Mab-3 Related Transcription Factor Research Articles

Curcumin Inhibits the Progression of Non-small Cell Lung Cancer by Regulating DMRT3/SLC7A11 Axis.

Non-small cell lung cancer (NSCLC) is a fatal malignancy all over the world. Emerging studies have shownthat curcumin might repress NSCLC progression by regulating ferroptosis, but the underlying mechanism remains unclear. 16HBE, LK-2, and H1650 cell viability was detected using Cell Counting Kit-8 assay. LK-2 and H1650 cell proliferation, apoptosis, and angiopoiesis were measured using 5-ethynyl-2'-deoxyuridine, flow cytometry, and tube formation assay. Superoxide dismutase, Malondialdehyde, Glutathione, and lactate dehydrogenase levels in LK-2 and H1650 cells were examined using special assay kits. Fe+ level was assessed using an iron assay kit. Doublesex and Mab-3 related Transcription Factor 3 (DMRT3) and solutecarrierfamily7member11(SLC7A11) protein levels were detected using western in NSCLC tissues, adjacent matched normal tissues, 16HBE cells, LK-2 cells, H1650 cells, and xenograft tumor tissues. Glutathione peroxidase 4, Acyl-CoA Synthetase Long Chain Family Member 4, and transferrinreceptor1 protein levels in LK-2 and H1650 cells were examined by western blot assay. DMRT3 and SLC7A11 levels were determined using real-time quantitative polymerase chain reaction. After JASPAR prediction, binding between DMRT3 and SLC7A11 promoter was verified using Chromatin immunoprecipitation and dual-luciferase reporter assays in LK-2 and H1650 cells. Role of curcumin on NSCLC tumor growth was assessed using the xenograft tumor model in vivo. Curcumin blocked NSCLC cell proliferation and angiopoiesis, and induced apoptosis and ferroptosis. DMRT3 or SLC7A11 upregulation partly abolished the suppressive role of curcumin on NSCLC development. In mechanism, DMRT3 was a transcription factor of SLC7A11 and increased the transcription of SLC7A11 via binding to its promoter region. Curcumin inhibited NSCLC growth in vivo by modulating DMRT3. Curcumin might constrain NSCLC cell malignant phenotypes partly through the DMRT3/SLC7A11 axis, providing a promising therapeutic strategy for NSCLC.

Identification and characterization of the Dmrt1B gene in the oriental river prawn, Macrobrachium nipponense.

Dmrt (doublesex and mab-3 related transcription factor) is a protein family of transcription factors implicated in sexual regulation. Dmrt proteins are widely conserved and known for their involvement in sex determination and differentiation across species, from invertebrates to humans. In this study, we identified a novel gene with a DM (doublesex/Mab-3)-domain gene in the river prawn, Macrobrachium nipponense, which we named MniDmrt1B due to its similarities and close phylogenetic relationship with Dmrt1B in Macrobrachium rosenbergii. Through amino acid alignments and structural predictions, we observed conservation and identified putative active sites within the DM domain. qRT-PCR analysis revealed that MniDmrt1B exhibited high expression levels in the testis, with consistently higher expression in males compared to females during development. Additionally, similar to other sex-regulated genes, the MniDmrt1B gene exhibited high expression levels during the sex differentiation-sensitive periods in M. nipponense. These results strongly indicated that MniDmrt1B probably plays an important role in testis development and sex differentiation in M. nipponense.

DMRTA2 supports glioma stem-cell mediated neovascularization in glioblastoma

Glioblastoma (GBM) is the most common and lethal brain tumor in adults. Due to its fast proliferation, diffusive growth and therapy resistance survival times are less than two years for patients with IDH-wildtype GBM. GBM is noted for the considerable cellular heterogeneity, high stemness indices and abundance of the glioma stem-like cells known to support tumor progression, therapeutic resistance and recurrence. Doublesex- and mab-3–related transcription factor a2 (DMRTA2) is involved in maintaining neural progenitor cells (NPC) in the cell cycle and its overexpression suppresses NPC differentiation. Despite the reports showing that primary GBM originates from transformed neural stem/progenitors cells, the role of DMRTA2 in gliomagenesis has not been elucidated so far. Here we show the upregulation of DMRTA2 expression in malignant gliomas. Immunohistochemical staining showed the protein concentrated in small cells with high proliferative potential and cells localized around blood vessels, where it colocalizes with pericyte-specific markers. Knock-down of DMRTA2 in human glioma cells impairs proliferation but not viability of the cells, and affects the formation of the tumor spheres, as evidenced by strong decrease in the number and size of spheres in in vitro cultures. Moreover, the knockdown of DMRTA2 in glioma spheres affects the stabilization of the glioma stem-like cell-dependent tube formation in an in vitro angiogenesis assay. We conclude that DMRTA2 is a new player in gliomagenesis and tumor neovascularization and due to its high expression in malignant gliomas could be a biomarker and potential target for new therapeutic strategies in glioblastoma.

New insights into the all-testis differentiation in zebrafish with compromised endogenous androgen and estrogen synthesis.

The regulatory mechanism of gonadal sex differentiation, which is complex and regulated by multiple factors, remains poorly understood in teleosts. Recently, we have shown that compromised androgen and estrogen synthesis with increased progestin leads to all-male differentiation with proper testis development and spermatogenesis in cytochrome P450 17a1 (cyp17a1)-/- zebrafish. In the present study, the phenotypes of female-biased sex ratio were positively correlated with higher Fanconi anemia complementation group L (fancl) expression in the gonads of doublesex and mab-3 related transcription factor 1 (dmrt1)-/- and cyp17a1-/-;dmrt1-/- fish. The additional depletion of fancl in cyp17a1-/-;dmrt1-/- zebrafish reversed the gonadal sex differentiation from all-ovary to all-testis (in cyp17a1-/-;dmrt1-/-;fancl-/- fish). Luciferase assay revealed a synergistic inhibitory effect of Dmrt1 and androgen signaling on fancl transcription. Furthermore, an interaction between Fancl and the apoptotic factor Tumour protein p53 (Tp53) was found in vitro. The interaction between Fancl and Tp53 was observed via the WD repeat domain (WDR) and C-terminal domain (CTD) of Fancl and the DNA binding domain (DBD) of Tp53, leading to the K48-linked polyubiquitination degradation of Tp53 activated by the ubiquitin ligase, Fancl. Our results show that testis fate in cyp17a1-/- fish is determined by Dmrt1, which is thought to stabilize Tp53 by inhibiting fancl transcription during the critical stage of sexual fate determination in zebrafish.

A novel sorting method for the enrichment of early human spermatocytes from clinical biopsies

To determine if early spermatocytes can be enriched from a human testis biopsy using fluorescence-activated cell sorting (FACS). Potential surface markers for early spermatocytes were identified using bioinformatics analysis of single-cell RNA-sequenced human testis tissue. Testicular sperm extraction samples from three participants with normal spermatogenesis were digested into single-cell suspensions and cryopreserved. Two to four million cells were obtained from each and sorted by FACS as separate biologic replicates using antibodies for the identified surface markers. A portion from each biopsy remained unsorted to serve as controls. The sorted cells were then characterized for enrichment of early spermatocytes. A laboratory study. Three men with a diagnosis of obstructive azoospermia (age range, 30-40 years). None. Sorted cells were characterized for RNA expression of markers encompassing the stages of spermatogenesis. Sorting markers were validated by their reactivity on human testis formalin-fixed paraffin-embedded tissue. Serine protease 50 (TSP50) and SWI5-dependent homologous recombination repair protein 1 were identified as potential surface proteins specific for early spermatocytes. After FACS sorting, the TSP50-sorted populations accounted for 1.6%-8.9% of total populations and exhibited the greatest average-fold increases in RNA expression for the premeiotic marker stimulated by retinoic acid (STRA8), by 23-fold. Immunohistochemistry showed the staining pattern for TSP50 to be strong in premeiotic undifferentiated embryonic cell transcription factor 1-/doublesex and Mab-3 related transcription factor 1-/STRA8+ spermatogonia as well as SYCP3+/protamine 2- spermatocytes. This work shows that TSP50 can be used to enrich early STRA8-expressing spermatocytes from human testicular biopsies, providing a means for targeted single-cell RNA sequencing analysis and invitro functional interrogation of germ cells during the onset of meiosis. This could enable investigation into details of the regulatory pathways underlying this critical stage of spermatogenesis, previously difficult to enrich from whole tissue samples.

Deleterious variants in X-linked RHOXF1 cause male infertility with oligo- and azoospermia.

Oligozoospermia and azoospermia are two common phenotypes of male infertility characterized by massive sperm defects owing to failure of spermatogenesis. The deleterious impact of candidate variants with male infertility is to be explored. In our study, we identified three hemizygous missense variants (c.388G>A: p.V130M, c.272C>T: p.A91V, and c.467C>T: p.A156V) and one hemizygous nonsense variant (c.478C>T: p.R160X) in the Rhox homeobox family member 1 gene (RHOXF1) in four unrelated cases from a cohort of 1201 infertile Chinese men with oligo- and azoospermia using whole-exome sequencing and Sanger sequencing. RHOXF1 was absent in the testicular biopsy of one patient (c.388G>A: p.V130M) whose histological analysis showed a phenotype of Sertoli cell-only syndrome. In vitro experiments indicated that RHOXF1 mutations significantly reduced the content of RHOXF1 protein in HEK293T cells. Specifically, the p.V130M, p.A156V, and p.R160X mutants of RHOXF1 also led to increased RHOXF1 accumulation in cytoplasmic particles. Luciferase assays revealed that p.V130M and p.R160X mutants may disrupt downstream spermatogenesis by perturbing the regulation of doublesex and mab-3 related transcription factor 1 (DMRT1) promoter activity. Furthermore, ICSI treatment could be beneficial in the context of oligozoospermia caused by RHOXF1 mutations. In conclusion, our findings collectively identified mutated RHOXF1 to be a disease-causing X-linked gene in human oligo- and azoospermia.

Effects of Alkalinity Exposure on Antioxidant Status, Metabolic Function, and Immune Response in the Hepatopancreas of Macrobrachium nipponense.

The oriental river prawn Macrobrachium nipponense is an important freshwater economic species in China, producing huge economic benefits. However, M. nipponense shows lower alkali tolerance than fish species, thus genetic selection is urgently needed in order to improve alkali tolerance in this species. In the present study, the effects of alkalinity exposure on the hepatopancreas of M. nipponense were measured under the alkali concentrations of 0 (control), 4, 8, and 12 mmol/L with the exposure time of 96 h through histological observations, measurement of antioxidant enzymes, metabolic profiling analysis, and transcriptome profiling analysis. The present study identified that the low concentration of alkali treatment (<4 mmol/L) did not result in morphological changes in the hepatopancreas and activity changes in antioxidant enzymes, while high-alkali treatment (>8 mmol/L) damaged the normal structures of the lumen and vacuoles and significantly stimulated the levels of superoxide dismutase, catalase, and total antioxidant capacity, indicating these antioxidant enzymes play essential roles in the protection of the body from the damage caused by the alkali treatment. Metabolic profiling analysis revealed that the main enriched metabolic pathways of differentially expressed metabolites in the present study were consistent with the metabolic pathways caused by environmental stress in plants and other aquatic animals. Transcriptome profiling analysis revealed that the alkali concentration of <8 mmol/L did not lead to significant changes in gene expression. The main enriched metabolic pathways were selected from the comparison between 0 mmol/L vs. 12 mmol/L, and some significantly up-regulated genes were selected from these metabolic pathways, predicting these selected metabolic pathways and genes are involved in the adaptation to alkali treatment in M. nipponense. The expressions of Ras-like GTP-binding protein, Doublesex and mab-3 related transcription factor 1a, and Hypothetical protein JAY84 are sensitive to changes in alkali concentrations, suggesting these three genes participated in the process of alkali adaptation in M. nipponense. The present study identified the effects of alkalinity exposure on the hepatopancreas of M. nipponense, including the changes in antioxidant status and the expressions of metabolites and genes, contributing to further studies of alkali tolerance in this species.

A chromosome-level assembly supports genome-wide investigation of the DMRT gene family in the golden mussel (Limnoperna fortunei).

The golden mussel (Limnoperna fortunei) is a highly invasive species that causes environmental and socioeconomic losses in invaded areas. Reference genomes have proven to be a valuable resource for studying the biology of invasive species. While the current golden mussel genome has been useful for identifying new genes, its high fragmentation hinders some applications. In this study, we provide the first chromosome-level reference genome for the golden mussel. The genome was built using PacBio HiFi, 10X, and Hi-C sequencing data. The final assembly contains 99.4% of its total length assembled to the 15 chromosomes of the species and a scaffold N50 of 97.05 Mb. A total of 34,862 protein-coding genes were predicted, of which 84.7% were functionally annotated. A significant (6.48%) proportion of the genome was found to be in a hemizygous state. Using the new genome, we have performed a genome-wide characterization of the Doublesex and Mab-3 related transcription factor gene family, which has been proposed as a target for population control strategies in other species. From the applied research perspective, a higher-quality genome will support genome editing with the aim of developing biotechnology-based solutions to control invasion. From the basic research perspective, the new genome is a high-quality reference for molecular evolutionary studies of Mytilida and other Lophotrochozoa, and it may be used as a reference for future resequencing studies to assess genomic variation among different golden mussel populations, unveiling potential routes of dispersion and helping to establish better control policies.

Comprehensive Analysis of DMRT3 as a Potential Biomarker Associated with the Immune Infiltration in a Pan-Cancer Analysis and Validation in Lung Adenocarcinoma.

Doublesex and Mab-3 related Transcription Factor 3 (DMRT3) is associated with the prognosis of some tumors. It is possible to explore the role of DMRT3 in the cancer process using bioinformatic approaches and experimental validation. We comprehensively explored the clinical and immunological characteristics of DMRT3. The DMRT3 expression is abnormal in human cancers and correlates with clinical staging. A high DMRT3 expression is significantly associated with poor overall survival (OS) in KIRC, KIRP, LUAD, and UCEC. Amplification was the greatest frequency of the DMRT3 alterations in pan-cancer. The OS was significantly lower in the DMRT3 altered group than in the DMRT3 unaltered group (P = 0.0276). The DMRT3 expression was significantly associated with MSI in three cancer types and TMB in six cancer types. The DMRT3 expression was significantly correlated with the level of the immune cell infiltration and the immune checkpoint genes. The DMRT3 was involved in some pathways in pan-cancer. DMRT3 may play a role in chemotherapy and may be associated with chemoresistance. A ceRNA network of KCNQ1OT1/miR-335-5p/DMRT3 was constructed in LUAD. DMRT3 was significantly upregulated in the LUAD cell lines. DMRT3 was aberrantly expressed in pan-cancer and may promote tumorigenesis and progression via different mechanisms. DMRT3 can be used as a therapeutic target to treat cancer in humans.

Testicular Transcriptome of Males and Pseudo-Males Provides Important New Insight into Sex Reversal of Rana dybowskii.

Simple SummaryAquaculture in Northeast China has favored Rana dybowskii (R. dybowskii) because of its regional specificity and the economic value of the female frog’s ovary, and the increasing proportion of female frogs is beneficial to improve the economic efficiency of aquaculture. Unfortunately, the sexual reversal from genotypic female to phenotypic male has led to a decrease in the proportion of females in the breeding population. In this study, differentially expressed genes between testes of sex reversal and male testes were screened by transcriptome sequencing. These findings provide a comprehensive analysis of gene expression in R. dybowskii sex reversal from genotypic female to phenotypic male and provide new ideas for sex control of R. dybowskii.Rana dybowskii (R. dybowskii) is an ecological species found in China, Japan, Korea, and Russia. Like most amphibians, R. dybowskii lacks heterotypic sex chromosomes, limiting the in-depth study of sex determination and sex reversal mechanisms. Previous studies have shown that certain environmental factors can modify R. dybowskii genotypic females into phenotypic males, but the mechanism is still unknown. Considering the difficulties in identifying and collecting sex reversal gonads at different stages of differentiation under natural conditions, testes from sexually mature wild adult R. dybowskii were taken in this study, and the genotypic sex of individuals and sex reversal were identified by two male-linked genetic markers reported in our most recent findings. Transcriptome sequencing was performed on testicular tissue from males and pseudo-males, as well as female ovary tissue. The results show that the gene expression patterns of pseudo-males’ testes were similar to those of the males but highly differed from females’ ovaries. One hundred and seventeen differentially expressed genes between testes of pseudo-males and males were found, and the up-regulation of doublesex and mab-3 related transcription factor 1 (Dmrt1) in testes of pseudo-males may play a key role in R. dybowskii sex reversal.

Adult spinal Dmrt3 neurons receive direct somatosensory inputs from ipsi- and contralateral primary afferents and from brainstem motor nuclei.

In the spinal cord, sensory-motor circuits controlling motor activity are situated in the dorso-ventral interface. The neurons identified by the expression of the transcription factor Doublesex and mab-3 related transcription factor 3 (Dmrt3) have previously been associated with the coordination of locomotion in horses (Equus caballus, Linnaeus, 1758), mice (Mus musculus, Linnaeus, 1758), and zebrafish (Danio rerio, F. Hamilton, 1822). Based on earlier studies, we hypothesized that, in mice, these neurons may be positioned to receive sensory and central inputs to relay processed commands to motor neurons. Thus, we investigated the presynaptic inputs to spinal Dmrt3 neurons using monosynaptic retrograde replication-deficient rabies tracing. The analysis showed that lumbar Dmrt3 neurons receive inputs from intrasegmental neurons, and intersegmental neurons from the cervical, thoracic, and sacral segments. Some of these neurons belong to the excitatory V2a interneurons and to plausible Renshaw cells, defined by the expression of Chx10 and calbindin, respectively. We also found that proprioceptive primary sensory neurons of type Ia2, Ia3, and Ib, defined by the expression of calbindin, calretinin, and Brn3c, respectively, provide presynaptic inputs to spinal Dmrt3 neurons. In addition, we demonstrated that Dmrt3 neurons receive inputs from brain areas involved in motor regulation, including the red nucleus, primary sensory-motor cortex, and pontine nuclei. In conclusion, adult spinal Dmrt3 neurons receive inputs from motor-related brain areas as well as proprioceptive primary sensory neurons and have been shown to connect directly to motor neurons. Dmrt3 neurons are thus positioned to provide sensory-motor control and their connectivity is suggestive of the classical reflex pathways present in the spinal cord.

CLINICAL AND GENETIC FACTORS ASSOCIATED WITH OSTEONECROSIS IN BRAZILIAN SICKLE CELL DISEASE PATIENTS: CASE-CONTROL STUDY

Sickle cell disease (SCD) includes a group of hereditary disorders characterized by mutations on the gene that codes hemoglobin (Hb). Patients with SCD are vulnerable to vascular occlusion within bone tissue, which may result in complications such as medullary infarction in the diaphysis and osteonecrosis (ON), which affects the epiphysis. Both processes may result in substantial morbidity and pain for the patients. This study aims to identify factors associated with the development of ON in patients with SCD. A nested case-control study of patients with SCD from six Brazilian blood centers, participating in the Recipient Epidemiology and Donor Evaluation Study-III (REDS-III) Brazil SCD Cohort was performed. A history of ON was abstracted from participants medical record using standardized definitions to define cases. Cases were matched to controls without ON in a 1:3 ratio matched by age, genotype and blood center using propensity score. A genome-wide association study (GWAS) was conducted to identify single nucleotide polymorphisms (SNPs) associated with ON. There were 307 ON cases in the 2793 cohort participants (prevalence 11%) who were compared to 921 controls without ON. The most common locations of ON were the proximal femur (91.2%) and proximal humerus (7.7%). Variables independently associated with ON in a multivariable model included patients with more severe sickle cell disease (SS, Sβ0, severe Sβ+ or SD genotypes, OR 1.81, 95% CI 1.11; 2.95 compared to less severe genotypes SC/SB+), higher values of hemoglobin (OR 1.25, 1.12; 1.39), hospitalization by pain crisis (OR 1.44, 1.06; 1.94), chronic transfusion treatment (OR 1.53, 1.10; 2.11), acute chest syndrome (OR 1.46, 1.06; 2.00), sepsis/bacteremia (OR 1.67, 1.03; 2.70) and osteomyelitis (OR 1.72, 1.05; 2.80). Increased indirect bilirubin and stroke were associated with a lower chance of developing ON (OR 0.81, 0.71; 0.93 for indirect bilirubin and OR 0.42, 0.24; 0.72 for stroke). The GWAS suggested an association of ON with the gene doublesex and mab-3 related transcription factor 2 (DMRT2) (p < 5 × 10−8). SCD is the most common monogenic disorder in Brazil and is associated with multisystem complications. Orthopedic complications represent 31.0% of these manifestations, with ON being the most common non-infectious joint complication. Knowing the risk factors that precede ON may help to diagnose and treat it early in order to improve the quality of life of patients. The few studies carried out in the Brazilian population on the occurrence of ON in SCD patients had small sample sizes. A higher chance of having ON was associated with patients with higher hemoglobin levels, more hospitalizations for pain crises and acute chest syndrome. Also the relationship of treatment with chronic transfusion, sepsis/bacteremia and osteomyelitis were associated with ON, which may indicate that the occurrence of this complication may be multifactorial or related to the severity of SCD. SNPs near DMRT2, a gene previously associated with skeletal disorders, rib malformation in a newborn and transition of endochondral bone formation, showed an altered risk for ON.

A novel Dmrt gene of crustacean: Functional analysis of idmrt-2 gene in the male reproductive system from Scylla paramamosain

The Dmrt (double-sex and mab-3 related transcription factor) gene family is considered to be a highly conserved gene family related to sex determination and sexual differentiation across species. In order to better understand the role of the idmrt-2 gene in gonad development in Scylla paramamosain, the idmrt-2 gene was cloned and analyzed. The cDNA contains a 1659 bp ORF region encoding 552 amino acids. The qRT-PCR results showed that idmrt-2 was significantly more expressed in the testis than in other tissues (p < 0.05). The expression of idmrt-2 was highest in the spermatids stage (T2 stage), followed by the mature sperms stage (T3 stage) and significantly higher than in the spermatocytes stage (T1 stage) (p < 0.05) during testicular development and the expression difference was not significant in different stages of ovarian development. RNAi studies revealed that after idmrt-2 was knocked down, the expression of Dmrt-like and foxl-2 genes in the testis decreased, as well as IAG expression in the androgenic gland. The findings suggest that idmrt-2 may be an IAG regulator and involved in testicular development.

Dmrt2 and myf5 Link Early Somitogenesis to Left-Right Axis Determination in Xenopus laevis.

The vertebrate left-right axis is specified during neurulation by events occurring in a transient ciliated epithelium termed left-right organizer (LRO), which is made up of two distinct cell types. In the axial midline, central LRO (cLRO) cells project motile monocilia and generate a leftward fluid flow, which represents the mechanism of symmetry breakage. This directional fluid flow is perceived by laterally positioned sensory LRO (sLRO) cells, which harbor non-motile cilia. In sLRO cells on the left side, flow-induced signaling triggers post-transcriptional repression of the multi-pathway antagonist dand5. Subsequently, the co-expressed Tgf-β growth factor Nodal1 is released from Dand5-mediated repression to induce left-sided gene expression. Interestingly, Xenopus sLRO cells have somitic fate, suggesting a connection between LR determination and somitogenesis. Here, we show that doublesex and mab3-related transcription factor 2 (Dmrt2), known to be involved in vertebrate somitogenesis, is required for LRO ciliogenesis and sLRO specification. In dmrt2 morphants, misexpression of the myogenic transcription factors tbx6 and myf5 at early gastrula stages preceded the misspecification of sLRO cells at neurula stages. myf5 morphant tadpoles also showed LR defects due to a failure of sLRO development. The gain of myf5 function reintroduced sLRO cells in dmrt2 morphants, demonstrating that paraxial patterning and somitogenesis are functionally linked to LR axis formation in Xenopus.

Abstract 769: Genomic and transcriptomic characterization of benign and malignant struma ovarii

Abstract Background: Struma Ovarii (SO) is a rare ovarian teratoma characterized by the presence of thyroid tissue in &amp;gt;50% of the tumor. The majority of SO are benign; however, malignant transformation occurs in up to 5% of the cases. The molecular foundations of benign and malignant SO are grossly unknown. Therefore, the goal of this study was to perform the first comprehensive genomic and transcriptomic analysis of the benign and malignant SO. Material and Method: We performed whole-exome sequencing (WES) and targeted RNA-sequencing (seq) on the DNA and RNA extracted from formalin-fixed paraffin-embedded SO tumor tissue samples. WES library was prepared using Agilent’s SureSelect XT HS2 kit, with 4 samples failing the quality assessment (QA). Variants were called from GATK processed WES data and annotated using VEP (with ClinVar and COSMIC databases). The targeted RNA-Seq library was prepared using the TruSight RNA Pan-Cancer Panel kit covering 1385 cancer genes, with all samples passing QA. The clinical characteristics of the study cohort were summarized by percentages for categorical variables and medians with 25-75% interquartile ranges for continuous variables. Results: The study included 31 tissue samples - 21 benign and 10 malignant, including 6 cases of papillary thyroid cancer (PTC), 3 of follicular variant of PTC, and 1 of follicular thyroid cancer. Patients with benign SO were characterized by the median age at diagnosis of 39 years [33-54], tumor size of 3.1 cm [2.5-5.8], while the patients with malignant SO presented at age of 45 [28-54], tumor size of 6 cm [0.85-14] and metastatic disease in 30% (3/10) - 2 patients with peritoneal metastases and 1 patient with pelvic lymph node metastases. The E1A Binding Protein P300, EP300 (6/27), and Isocitrate dehydrogenase, IDH2 (5/27) were the topmost mutated genes in the SO samples. Malignant SO samples were characterized with the presence of pathogenic variants of KRAS (pQ61L and pG12V), NRAS (pQ61R), TP53 (splice site) mutations, and Nuclear Receptor Binding SET Domain Protein 1 (NSD1) fusion as the most common molecular drivers. Among benign SO samples, the most common driver was Thyroglobulin (TG) fusion with either Guanine Nucleotide binding protein (GNAS) or Rac Family Small GTPase 1 (RAC1). Differential expression analysis showed that the member of tumor suppressor family - tumor protein 63 (TP63) was the most downregulated (Log2FC = -3), while Double-sex and Mab-3 Related Transcription Factor 1 (DMRT1), implicated in the development of germ cell tumors, was the most upregulated gene in malignant SO samples over benign (Log2FC = 2.1; padj&amp;lt;0.05). Conclusions: In contrast to cancer arising from the thyroid gland, characterized by BRAFV600E as the most common mutation, malignant SO belongs to RAS-like tumors. The downregulation of tumor suppressors and upregulation of DMRT1 might be implicated in the malignant transformation of SO. Citation Format: Shilpa Thakur, Darryl Nousome, Kshama Aswath, Stephanie Cardenas, Sonam Kumari, Ruth Adewale, Maria Merino, Esra Dikoglu, Padmasree Veeraraghavan, Sriram Gubbi, Joanna Klubo-Gwiezdzinska. Genomic and transcriptomic characterization of benign and malignant struma ovarii [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 769.

Reproductive toxicity of InP/ZnS QDs in male rare minnow (Gobiocypris rarus)

InP/ZnS quantum dots (QDs) stand out among cadmium-free alternatives for higher exciton Bohr radius and strong quantum confined effect. In this study, the reproductive toxicity and mechanism of InP/ZnS QDs at different concentrations in male Chinese rare minnows (Gobiocypris rarus) were investigated. The results showed that QDs in 800 nmol/L concentration group could enter the testes after 1 d of exposure and caused changes in the structure of the testes, including the scattered distribution of seminal vesicles, reduction in germ cells and vacuolation in some areas of interstitial cells. The expression levels of androgen receptor (Ar) and doublesex and mab-3 related transcription factor 1 (Dmrt1) and the tight junction protein-related genes β-catenin and occludin were upregulated in rare minnows. The sperm quality and ATP content of parents in the 800 nmol/L treatment group were significantly decreased. Continuous detection of the development of F1 generation embryos showed that parental exposure to InP/ZnS QDs reduced the heart rate and spontaneous movement frequency of F1 generation embryos, and the fertilization rate of the F1 generation in the 800 nmol/L treatment group was significantly reduced. In general, the sperm quality and testicular structure of adult rare minnows were not significantly affected by concentrations below 400 nmol/L. High-concentration InP/ZnS QDs exposure can damage the integrity of the blood–testis barrier (BTB) and cause reproductive damage to the parents of rare minnows, which will continue to the next generation and affect their development.

The miR-200 Family Targeting amh Affects the Gonadal Development of Japanese Flounder

Four members of the miR-200 family in Japanese flounder (Paralichthys olivaceus) have sex-biased expression patterns, but their target genes and how they work in the development of the gonads are rarely known. Anti-Müllerian hormone (AMH) can inhibit the development of Muller’s duct in female mammals and regulate the formation of gametes after sexual maturity. There is no Muller’s duct in teleosts, but the amh gene still exists. Knockout of amh results in sex reversal from male to female. Therefore, it is essential to explore the relationship between the miR-200 family and amh to clarify what role miR-200 plays in the development of the gonads. In Japanese flounder, the two binding sites for the miR-200 family in the 3′UTR of amh were found through bioinformatic prediction. Double luciferase and green fluorescent protein reporter experiments demonstrated amh to be directly targeted by miR-200a and miR-200b. Moreover, miR-200a and miR-200b reduced the expression of amh through site 1 rather than site 2. To explore the regulatory role of miR-200a in gonadal development, we further overexpressed miR-200a in the primary Sertoli cells of the testis. With the overexpression of miR-200a, the expression of amh decreased, while the expression of the other two male sex-related genes, dmrt1 (doublesex and mab-3 related transcription factor 1) and gsdf (diagonal soma driven factor), increased significantly. This result indicates that the miR-200 family regulates the gonadal differentiation and development by targeting amh in Japanese flounder.

Genome-Wide Identification, Phylogeny, and Expression Profile of the Dmrt (Doublesex and Mab-3 Related Transcription Factor) Gene Family in Channel Catfish (Ictalurus punctatus).

The Dmrt (Doublesex and Mab-3 related transcription factor) gene family is a class of crucial transcription factors characterized by a conserved DM domain related to sex determination and differentiation, which has been systematically described in various teleost fish, but less in channel catfish (Ictalurus punctatus), an important global aquaculture species in the US and China. In this study, seven Dmrt genes from channel catfish genome were identified and analyzed using bioinformatics methods. Seven IpDmrt genes were distributed unevenly across five chromosomes. Synteny analysis revealed that Dmrt1, Dmrt2a, Dmrt2b, Dmrt3, Dmrt4, and Dmrt5 were relatively conserved in teleost fish. Tissue distribution analysis showed that IpDmrt1, IpDmrt2b, IpDmrt5, and IpDmrt6 exhibited sexually dimorphic expression patterns and, among them, IpDmrt1 and IpDmrt6 had high expression levels in the testes, while IpDmrt2b and IpDmrt5 had more significant expression levels in the ovaries than in other tissues. After 17β-estradiol treatment, IpDmrt2b and IpDmrt5 were significantly up regulated, while the expression of IpDmrt1 and IpDmrt6 was significantly repressed in XY channel catfish ovaries compared with XX channel catfish ovaries. The present study provides a comprehensive insight into the Dmrt gene family of channel catfish. The results suggest that IpDmrt1 and IpDmrt6 may play an important role in testis differentiation/development, while IpDmrt2b and IpDmrt5 are critical in ovary development in this species.

Single Cell Transcriptomic Analysis of Spinal Dmrt3 Neurons in Zebrafish and Mouse Identifies Distinct Subtypes and Reveal Novel Subpopulations Within the dI6 Domain.

The spinal locomotor network is frequently used for studies into how neuronal circuits are formed and how cellular activity shape behavioral patterns. A population of dI6 interneurons, marked by the Doublesex and mab-3 related transcription factor 3 (Dmrt3), has been shown to participate in the coordination of locomotion and gaits in horses, mice and zebrafish. Analyses of Dmrt3 neurons based on morphology, functionality and the expression of transcription factors have identified different subtypes. Here we analyzed the transcriptomes of individual cells belonging to the Dmrt3 lineage from zebrafish and mice to unravel the molecular code that underlies their subfunctionalization. Indeed, clustering of Dmrt3 neurons based on their gene expression verified known subtypes and revealed novel populations expressing unique markers. Differences in birth order, differential expression of axon guidance genes, neurotransmitters, and their receptors, as well as genes affecting electrophysiological properties, were identified as factors likely underlying diversity. In addition, the comparison between fish and mice populations offers insights into the evolutionary driven subspecialization concomitant with the emergence of limbed locomotion.

Comparative phylotranscriptomics reveals putative sex differentiating genes across eight diverse bivalve species

Mollusks, especially bivalves, exhibit a great diversity of sex determining mechanisms, including both genetic and environmental sex determination. Some bivalve species can be gonochoristic (separate sexes), while others are hermaphroditic (sequential or simultaneous). Several models have been proposed for specific bivalve species, utilizing information gained from gene expression data, as well as limited RAD-seq data (e.g., from Crassostrea gigas). However, these mechanisms are not as well studied as those in model organisms (e.g., Mus musculus, Drosophila melanogaster, Caenorhabditis elegans) and many genes involved in sex differentiation are not well characterized. We used phylotranscriptomics to better understand which possible sex differentiating genes are in bivalves and how these genes relate to similar genes in diverse phyla. We collected RNAseq data from eight phylogenetically diverse bivalve species: Argopecten irradians, Ensis directus, Geukensia demissa, Macoma tenta, Mercenaria mercenaria, Mya arenaria, Mytilus edulis, and Solemya velum. Using these data, we assembled representative transcriptomes for each species. We then searched for candidate sex differentiating genes using BLAST and confirmed the identity of nine genes using phylogenetics analyses from nine phyla. To increase the confidence of identification, we included ten bivalve genomes in our analyses. From the analysis of doublesex and mab-3 related transcription factor (DMRT) genes, we confirmed the identify of a Mollusk-specific sex determining DMRT gene: DMRT1L. Based on gene expression data from M. edulis and previous research, DMRT1L and FoxL2 are key genes for male and female development, respectively.