Gene

BackgroundHeterochromatic long arm of mouse Y chromosome harbors the multicopy species-specific sequences Ssty, Sly, Asty and Orly that are transcribed in testis and have known functions in male fertility. Of these Ssty and Sly encode proteins - yet all the transcripts are not translated. To investigate the roles of these Y-heterochromatic transcripts further, we analyzed them. MethodsMice with 2/3rd deletion of the Y-chromosome (XYRIIIqdel) and its wild type (XYRIII) were used in this study. Bioinformatic approaches, small RNA northern blots, Electrophoretic Mobility Shift Assays, Luciferase reporter assays, dPCR analysis, RT-qPCR assays and western blotting techniques were used to identify piRNAs that regulate autosomal genes. ResultsWe demonstrate that the multicopy gene families from mouse Y-long arm generate piRNAs predominantly in testis. We observed sequences homologous to these piRNAs in the UTRs of a few autosomal genes, which are differentially expressed in the sperms of XYRIIIqdel mice. Furthermore, the Endogenous Retrovirus Element (ERV) LTR, found in the Orly1 transcript identified piRNAs in the database, showed homology to UTRs and associated genomic regions of a few autosomal genes. Orly1 showed a reduction in genomic copy number by digital PCR in XYRIIIqdel mice. One of the four autosomal genes containing the ERV segment in their UTRs, showed a differential testicular protein expression in the mutant mice. ConclusionsThus, we further elucidate that different classes of repeats from Y-chromosome regulate autosomal gene expression via piRNAs. Besides, this study also identified novel roles for a Y-derived ERV in autosomal gene regulation in testis.

In wheat, weak seed dormancy (SD) is related to an increased tendency for pre-harvest sprouting (PHS), which reduces yield and quality. However, the molecular mechanism underlying SD remains elusive. Here, we identified a wheat R2R3-MYB transcription factor (TaMYB83-7B) related to SD. Expression analysis showed that TaMYB83-7B was highly expressed in wheat seeds, and was more highly expressed in strong-dormancy varieties than in weak-dormancy varieties. Sequence and association analysis indicated that T/C mutations at -907 bp and -1133 bp in the TaMYB83-7B promoter were significantly associated with wheat SD, with C at both sites related to strong dormancy. Dual-luciferase reporter assays demonstrated that the transcriptional activity of the TaMYB83-7B promoter was significantly higher in strong-dormancy varieties than in weak-dormancy varieties. Further analyses indicated that TaMYB83-7B functions as a transcriptional inhibitor. Germination experiments revealed that overexpression of TaMYB83-7B significantly enhanced SD, while its loss-of-function reduced SD. Finally, TaMYB83-7B was found to regulate SD by influencing the balance between abscisic acid (ABA) and gibberellin (GA) in wheat seeds. Overall, the results of this study enhance our understanding of the complex regulatory mechanism underlying SD, and provide gene targets and molecular markers for the genetic improvement of PHS resistance in wheat.

Embryo implantation is early and complex stage of pregnancy begins when competent blastocyst makes a physiological attachment to receptive endometrium. Expression of numerous molecules are essential for initiation of pregnancy. leukemia inhibitory factor (LIF) is essential cytokines required for priming uterus to make it receptive for implantation. In mice, the ovarian estrogen regulated expression of LIF is absolutely required for implantation. Golden hamster showed ovarian estrogen independent process of embryo implantation. Hence, the regulation of LIF in uterus of golden hamster during early pregnancy is still ambiguous. In this study, we explored the possible regulation of LIF by uterine factor and their spatio-temporal localization and expression in the uterus of golden hamster during early pregnancy and pseudopregnancy. We further demonstrated their ability to activate prostaglandin synthesizing enzymes to achieve successful pregnancy. We used immunohistochemistry, quantitative and semiquantitative PCR to achieve the objectives. We observed the expression of LIF in all the day of early pregnancy and pseudopregnancy in the uterus of hamster. Their m-RNA was found to be upregulated around the day of implantation and decidualization. LIF showed high expression in D3 pseudopregnancy. LIF was found to be regulated by estrogen in ovariectomized uterus and significantly reduced expression of LIF was observed in letrozole treated uterine horn. Downregulated expression of prostaglandin synthesizing enzymes was observed in anti-LIF antibody treated uterus. Together, these findings highlights that uterine factor regulated LIF mediate their action via activating prostaglandin synthesizing enzymes to make uterus receptive for successful early pregnancy in hamster. HighlightO_LIExpression of LIF in uterus during pregnancy in golden hamster is independent from the presence of blastocyst C_LIO_LILIF is regulated by estrogen in ovariectomized hamster C_LIO_LIExpression of LIF mRNA is downregulated in letrozole treated uterine horn in day 5 of pregnancy indicating the possibility of their regulation by uterine estrogen in golden hamster C_LIO_LIProstaglandin synthesizing enzyme and LIF might be associated with the activation of inflammatory signals which are essential for successful establishment of early pregnancy in golden hamster. C_LI

Voltage-gated potassium channels (Kv) are a large family of potassium channels composed of 40 members across 12 subtypes. The KCNH genes encode 3 subfamilies of voltage-gated potassium channels: Kv10 (EAG, ether a go go), Kv11 (ERG, EAG-related gene), and Kv12 (ELK, EAG-like K). Kv channels play prominent roles in the neuronal and cardiovascular systems. Mutations in Kv channels have been linked to many human diseases, such as epilepsy, heart arrhythmias, and cancers. Significant progress has been made in understanding protein structures, physiological functions, and the pharmacological modifiers. However, the evolutionary history and gene expression of vertebrate KCNH genes during embryonic development remain largely unknown. We systematically identified and cloned 14 kcnh genes in zebrafish. Then, we examined vertebrate KCNH channel evolution by phylogenetic and syntenic analyses. Our data revealed that the three subtypes of the KCNH gene family have already evolved in invertebrates, long before the emergence of vertebrates. The number of vertebrate KCNH genes increased, most likely due to whole-genome duplications (WGDs). In addition, we examined zebrafish kcnh gene expression during early embryogenesis by in situ hybridization. Each subgroups genes showed similar but distinct gene expression domains with some exceptions. Most of them were expressed in neural tissues. Notably, kcnh6a showed robust expression in the developing heart, consistent with its conserved role in cardiac repolarization. Additionally, a few kcnh genes were transiently expressed in nonneural tissues, such as somites and the notochord, suggesting they may have a unique role in embryonic development. Our phylogenetic and developmental analyses of KCNH channels shed light on their evolutionary history and potential roles during embryogenesis, in line with their physiological functions and human channelopathies.

Nucleobindin 1 (NUCB1) is a multifunctional conserved protein located in Golgi luminal, nucleus, extracellular and cytosolic pools. NUCB1 is multidomain protein comprised of a signal peptide, a DNA-binding domain, a leucine zipper and Ca2+ -binding domain. The multiple domains and localization of NUCB1 potentiates its interactions with various partners, such as DNA, Gi3 protein, cyclooxygenase 2, LRP10 and RNA suggests its importance in the regulation of many cellular events. We revealed that NUCB1 contains three RNA-binding regions and able to interact with two RNA fragments. It was suggested possible variants of the participation of NUCB1 in the interaction of the two partially complementary RNAs. The RNA-binding properties of the NUCB1 were also confirmed in vivo experiments.

Understanding the genetic structure of keystone species within river networks is essential for effective conservation and management. While population differentiation of anadromous species often occurs between river systems, less research has been conducted on differentiation within rivers with smaller catchment areas. In this study, we investigated the population genetic structure of wild Atlantic salmon (Salmo salar) across the small-scale river Ronne [a] system in southernmost Sweden using Restriction-site Associated DNA sequencing (RADseq). Although the Admixture analysis did not detect clearly defined genetic clusters, significant pairwise FST values and DAPC revealed emerging population differentiation among the Ronne [a] tributaries. The observed patterns are consistent with a system characterized by connectivity, where genetic flow is present but can be reduced by behavioral and ecological factors such as spawning homing behavior and selective movements. These findings suggest that, despite overall connectivity, Atlantic salmon populations in the Ronne [a] catchment area may function as partially independent sub-populations. This highlights the importance of conservation and management strategies in fragmented river systems to consider population genetic structure to support resilient salmon populations under ongoing anthropogenic pressures.

Currently, the genetic architecture of Middle Eastern populations is underrepresented in global genomic databases. This gap increases the rate of Variants of Uncertain Significance (VUSs) and clinical misinterpretations of genomic data especially in Middle Eastern populations. Whole exome sequencing was conducted on 90 healthy individuals from Jordan and the data were analysed using Principal Component Analysis (PCA) and multi-computational filtering. PCA revealed a double ancestry (EUR-AFR) admixture rather than a triple admixture (EUR-AFR-AMR). More than 3,500 populations-specific variants (PSVs) were identified, of which 72% were singletons. Additionally, 19 variants were significantly enriched compared to the maximum allele frequencies in public global databases (Fisher's exact test with Benjamini-Hochberg false discovery rate correction, p-value < 0.05). Consequently, the results suggest the reclassification of variants of Uncertain Significance (VUS) which reside in the ECE2 gene to likely benign and the variants of Conflicting Classification of Pathogenicity in the genes IL1RN and THPO to benign based on the significant allele frequency (AF=0.0389, p-value < 0.05). Furthermore, a pathogenic ClinVar variant was identified in a healthy individual, warranting careful interpretation. The findings underscore the importance of identifying PSVs in order to minimize or even prevent clinical misdiagnosis and highlight the unique genetic signature in Jordan. The study serves as a foundational resource for precision medicine in the region.

The Cytochrome P450 Family 11 Subfamily B Member 2 (CYP11B2) is the gene responsible for the synthesis of aldosterone synthase, the enzyme catalysing the terminal steps in the production of aldosterone. Polymorphism at the promoter region of this gene has been implicated to upregulate the synthesis of aldosterone synthase and the downstream production of aldosterone. A high aldosterone (hyperaldosteronism) is a known risk factor for hypertension. However, the association of the T-344C polymorphism with aldosterone production and the development of hypertension in the Ghanaian population has not been explored. Consequently, this study aims to unravel the relationship between T-344C (rs1799998) polymorphism, hypertension and serum aldosterone level. This study employed a case-control design enrolling 200 subjects of which 100 were hypertensive patients and 100 healthy controls in the Tamale Metropolis. Using a combination of genotyping, biochemical blood analysis and logistic modelling, we reveal that the frequency of the risk allele of rs1799998 (C) was higher amongst the patient group (0.565) than the control (0.315) group (p<0.0001). The adjusted (age and sex) logistic regression model revealed that the TC genotype [OR= 2.17 (1.08-4.38), p=0.0302] and the CC genotype [OR= 6.35 (2.60-15.54, p-value<0.0001] were significantly associated with hypertension. Under the genetic models, the recessive model (CC vs TC+TT) showed that the CC risk genotype was associated with hypertension [OR = 4.055 (1.838-8.949) p<0.001] after adjusting for age and sex. Furthermore, the CC risk genotype was associated with an elevated plasma aldosterone. In conclusion, the CYP11B2 gene polymorphism (T-344C) was associated with high plasma aldosterone and hypertension.

Rana dybowskii is distributed across Northeast Asian and represents a valuable medical resource. A high-quality assembly of the genome has not yet been reproted. This species has 2n=24 chromosomes, but a huge genome size that estimated at 3.5 ~4.6 Gb in the previous studies. The relatively large chromosome size, exceeding hundreds of megabases, may result in difficulties of obtaining a complete chromosome level genome. Here, we constructed a chromosome-level genome assembly of R. dybowskii by integrating PacBio HiFi long-read sequencing for de novo assembly and CiFi (3C coupled with HiFi sequencing) for scaffolding. The final assembly consists of 12 chromosomes with a total of 3.95 Gb and a scaffold N50 length of 455 Mb. BUSCO assessment using the tetrapoda_odb12 database identified 94.2% complete and 0.5% fragmented orthologs, suggesting a high level of completeness of the assembly. Genomic annotation revealed that repetitive sequences comprise over 53% of the assembly, with retroelements and DNA transposons accounting for 22% and 25%, respectively. A total of 43,999 protein-coding genes were predicted with the assistance of RNA-seq reads from four tissues (muscle, eye, testis and skin). This high-quality chromosome-level reference genome provides a valuable genomic resource for advancing genetic studies of the species.

Nuclear receptors, transcription factors essential for organism growth, development, and reproduction, are expressed in a variety of tissues, with some exhibiting differential expression between males and females. The Estrogen-related receptor (ERR) is a conserved metabolic nuclear receptor required for energy metabolism and lipid accumulation. While previous studies in Drosophila have identified potential ERR targets from mixed sex larval populations and adult males, it is unclear whether transcriptional targets and biological pathways downstream of ERR are altered in a sex-specific manner. Here, we took an RNA sequencing approach to identify candidate ERR targets specifically in adult females and compared differentially expressed genes to a published male-specific dataset. Whole body conditional knockout of ERR significantly downregulated transcription of enzymes associated with glycolysis and the pentose phosphate pathway. In contrast, components of the DNA replication machinery were selectively downregulated in adult females, whereas ribosome biogenesis transcription was increased. Our results have further defined the metabolic targets of ERR between males and females, as well as suggest that ERR regulates DNA replication and global translation in females. SUMMARYIn this manuscript, we used RNA sequencing to identify differential expression of transcripts dependent on the nuclear receptor ERR in Drosophila adult females. We find that ERR is required for activating transcription of glycolytic and pentose phosphate pathway enzymes, as observed in larvae and adult males. Furthermore, compared to males, loss of ERR in females specifically decreased DNA replication enzyme components while upregulating ribosomal components. Our results suggest that nuclear receptors have common and sex-specific targets, which will be of interest for those in the nuclear receptor and sexual dimorphism fields.

Humans and other vertebrates contain two estrogen receptors (ERs), ER-alpha and ER-beta, which mediate the physiological actions of three estrogens: estrone (E1), estradiol (E2) and estriol (E3). Of these three estrogens, in vivo, E2 is the strongest transcriptional activator of ER-alpha and ER-beta, E1 is next most active, followed by E3. We studied transcriptional activation of human ER-alpha and ER-beta by E2, E1 and E3 in African green monkey kidney (COS-7) cells, which we compared with studies of estrogen stimulation of ER transcription in human em-bryonic kidney (HEK-293) cells. To our surprise, in COS-7 cells, E3 had the lowest half-maximal response (EC50) for human ER-alpha and ER-beta than either E2, which was second most active estrogen, or E1. In contrast, for human ER-alpha and ER-beta transfected into HEK-293 cells, E2 was the most active estrogen, followed by E1 and E3. Similar results were found in COS-7 cells and HEK-293 cells transfected with elephant shark ER-alpha and ER-beta. Thus, under some conditions, E3 is a more active estrogen than either E2 or E1. This suggests that E3 may be a novel physiological ligand for the ER in some mammalian cells.

Anoxia is a major stress for most vertebrates and frequently accompanies harsh winter conditions, particularly in species that spend much of the season frozen solid. North American freeze-tolerant wood frogs (Rana sylvatica) can survive several months without oxygen and endure whole-body freezing for up to eight months of the year, with [~]70% of total body water frozen as extracellular ice, yet revive when temperatures rise in spring. Survival depends on multiple adaptations, including tolerance of prolonged oxygen deprivation while frozen, when breathing and circulation are halted. A key strategy involves hepatic glycogen mobilization, producing large amounts of glucose that are distributed to tissues where it functions both as a cryoprotectant and as a substrate for anaerobic ATP production. The present study examines the role of histone lysine methylation and demethylation in regulating liver proteins under anoxic conditions. Relative protein expression of seven histone methyltransferases (ASH2L-S, ASH2L-L, RBBP5, SETD8, SMYD2, ESET, SETD1), six lysine demethylases (KDM1A, KDM3B, KDM4A, KDM4B, KDM5A, KDM5C), and eight histone marks (H3K4me1, H3K4me2, H3K9me3, H3K27me3, H3K36me3, H3K79me3, H4K20me1, H4K20me3) were evaluated in wood frog liver under control, 4-hour, and 24-hour anoxia exposures. The data indicate that histone lysine methylation and demethylation contribute significantly to transcriptional regulation under anoxia. Specifically, H3K4, H3K36, and H3K79 methylation were associated with transcriptional activation, whereas H3K9, H3K27, and H4K20 methylation correlated with transcriptional repression. These findings highlight the dynamic role of epigenetic regulation in supporting hypometabolism and stress adaptation in freeze-tolerant wood frogs.

BackgroundNeurulation is a fundamental process in the formation of the central nervous system (CNS). The process begins with the folding and fusion of the neural plate to form the neural tube which subsequently gives rise to the development of the brain and spinal cord. Environmental and genetic factors that disrupt neurulation can induce neural tube defects (NTDs) and consequently cause additional developmental complications, including motor impairments. Purinergic signaling is a conserved form of extracellular communication (i.e. paracrine, synaptic signaling) that plays a role in early development. This signaling is mediated by purine nucleotides and nucleosides, which activate metabotropic P2Y and ionotropic P2X purinoceptors, respectively. Distinct patterns of intracellular calcium dynamics are observed throughout vertebrate development, from fertilization through organogenesis, including neurulation. Among P2X receptors, P2X4 is an ATP-modulated, Ca2+-permeable, ligand-gated ion channel characterized by having the highest Ca2+ permeability and is known to be modulated by ivermectin (IVM). ObjectiveOur investigation focuses on assessing the effects of IVM treatment during neurulation and evaluating the impact of this drug on phenotype, motor behavior and neuromuscular junction (NMJ) structure at tadpole stage. These results were compared with those obtained following separate treatments with compounds that specifically block glycine, GABA(A) and nACh receptors, all which have been described as IVM targets. ResultsIn this study we demonstrate the transcriptional expression for both P2X and P2Y purinergic receptors during neurulation, as well as the expression of P2X4. Following IVM neurula-treatments, we observed neural tube defects (NTDs), pigmentation changes, motor paralysis and alterations in neuromuscular junction (NMJ) structure, particularly affecting axonal branching. In contrast, treatment with the blockers strychnine, bicuculline and -bungarotoxin, used to assess the involvement of GlyR, GABA(A)R and 7nAChR, respectively, failed to show similar outcomes. ConclusionsIn summary, our results highlight the critical role of purinergic signaling during early development, particularly P2X4 receptor mediated signaling during neurulation which may account for the pharmacological effects induced by the positive allosteric modulator ivermectin.

Telomere length (TL) is increasingly used in ecology as a biomarker of individual quality and environmental stress, yet research on non-model species with complex life histories remains limited. Because TL varies among tissues and across ages in a species-specific manner, identifying non-lethal tissues that reliably reflect whole-organism telomere dynamics is essential for longitudinal telomere studies in the field. This study aimed to evaluate tissue-specific TL in Japanese eel (Anguilla japonica), an endangered catadromous fish. We first mapped the chromosomal distribution of telomeric sequences using fluorescent in situ hybridization (FISH), the first application of this method in this species. We then tested whether muscle and caudal fin, which can be sampled easily and non-lethally, can serve as suitable proxy tissues for TL measurements in wild individuals. Relative telomere length (RTL) was quantified by qPCR in blood, brain, caudal fin, gonads, heart, liver, and muscle. FISH analysis confirmed telomeric repeats at all chromosomal ends, with only weak interstitial signals on three chromosomal pairs unlikely to affect qPCR-based estimates. A generalized additive mixed model and Wilcoxons signed-rank tests revealed significant inter-tissue differences: RTL was shortest in the brain and muscle and longest in liver, blood and caudal fin. Muscle and caudal fin RTL were significantly correlated with RTL in many other tissues, supporting their use as proxy tissues for longitudinal TL monitoring, including responses to environmental variation. Both total length and age were tested as explanatory variables for RTL, and the model including total length showed a better fit than the age-based model. Non-linear relationships between RTL and total length observed in several tissues suggest physiological shifts associated with growth and sexual differentiation. Overall, these findings advance understanding of telomere dynamics in eels and establish muscle and caudal fin as suitable tissues for repeated, non-lethal TL assessment in ecological and conservation contexts.

In some reptiles, genetic and environmental sex determination interact whereby extreme incubation temperatures override genetic sex determination (GSD) to produce sex-reversed individuals. In one lizard with temperature-influenced GSD, the central bearded dragon, intron retention in the histone-modifier genes Kdm6b and Jarid2 has been implicated as a candidate signal linking temperature to sex. Equivalent intron retention is also present in two species with temperature-dependent sex determination, the red-eared slider turtle and the American alligator. The eastern three-lined skink, Bassiana duperreyi, represents another lizard with temperature induced sex reversal. It has an XY sex determination system in which low temperature incubation causes sex reversal of XX embryos to produce phenotypic males. In this study, we performed splice-aware analysis of RNA sequencing from hatchling brains of the three-lined skink. We investigated differences in alternative splicing and gene expression between the three sex conditions: XY males (XYm), XX females (XXf), and sex-reversed XX males (XXm). Sex reversal specific intron retention was observed in the gene, Ttll7, which only occurred in XXm and not in XYm or XXf. Intron retention in Ttll7 could alter the function of the encoded protein, a tubulin polyglutamylase, but its effect on sex reversal here is unknown. In addition, intron retention in the histone-modifier genes Jarid2 and Kdm6b occurred in all conditions. The presence of Kdm6b and Jarid2 intron retention in all sex conditions suggests that the pattern of intron retention in sex reversal in the eastern three-lined skink is distinct compared to the bearded dragon. We conclude that a different molecular pathway for sex reversal is induced in the three-lined skink, the details of which remain elusive.

Senescence human bone marrow mesenchymal stem cells (BMSCs), vulnerable to age-related defects, is poor in tissue regeneration. Cells in bone marrow accumulated senescent contributing to the development of metabolic energy regulation hold prospects for therapeutic advances. This study aimed to evaluate energy metabolic changes in male bone marrow mesenchymal stem cells senescence process. Our research established cell specific surface marker and enzymes expression level changes, as well as ECAR and OCR resonance. Notably, CD14, HLA-DRB1 and CD90 upregulated, glycolysis-related genes are increased, tricarboxylic acid cycle-related genes are decreased. We firstly identified links between time-dependent cell aging process and energy metabolism in BMSCs.

The c-MYC proto-oncogene regulates cellular proliferation, and its aberrant expression drives a range of human cancers. It also has a bidirectional regulatory relationship with the mammalian core circadian clock, with emerging evidence suggesting that MYC overexpression leads to clock disruption and loss of rhythms. While prior studies have probed MYCs role in clock disruption by overexpressing or mutating the c-MYC gene, our understanding of the endogenous nature of c-MYC is limited. A major gap in knowledge is whether MYC itself is expressed rhythmically and if so, how its timing relates to that of core clock components. To address these shortcomings, we generated a c-MYC reporter and assessed its circadian nature, comparing it to BMAL1 and PER2, and developed a computational model based on these and previous findings to evaluate its role(s). We developed lentiviral constructs for and established a U2OS (common circadian model) reporter cell line expressing luciferase (luc) driven by a human-derived c-MYC promoter sequence. To facilitate comparisons, as part of this work, we also developed a human-sequence derived BMAL1 promoter reporter to more readily recapitulate its behaviors. Using luminometry studies and subsequent data analyses, we demonstrated that the c-MYC promoter oscillated rhythmically in U2OS cells, which possess inherently low levels of c-MYC. Furthermore, we found that c-MYC oscillates out-of-phase relative to BMAL1 and PER2. Using this information, we built a mathematical model to better understand how c-MYCs oscillations at both basal and over-expressed levels affect the clock and vice versa. The model reproduced expected alterations to the core clock resulting from c-MYC overexpression and showed that MYCs role is as a disruptor, although the timing of MYC regulation can minimize its negative impact(s) on circadian timekeeping. This work is the first to assess c-MYCs phase relationships relative to the core clock and to provide evidence for its circadian nature. Author summaryc-MYC is a transcription factor that is highly regulated and plays an important role in cellular proliferation. In cancers, deregulation of c-MYC causes its overexpression, resulting in tumorigenesis. There have been multiple connections demonstrated between MYC and the circadian clock, including the clocks role in MYC expression and that its overexpression can lead to disruptions to the core circadian clock. However, knowledge of the expression patterns of MYC are limited, including whether they occur in a circadian manner. To address this, we developed a c-MYC-luciferase reporter in a human circadian cell model (U2OS). For the first time, we were able to directly assess the rhythmic nature of c-MYC using this tool. Subsequently, we developed a mathematical model to gain insights into the disruptive role of MYC in clock regulation under disease-like conditions and, in turn, the effects of the circadian clock on MYC. We found that c-MYC oscillated in a circadian manner in U2OS cells and that the MYC proteins role is as a disruptor, but its timing can minimize its negative impact(s) on circadian rhythms.

Polycystic ovary syndrome (PCOS) is a prevalent endocrine disorder characterized by hyperandrogenism, ovulatory dysfunction, and polycystic ovaries, with granulosa cell dysfunction being a key pathological feature. This study aimed to investigate the role of microRNA-6818-5p in PCOS pathogenesis. Quantitative PCR revealed a significant upregulation of circulating miR-6818-5p in PCOS patients compared to healthy controls. In vitro, functional assays in the human granulosa cell line KGN demonstrated that miR-6818-5p overexpression markedly inhibited cell proliferation (assessed by CCK-8 assay) and promoted apoptosis (measured by Annexin V/PI flow cytometry). Mechanistically, dual-luciferase reporter assay and Western blotting identified HSD17B2 as a direct target of miR-6818-5p, with miR-6818-5p mimics significantly suppressing HSD17B2 protein expression. In conclusion, our findings reveal that elevated miR-6818-5p in PCOS may contribute to follicular development dysfunction by targeting HSD17B2 to disrupt granulosa cell proliferation and apoptosis balance, offering novel insights into PCOS pathology and highlighting miR-6818-5p as a potential diagnostic biomarker and therapeutic target.

In the multitude of suitable experimental systems used for functional studies in the field of developmental biology, the sea urchin plays a central role due to its amenability to various methods, including both transient and stable transgenesis. Among others, transposable elements represent powerful tools for generating stable transgenic specimens, and Minos transposon turned out to be an excellent genetic tool in marine organisms, despite its efficiency being host-dependent. This study provides new evidence for the activity of Minos transposable elements and their stable integration into the genome of the Mediterranean sea urchin Paracentrotus lividus. Using the Minos-based technology coupled with a fully-automated system used for the qPCR screening of the Minos transposon integration, we devised a new pipeline for performing transgenesis-based functional studies in P. lividus.

During embryonic development bipotential gonads differentiate into either testes or ovaries under the direction of mutually exclusive gene networks. DNA methylation has been shown to regulate gene expression and to play a role in many developmental processes. This includes sex differentiation in which DNA methylation is linked to control of key sex genes. Whether DNA methylation regulates sexual differentiation in species where environmental influences such as temperature are involved is less clear. We conducted a genome-wide study in embryonic gonads of the central bearded dragon (Pogona vitticeps), a lizard with temperature induced sex reversal, and compared DNA methylation patterns to gene expression profiles at a stage of early sex differentiation. Overall, sex reversed ZZf females were found to have lower global methylation than both canonical sexes, ZZm males and ZWf females. We found that the expression of a key gene in sex differentiation, Amh, is regulated via DNA methylation. Amh is a driver of testis differentiation and is repressed in genetically determined females, as well as in temperature sex-reversed females, by hypermethylation around its transcription start site. Although the trigger of ovary determination is different in both groups of females, one by genetic complement and one by a temperature signal, downstream regulation of gene expression converges and seems to follow similar mechanisms.