, on adult neurogenesis and spatial learning and memory, in intracerebroventricular streptozotocin (ICV STZ)-induced dementia in rats. We have found that SGZR at the dose of 4 mg/kg per oral showed significant improvement in learning and memory compared to ICV STZ-treated rats. A substantial increase in neurogenesis was observed in the subventricular zone (SVZ) and the dentate gyrus (DG), as indicated by an increase in the number of 5-bromo-2'-deoxyuridine (BrdU)+ cells, BrdU+ nestin+ cells, and doublecortin (DCX)+cells. Treatment with SGZR also decreased the active form of glycogen synthase kinase 3β (GSK3β) and hence enhanced the nuclear translocation of the β-catenin. Enhanced expression of Wnt transcription factors and target genes indicates that the up-regulation of Wnt signaling might be involved in the observed increase in neurogenesis. Hence, it can be concluded that the SGZR enhances memory functions and adult neurogenesis via the upregulation of Wnt β-catenin signaling in ICV STZ-treated rats.

A comprehensive bioinformatic analysis of the role of TGF-β1-stimulated activating transcription factor 3 by non-coding RNAs during breast cancer progression

A potent growth inhibitor for normal mammary epithelial cells is transforming growth factor beta 1 (TGF-β1). When breast tissues lose the anti-proliferative activity of this factor, invasion and bone metastases increase. Human breast cancer (hBC) cells express more activating transcription factor 3 (ATF3) when exposed to TGF-β1, and this transcription factor is essential for BC development and bone metastases. Non-coding RNAs (ncRNAs), including circular RNAs (circRNAs) and microRNAs (miRNAs), have emerged as key regulators controlling several cellular processes. In hBC cells, TGF-β1 stimulated the expression of hsa-miR-4653–5p that putatively targets ATF3. Bioinformatics analysis predicted that hsa-miR-4653–5p targets several key signaling components and transcription factors, including NFKB1, STAT1, STAT3, NOTCH1, JUN, TCF3, p300, NRF2, SUMO2, and NANOG, suggesting the diversified role of hsa-miR-4653–5p under physiological and pathological conditions. Despite the high abundance of hsa-miR-4653–5p in hBC cells, the ATF3 level remained elevated, indicating other ncRNAs could inhibit hsa-miR-4653–5p’s activity. In silico analysis identified several circRNAs having the binding sites for hsa-miR-4653–5p, indicating the sponging activity of circRNAs towards hsa-miR-4653–5p. The study's findings suggest that TGF-β1 regulates circRNAs and hsa-miR-4653–5p, which in turn affects ATF3 expression, thus influencing BC progression and bone metastasis. Therefore, focusing on the TGF-β1/circRNAs/hsa-miR-4653–5p/ATF3 network could lead to new ways of diagnosing and treating BC.

Elucidating the molecular symphony: unweaving the transcriptional & epigenetic pathways underlying neuroplasticity in opioid dependence and withdrawal.

The persistent use of opioids leads to profound changes in neuroplasticity of the brain, contributing to the emergence and persistence of addiction. However, chronic opioid use disrupts the delicate balance of the reward system in the brain, leading to neuroadaptations that underlie addiction. Chronic cocaine usage leads to synchronized alterations in gene expression, causing modifications in the Nucleus Accumbens (NAc), a vital part of the reward system of the brain. These modifications assist in the development of maladaptive behaviors that resemble addiction. Neuroplasticity in the context of addiction involves changes in synaptic connectivity, neuronal morphology, and molecular signaling pathways. Drug-evoked neuroplasticity in opioid addiction and withdrawal represents a complicated interaction between environmental, genetic, and epigenetic factors. Identifying specific transcriptional and epigenetic targets that can be modulated to restore normal neuroplasticity without disrupting essential physiological processes is a critical consideration. The discussion in this article focuses on the transcriptional aspects of drug-evoked neuroplasticity, emphasizing the role of key transcription factors, including cAMP response element-binding protein (CREB), ΔFosB, NF-kB, Myocyte-enhancing factor 2 (MEF2), Methyl-CpG binding protein 2 (MeCP2), E2F3a, and FOXO3a. These factors regulate gene expression and lead to the neuroadaptive changes observed in addiction and withdrawal. Epigenetic regulation, which involves modifying gene accessibility by controlling these structures, has been identified as a critical component of addiction development. By unraveling these complex molecular processes, this study provides valuable insights that may pave the way for future therapeutic interventions targeting the mechanisms underlying addiction and withdrawal.

Full-length target sequences of GeoMx digital spatial profiling probes reveal that gene-promiscuity predicts probe sensitivity to EDTA tissue decalcification

GeoMx Digital Spatial Profiling (Nanostring) is a commercial spatial transcriptomics method to selectively analyze regions of interest within intact tissue sections. We show that decalcification with ethylene-diamine-tetra-acetic (EDTA) variably attenuates probe counts, while probes that are more resistant to this effect consequently appear overexpressed after quantile normalization. By determining the undisclosed full-length target sequences of probes used in the human whole transcriptome panel, hereby updating target transcripts and genes, we find that the gene-promiscuity of probes is an important factor that determines sensitivity to EDTA incubation.

Bidirectional Polymerization-Transcription Amplification-Encoded Dual-Color Fluorescent Biosensor for Label-Free and Primer-Free Detection of Multiple piRNAs.

PIWI-interacting RNAs (piRNAs) are a type of endogenous noncoding RNAs with a length of 24-31 nucleotides, and they can specifically bind with PIWI proteins to form the piRNA/PIWI complexes for regulating multiple physiological and pathological processes. Herein, we develop a bidirectional polymerization-transcription amplification-encoded dual-color fluorescent biosensor for label-free and primer-free measurements of multiple piRNAs. The designed hairpin probe contains a palindromic tail, and it can serve as the target recognition unit, polymerization primer, and transcription template. In the presence of target piRNAs, the hairpin probes are opened to expose a palindromic sequence that can trigger bidirectional polymerization and transcription reaction with the assistance of KF polymerase and T7 RNA polymerase for the production of numerous RNA aptamers. The aptamers subsequently bind with the corresponding fluorophores (DFHBI-1T/MG) to form the RNA aptamer-fluorophore complexes for the generation of enhanced fluorescence signals. This biosensor can sensitively detect piR-36026 with a limit of detection (LOD) of 82.08 aM and piR-36743 with a LOD of 44.44 aM. Moreover, it can quantify cellular piRNAs with single-cell sensitivity and distinguish cancer cells from normal cells. Furthermore, it has the capability of distinguishing the expression of piRNAs in the tissues of breast cancer patients and healthy individuals. By simply altering the target recognition site of the hairpin probe, this biosensor can be extended to detect various piRNAs, offering a powerful platform for piRNA-related clinical diagnostics and therapeutics.

Sex-Specific Skeletal Muscle Gene Expression Responses to Exercise Reveal Novel Direct Mediators of Insulin Sensitivity Change.

Understanding the causal pathways, systems, and mechanisms through which exercise impacts human health is complex. This study explores molecular signaling related to whole-body insulin sensitivity (Si) by examining changes in skeletal muscle gene expression. The analysis considers differences by biological sex, exercise amount, and exercise intensity to identify potential molecular targets for developing pharmacologic agents that replicate the health benefits of exercise. The study involved 53 participants from the STRRIDE I and II trials who completed eight months of aerobic training. Skeletal muscle gene expression was measured using Affymetrix and Illumina technologies, while pre- and post-training Si was assessed via an intravenous glucose tolerance test. A novel gene discovery protocol, integrating three literature-derived and data-driven modeling strategies, was employed to identify causal pathways and direct causal factors based on differentially expressed transcripts associated with exercise intensity and amount. In women, the transcription factor targets identified were primarily influenced by exercise amount and were generally inhibitory. In contrast, in men, these targets were driven by exercise intensity and were generally activating. Transcription factors such as ATF1, CEBPA, BACH2, and STAT1 were commonly activating in both sexes. Specific transcriptional targets related to exercise-induced Si improvements included TACR3 and TMC7 for intensity-driven effects, and GRIN3B and EIF3B for amount-driven effects. Two key signaling pathways mediating aerobic exercise-induced Si improvements were identified: one centered on estrogen signaling and the other on phorbol ester (PKC) signaling, both converging on the epidermal growth factor receptor (EGFR) and other relevant targets. The signaling pathways mediating Si improvements from aerobic exercise differed by sex and were further distinguished by exercise intensity and amount. Transcriptional adaptations in skeletal muscle related to Si improvements appear to be causally linked to estrogen and PKC signaling, with EGFR and other identified targets emerging as potential skeletal muscle-specific drug targets to mimic the beneficial effects of exercise on Si.