Expression Patterns Research Articles

IMMU-07. THE IMMUNE LANDSCAPE OF PEDIATRIC BRAIN TUMORS: A TRANSCRIPTOMIC ANALYSIS

Abstract Despite standard-of-care therapy, pediatric brain tumors are now the leading cause of cancer-related death in children, highlighting an urgent need for the development of new treatments. Immunotherapy is actively being evaluated as a therapeutic approach to pediatric brain tumors, however, more research is needed to reveal additional antigenic targets that exist in these malignancies. In this study, our team performed a transcriptomic analysis of pediatric brain tumor RNA sequencing data within the Children’s Brain Tumor Network (CBTN) to identify intra and extracellular antigens that are highly expressed across major tumor types. Interestingly, our team observed that most tumors had high gene expression of a group of antigens that were unique to that specific tumor type. Specifically, our analysis revealed that diffuse midline gliomas (DMGs) highly expressed the extracellular antigens CA9, CTCFL, and ST8SIA1, whereas atypical rhabdoid tumors (ATRTs) highly expressed MUC1 and TEK. Other tumors that displayed high gene expression for a variety of antigenic targets included H3-wildtype high-grade gliomas (HGGs), and ependymomas (EPNs). Strikingly, we observed minimal difference in antigen expression in primary tumors when compared to patient-matched recurrences. Beyond antigen expression, we evaluated the expression of antigen processing machinery (APM) which includes major histocompatibility complexes (MHCs). Our findings revealed that DMGs had the lowest level of APM expression, however, these tumors uniquely expressed high levels of the immune checkpoints ADORA2A, CD276, and KLRC1. Contrarily, ATRTs showed a high expression of immune checkpoints CD274 (PD-L1), PVR, and CD80. Other tumors included in our analysis had unique expression patterns of antigen, as well as APM and immune checkpoints. Our findings illustrate that pediatric brain tumors have distinct expression patterns of antigens, APM, and immune checkpoints that are specific to tumor type, emphasizing diverse, rather than general, immunotherapeutic approaches must be considered for childhood brain tumors.

The potential of ESCO2 as a prognostic and immunotherapeutic marker of pan-cancer and its role in anti-PD1 treatment of bladder cancer.

Background Establishment of sister chromatid cohesion N-acetyltransferase 2 (ESCO2), a member of the EFO2 family, is implicated in the pathogenesis and progression of various cancers. However, there has been limited comprehensive pan-cancer analysis conducted on ESCO2 thus far. Methods Publicly available databases, such as the UCSC Xena database, were utilized to examine differential expression patterns across various cancer types. In addition, variations in expression levels were investigated across distinct clinical stages. Univariate Cox regression and Kaplan-Meier survival analyses were conducted to evaluate the impact on overall survival (OS), disease-specific survival (DSS), disease-free interval (DFI), and progression-free interval (PFI) at a pan-cancer level. The correlation between ESCO2 expression and immune cell infiltration was examined to gain insight into the tumor microenvironment in different cancers. The results of the bioinformatic analysis were validated using immunotherapy clinical trials and pathological specimens. CCK-8 and transwell assay experiments were performed to investigate the biological function of ESCO2. Results ESCO2 expression was found to be up-regulated in most cancers, with a correlation to TNM stages. Prognostic analysis indicated that overexpression of ESCO2 was associated with poor prognosis in various cancers. Furthermore, the correlation between ESCO2 expression and immune cell infiltration suggested its potential as a predictor for immunotherapy efficacy. Notably, ESCO2 expression showed positive associations with immunoinhibitor, immnostimulator, MHC molecule, chemokine receptor, tumor mutation burden (TMB) and microsatellite instability (MSI) levels in bladder cancer (BLCA). The validation cohort for immunotherapy corroborated these findings and substantiated that ESCO2 could function as an autonomous prognostic biomarker and a promising target for cancer treatment via immunotherapy. In addition, in vitro experiments confirmed the role of ESCO2 in influcing the proliferation, invasion and migration of BLCA cells. Conclusion ESCO2 participates in regulating the immune infiltration and affecting the prognosis of patients in many cancers, especially in BLCA. ESCO2 may serve as a prognostic and immunotherapy biomarker in future treatment of human cancer.

Discovery of key molecular signatures for diagnosis and therapies of glioblastoma by combining supervised and unsupervised learning approaches.

Glioblastoma (GBM) is the most malignant brain cancer and one of the leading causes of cancer-related death globally. So, identifying potential molecular signatures and associated drug molecules are crucial for diagnosis and therapies of GBM. This study suggested GBM-causing ten key genes (ASPM, CCNB2, CDK1, AURKA, TOP2A, CHEK1, CDCA8, SMC4, MCM10, and RAD51AP1) from nine transcriptomics datasets by combining supervised and unsupervised learning results. Differential expression patterns of key genes (KGs) between GBM and control samples were verified by different independent databases. Gene regulatory network (GRN) detected some important transcriptional and post-transcriptional regulators for KGs. The KGs-set enrichment analysis unveiled some crucial GBM-causing molecular functions, biological processes, cellular components, and pathways. The DNA methylation analysis detected some hypo-methylated CpG sites that might stimulate the GBM development. From the immune infiltration analysis, we found that almost all KGs are associated with different immune cell infiltration levels. Finally, we recommended KGs-guided four repurposable drug molecules (Fluoxetine, Vatalanib, TGX221 and RO3306) against GBM through molecular docking, drug likeness, ADMET analyses and molecular dynamics simulation studies. Thus, the discoveries of this study could serve as valuable resources for wet-lab experiments in order to take a proper treatment plan against GBM.

ANGI-08. SPATIAL TRANSCRIPTOMIC COMPARISONS OF GLIOBLASTOMA TISSUE REVEAL UPREGULATION OF NEURODEVELOPMENTAL PATHWAYS AND SYNAPTIC GENES IN INFILTRATING TUMOR CELLS

Abstract Glioblastoma remains one of the deadliest brain malignancies. First-line therapy consists of maximal surgical tumor resection, chemotherapy, and radiotherapy. Malignant cells escape surgical resection by migrating into the surrounding healthy brain tissue, where they give rise to the recurrent tumor. Gene expression profiling allows glioblastoma tumor cores to be classified into mesenchymal, proneural, and classical subtypes, each with distinct genetic alterations and cellular compositions. In contrast, the adjacent brain parenchyma where infiltrating malignant cells escape surgical resection is less characterized in patients. Using single-cell and multicellular resolution spatial transcriptomics on tissues from both the tumor core and infiltrated brain tissue (n = 11), we compared the transcriptional profiles of malignant cells in these regions. Malignant cells near regions with microvascular proliferation showed increased expression of genes related to vascular homeostasis. Within tumor cores, proneural and mesenchymal subtypes exhibited distinct spatial gene expression patterns, although these differences were reduced in the infiltrated brain tissue, apart from gene expression due to chromosomal alterations specific to the proneural subtype. We identified two transcriptional patterns of brain infiltration, with the dominant pattern showing a transition from mesenchymal to proneural states. This transition was accompanied by increased expression of genes linked to neurodevelopmental pathways and glial cell differentiation. Additionally, one gene module upregulated in infiltrating malignant cells was predictive of poor patient survival and enriched for genes associated with differentiated neuronal cells. Together, our findings provide an updated view of the spatial landscape of glioblastomas and infiltrated brain tissue, furthering our understanding of the malignant cells that invade healthy brain. This insight offers new avenues for targeted therapy after surgical resection of the primary tumor.

Gene expression-based modeling of overall survival in Black or African American patients with lung adenocarcinoma

IntroductionLung cancer is a leading cause of cancer-related deaths worldwide. Black/African American (B/AA) populations, in particular, exhibit the highest incidence and mortality rates of lung adenocarcinoma (LUAD) in the United States.MethodsThis study aims to explore gene expression patterns linked to LUAD in B/AA and case-matched white patients, with the goal of developing predictive models for prognosis. Leveraging RNA sequencing data from The Cancer Genome Atlas (TCGA) database, genes and pathways associated with overall survival (OS) were identified.ResultsThe OS-associated genes in B/AA patients were distinct from those in white patients, showing predominant enrichment in immune-related pathways. Furthermore, mRNA co-expression network analysis revealed that OS-associated genes in B/AA patients had higher levels of interaction with various pathways, including those related to immunity, cell-ECM interaction, and specific intracellular signaling pathways. Notably, a potential B/AA-specific biomarker, C9orf64, demonstrated significant correlations with genes involved in immune response. Unsupervised machine learning algorithms stratified B/AA patients into groups with distinct survival outcomes, while supervised algorithms demonstrated a higher accuracy in predicting survival for B/AA LUAD patients compared to white patients.DiscussionIn total, this study explored OS-associated genes and pathways specific for B/AA LUAD patients. Further validation and clinical application of these findings are warranted to address disparities and improve outcomes in diverse patient populations.

Detection of Candidate Genes and Development of KASP Markers for Pod Length and Pod Width by Combining Genome-Wide Association and Transcriptome Sequencing in Vegetable Soybean

Vegetable soybeans are one of the most important vegetable types in East Asia. The yield of vegetable soybeans is considerably influenced by the size of their pods. To facilitate the understanding of the genetic basis of the pod length and width in vegetable soybeans, we conducted a genome-wide association study (GWAS) and transcriptome sequencing. Four quantitative trait loci, namely, qGPoL1, qGPoL2, qGPoW1, and qGPoW2, were mapped via GWAS analysis. Through the integration of gene function annotation, transcriptome sequencing, and expression pattern analysis, we identified Glyma.06G255000 and Glyma.13G007000 as the key determinants of the pod length and width in vegetable soybeans, respectively. Furthermore, two kompetitive allele-specific polymerase chain reaction (KASP) markers, namely, S06-42138365 (A/T) and S13_628331 (A/T), were developed and effectively validated in 27 vegetable soybean accessions. Overall, our research identified genes that regulate the pod length and width and determined KASP markers for molecular marker-assisted selection breeding. These findings have crucial implications for the improvement of soybean crops and can contribute to the development of efficient breeding strategies.

The E3 ubiquitin ligase RNF220 maintains hindbrain Hox expression patterns through regulation of WDR5 stability

The spatial and temporal linear expression of Hox genes establishes a regional Hox code, which is crucial for the antero-posterior (A-P) patterning, segmentation, and neuronal circuit development of the hindbrain. RNF220, an E3 ubiquitin ligase, is widely involved in neural development via targeting of multiple substrates. Here, we found that the expression of Hox genes in the pons was markedly up-regulated at the late developmental stage (post-embryonic day E15.5) in Rnf220-/- and Rnf220+/- mouse embryos. Single-nucleus RNA sequencing (RNA-seq) analysis revealed different Hox de-repression profiles in different groups of neurons, including the pontine nuclei (PN). The Hox pattern was disrupted and the neural circuits were affected in the PN of Rnf220+/- mice. We showed that this phenomenon was mediated by WDR5, a key component of the TrxG complex, which can be polyubiquitinated and degraded by RNF220. Intrauterine injection of WDR5 inhibitor (WDR5-IN-4) and genetic ablation of Wdr5 in Rnf220+/- mice largely recovered the de-repressed Hox expression pattern in the hindbrain. In P19 embryonal carcinoma cells, the retinoic acid-induced Hox expression was further stimulated by Rnf220 knockdown, which can also be rescued by Wdr5 knockdown. In short, our data suggest a new role of RNF220/WDR5 in Hox pattern maintenance and pons development in mice.

RADT-06. IDENTIFICATION OF NEW RADIATION RESPONSE PREDICTOR GENES FOR GLIOBLASTOMACRUCIAL CONTRIBUTION OF THE LUNATIC FRINGE GENE (LFNG)

Abstract Glioblastoma (GBM) is adults’ most common aggressive primary intracranial tumor. The current standard of care involves maximal safe surgery followed by radiochemotherapy. OBJECTIVE our study was to identify genes that predict response to radiochemotherapy. MATERIALS AND METHODS We collected molecular and clinical data, including primary glioma mRNA expression, from two international online databases - TCGA and CCGA. A total of 76 cases were examined, with 41 patients in the TCGA cohort and 35 in the CCGA cohort. The patients were divided into two groups before assessing the data. Group 1 included patients who were alive at the time of the analysis, and Group 2 included patients who had passed away. Then, we performed a single-center Moroccan transcriptomic study involving 28 cases to assess the impact of the individualized gene in our environment. RESULTS The following genes, namely SRPX, S100A16, LFNG, CD5, CMBL, and TCTN2, are among the top genes co-overexpressed in TCGA and CGGA patients. We conducted a multivariate COX survival test, taking into account known prognostic clinical parameters, and found that the LFNG gene is the only marker of poor prognosis independent of other clinical markers in TCGA and CGGA patients treated with radiotherapy. Our results indicate that patients who overexpress LFNG have elevated IFNg levels and low granzyme B levels, while the levels of perforin and granzyme A remain unaffected by LFNG variation. Furthermore, our results demonstrate positive correlations between LFNG gene expression and four immune checkpoints (PD1, PDL1, CTLA4, and VISTA), but negative correlations only with TIM3. CONCLUSION Our findings suggest that LFNG could serve as a useful biomarker for identifying patients with poor prognosis and that LFNG expression may be associated with specific immune checkpoint expression patterns. These results have important implications for the development of new therapeutic approaches in the treatment of TCGA and CGGA patients.

BIOM-20. EVALUATING THE POTENTIAL OF PSMA TARGETING IN CNS TUMORS: INSIGHTS FROM LARGE-SCALE TRANSCRIPTOME PROFILING

Abstract PURPOSE Prostate-specific membrane antigen (PSMA) is a well-established target in prostate cancer therapy that has shown potential as a theranostic target across non-central nervous system (CNS) and CNS tumor types. We aimed to investigate the pan-tissue expression pattern of the PSMA-encoding gene FOLH1 to assess whether transcriptome profiling can inform tumor theranostics. EXPERIMENTAL DESIGN We assessed FOLH1 expression from the Open Pediatric Cancer Project (OpenPedCan, n = 2132 specimens), The Cancer Genome Atlas (TCGA, n = 10411 specimens), and the Genotype Tissue Expression Project (GTEx, n = 17382 specimens) in relation to published reports of PSMA radionuclide uptake. RESULTS When comparing FOLH1 expression across tumor versus normal tissues, we found that non-CNS tumors exhibiting elevated expression of at least two-fold (FDR < 0.05) were reported to have successful PSMA radionuclide uptake in contrast to tumors with less than a two-fold elevation or with lower expression of FOLH1 relative to normal. Notably, CNS tumors universally exhibited lower expression of FOLH1 relative to normal brain tissue, but we observed considerable variation in the expression of blood-tumor barrier (BTB) components associated with reports of BTB integrity and uptake of PSMA radiotracers. CONCLUSIONS Large-scale transcriptomics data may help guide the application of PSMA-based radionuclide therapies in non-CNS tumors, but care should be taken to account for BTB effects in CNS tumors when assessing the potential for radionuclide success. Our analysis demonstrated that FOLH1 showed a lack of tumor-specific expression for both adult and pediatric CNS tumors when compared to normal brain tissue, suggesting that PSMA is not a desirable target in this disease type. This study highlights the utility of utilizing large-scale molecular data to guide the selection of theranostic targets.

CD47 in Osteosarcoma: Correlation with Metastasis and Macrophage-Mediated Phagocytosis

CD47 is expressed on cell surfaces and acts as a “don’t eat me” signal by interacting with signal-regulatory protein-α on the macrophage surface. Some cancer cells express CD47 protein and can evade macrophage phagocytosis. Herein, we evaluated the feasibility of targeting CD47 for osteosarcoma by analyzing its expression patterns, clinicopathological correlations, and immunotherapeutic potential. We performed a retrospective analysis on 24 biopsy samples from patients with osteosarcoma to investigate correlations between CD47 protein positivity and clinicopathological characteristics. CD47 protein expression was detected in 20.8% of the biopsy samples. CD47 positivity correlated with metastasis at diagnosis. Patients with CD47-positive tumors were older than those with CD47-negative tumors. However, CD47 protein expression was not associated with sex, tumor size, or histologic response to preoperative chemotherapy. In vitro, CD47 antibody (B6H12) did not affect osteosarcoma cell viability or apoptosis. In a wound-healing assay, CD47 inhibited the migration of osteosarcoma cells. Differentiated macrophages exhibited higher phagocytic activity against osteosarcoma cells when pretreated with B6H12 compared with the isotype control. Our preliminary data suggest a possible interaction between CD47 protein and macrophage phagocytosis in osteosarcoma metastasis. A better understanding of the role of CD47 is necessary to develop an innovative immunotherapeutic approach against osteosarcoma.

Aerial litter mimicry: A novel form of floral deception mediated by a monoterpene synthase

Abstract Floral mimics deceive their pollinators by developing visual and olfactory resemblance to various models. We report a flower that exhibits phenotypes like aerial litter and deceives an aerial litter specialist beetle to achieve pollination. We assessed the floral phenology and the effective pollinators of an Australian understorey treelet, Meiogyne heteropetala (Annonaceae). The similarities of morphology, colour and odour between the flowers and co‐occurring aerial litter were investigated. The terpene synthase involved in floral scent emission was identified by expression patterns and product profile. The behavioural responses of the pollinator to various odours were assessed using bioassays. The erotylid beetle Loberus sharpi is the most likely effective pollinator because it was the only pollen‐laden visitor during the pistillate phase. Loberus sharpi was exclusively found in aerial litter and M. heteropetala flowers. The flowers offer an honest shelter reward. The beetle also oviposits there, but most larvae eventually perished as the petals dropped onto the forest floor. The morphology and spectral reflectance of the flowers overlap with aerial litter. The floral scent was dominated by monoterpenes, especially 1,8‐cineole. The cineole synthase MhCINS was the only highly expressed floral terpene synthase and possessed a highly similar product profile to the floral scent composition. The volatile composition of M. heteropetala flowers is distinct from other congeners and highly similar to aerial litter, indicating advergence to aerial litter. Visual and odour resemblance, coupled with low larval survivorship, provides evidence that the beetles were deceived into pollinating the flowers. Behavioural experiments showed that the pollinator was attracted to both aerial litter and M. heteropetala flowers. The beetles were also attracted to 1,8‐cineole and synthetic mixes of floral odour and MhCINS products. The beetles were unable to distinguish floral scent from MhCINS products nor from 1,8‐cineole, suggesting MhCINS alone sufficed to attract the pollinator olfactorily. The beetles, however, preferred aerial litter over flowers. The beetles likely categorised the flower as a general, but not the most preferred, brood substrate. Synthesis. This study reports the first case of floral mimicry of aerial litter and characterises the biochemical process responsible for olfactory mimicry.

Human-Brain-Derived Ischemia-Induced Stem Cell Transplantation Is Associated with a Greater Neurological Functional Improvement Compared with Human-Bone Marrow-Derived Mesenchymal Stem Cell Transplantation in Mice After Stroke

The transplantation of injury/ischemia-induced stem cells (iSCs) extracted from post-stroke human brains can improve the neurological functions of mice after stroke. However, the usefulness of iSCs as an alternative stem cell source remains unclear. The current study aimed to assess the efficacy of iSC and mesenchymal stem cell (MSC) transplantation. In this experiment, equal numbers of human brain-derived iSCs (h-iSCs) (5.0 × 104 cells/μL) and human bone marrow-derived MSCs (h-MSCs) (5.0 × 104 cells/μL) were intracranially transplanted into post-stroke mouse brains after middle cerebral artery occlusion. Results showed that not only h-iSC transplantation but also h-MSC transplantation activated endogenous neural stem/progenitor cells (NSPCs) around the grafted sites and promoted neurological functional improvement. However, mice that received h-iSC transplantation experienced improvement in a higher number of behavioral tasks compared with those that received h-MSC transplantation. To investigate the underlying mechanism, NSPCs extracted from the ischemic areas of post-stroke mouse brains were cocultured with h-iSCs or h-MSCs. After coincubation, NSPCs, h-iSCs, and h-MSCs were selectively collected via fluorescence-activated cell sorting. Next, their traits were analyzed via microarray analysis. The genes related to various neuronal lineages in NSPCs after coincubation with h-iSCs were enriched compared with those in NSPCs after coincubation with h-MSCs. In addition, the gene expression patterns of h-iSCs relative to those of h-MSCs showed that the expression of genes related to synapse formation and neurotransmitter-producing neurons increased more after coincubation with NSPCs. Hence, cell–cell interactions with NSPCs promoted transdifferentiation toward functional neurons predominantly in h-iSCs. In accordance with these findings, immunohistochemistry showed that the number of neuronal networks between NSPCs and h-iSCs was higher than that between NSPCs and h-MSCs. Therefore, compared with h-MSC transplantation, h-iSC transplantation is associated with a higher neurological functional improvement, presumably by more effectively modulating the fates of endogenous NSPCs and grafted h-iSCs themselves.

Assessing expression patterns of PTGR1, a potential biomarker for acylfulven sensitivity in urothelial carcinoma

BackgroundMetastatic urothelial carcinoma (mUC) has a poor prognosis, despite recent therapeutic advancements. Prostaglandin Reductase 1 (PTGR1) is essential for activating acylfulvens, a promising class of drugs for treating a subset of urothelial carcinoma (UC) patients. The efficacy of acylfulvens depends on PTGR1 activity and defects in the nucleotide excision repair (NER) pathway, notably ERCC2 mutations present in 10–15% of bladder tumors. This study identifies patients eligible to be included in acylfulven clinical trials based on the presence of PTGR-1 by immunohistochemistry (IHC) staining, RNA expression level and mutations in the NER pathway. Additionally, it evaluates PTGR-1 expression as a prognostic biomarker.MethodsThree PTGR-1 antibodies were tested in a kidney cancer cell line A498 and in tissues with known PTGR1 expression (liver, tonsils, pancreas, small intestine, etc.). Patients with untreated mUC receiving 1st line platinum from Dec. 2019 to Dec. 2021 in the Capital Region, Denmark, were included retrospectively. FFPE tumor samples were collected, and tissue microarrays (TMAs) were constructed. TMAs were stained with the best-performing PTGR1 antibody, RNA expression was analyzed using Nanostring nCounter PanCancer panel and gene mutations were assessed using a targeted genomic panel (TSO-500). Kaplan-Meier and multivariate Cox regression assessed overall survival and covariate impacts.ResultsThe AB181131 PTGR1 antibody was the most reliable in validation tissues. Tumors from 71 mUC patients were used to construct the TMA, and 40% of tumors scored positive for PTGR1 by IHC staining. A normalized PTGR1 RNA cutoff at 2,550 normalized counts achieved an AUC of 0.9, defining 35 samples as positive with a sensitivity of 96% and specificity of 85% in relation to IHC-positivity. Differential expression showed a significant upregulation of PTGR1 RNA in PTGR1 IHC-positive cases. NER-deficiency and PTGR1 positivity (mutations in ERCC1, ERCC2, ERCC3, ERCC4) was seen in 9 patients (13%). Median overall survival was 16 months in the cohort. Overall survival (OS) analysis indicates that overexpression of PTGR1 RNA was associated with a reduced median OS (12 months vs. 25 months, p = 0.039, log-rank).ConclusionPTGR1 IHC staining pattern using the Abcam AB181131 antibody is highly correlated with PTGR1 RNA expression. 13% in our cohort were identified as NER deficient and PTGR1 positive. Lower levels of PTGR1 indicates better outcome in this cohort.

Das also war des Pudels Kern oder: „Geflügelte Worte“ als Phrasemklasse? – Problemfelder und begriffliche Präzisierung

Abstract As a class belonging to phraseology, winged words are characterized above all by the fact they are also used in other contexts as quotations, especially from literary texts (e.g., Grau, teurer Freund, ist alle Theorie from Goethes Faust I), thus acquiring a life of their own. However, they can be referred to the original sources in the consciousness of the language user. Forms of expression from areas other than literary areas, such as movies and television, advertising, etc., are increasingly gaining a comparable status in public language use. The imprinting of these expressions often stems from the fact that winged words are not only or no longer used as quotations but also used as a means of commenting on facts, evaluating them, etc., and at the same time calling up knowledge scenarios that refer to the original context of use as backgrounds for understanding. Over time, new patterns of expression may be established by modifying parts of the original formulation so that – expressed in construction grammar – new situationally adaptable form-meaning pairs can emerge. Based on this “phraseologization mechanism”, problematic aspects of phraseology theory are identified and discussed using the example of Das also war des Pudels Kern. Finally, suggestions are made for the conceptual sharpening of the category “winged words”.

The perspective of ceRNA regulation of circadian rhythm on choroidal neovascularization

Abnormal growth of blood vessels (choroidal neovascularization) can lead to age-related macular degeneration (AMD) and eventually cause vision loss due to detachment of the retinal pigmented epithelium. This indicates that choroidal neovascularization is important for the treatment of AMD. The circadian clock in the mammalian retina is responsible for controlling various functions of the retina, enabling it to adjust to changes in light and darkness. Recent studies have revealed a potential connection between the circadian clock and eye diseases, although a cause-and-effect relationship has not been definitively established. C57BL/6J male mice (aged 6 weeks) were randomly divided into two groups (Control group: 9:00–21:00 light period (300 lx); Jet lag group: 8-hour phase advance once every 4 days). A laser-induced CNV model was created after 2 weeks of feeding in a controlled or jet-lagged environment. Then, full transcriptome sequencing was performed. The pathways regulated by differentially expressed mRNAs were identified by GO analysis and GSEA. Further protein networks were constructed with the STRING database and Cytoscape software. WGCNA was used to further explore the co-expression modules of these differential genes and the correlation between these differential genes and phenotypes. ceRNA networks were constructed with miRanda and TargetScan. The pathways associated with the overlapping differentially expressed mRNAs in the ceRNA network were identified, and the hub genes were validated by qPCR. A total of 661 important DEGs, 31 differentially expressed miRNAs, 106 differentially expressed lncRNAs and 87 differentially expressed circRNAs were identified. GO and GSEA showed that the upregulated DEGs were mainly involved in reproductive structure development and reproductive system development. The STRING database and Cytoscape were used to determine the protein interaction relationships of these DEGs. WGCNA divided the expression of these genes into several modules and screened the hub genes of each module separately. Furthermore, a ceRNA network was constructed. GO analysis and GSEA showed that these target DEmRNAs mainly function in wound healing, cell spreading, epiboly involved in wound healing, epiboly, and morphogenesis of an epithelial sheet. Finally, ten key genes were identified, and their expression patterns were confirmed by real-time qPCR. In this study, we investigated the regulatory mechanism of ceRNAs in choroidal neovascularization according to different light-dark cycles in the eyeball.

From the genome's perspective: Bearing somatic retrotransposition to leverage the regulatory potential of L1 RNAs.

Transposable elements (TEs) are mobile genomic elements constituting a big fraction of eukaryotic genomes. They ignite an evolutionary arms race with host genomes, which in turn evolve strategies to restrict their activity. Despite being tightly repressed, TEs display precisely regulated expression patterns during specific stages of mammalian development, suggesting potential benefits for the host. Among TEs, the long interspersed nuclear element (LINE-1 or L1) has been found to be active in neurons. This activity prompted extensive research into its possible role in cognition. So far, no specific cause-effect relationship between L1 retrotransposition and brain functions has been conclusively identified. Nevertheless, accumulating evidence suggests that interactions between L1 RNAs and RNA/DNA binding proteins encode specific messages that cells utilize to activate or repress entire transcriptional programs. We summarize recent findings highlighting the activity of L1 RNAs at the non-coding level during early embryonic and brain development. We propose a hypothesis suggesting a mutualistic relationship between L1 mRNAs and the host cell. In this scenario, cells tolerate a certain rate of retrotransposition to leverage the regulatory effects of L1s as non-coding RNAs on potentiating their mitotic potential. In turn, L1s benefit from the cell's proliferative state to increase their chance to mobilize.

Uncovering functional lncRNAs by scRNA-seq with ELATUS

Long non-coding RNAs (lncRNAs) play fundamental roles in cellular processes and pathologies, regulating gene expression at multiple levels. Despite being highly cell type-specific, their study at single-cell (sc) level is challenging due to their less accurate annotation and low expression compared to protein-coding genes. Here, we systematically benchmark different preprocessing methods and develop a computational framework, named ELATUS, based on the combination of the pseudoaligner Kallisto with selective functional filtering. ELATUS enhances the detection of functional lncRNAs from scRNA-seq data, detecting their expression with higher concordance than standard methods with the ATAC-seq profiles in single-cell multiome data. Interestingly, the better results of ELATUS are due to its advanced performance with an inaccurate reference annotation such as that of lncRNAs. We independently confirm the expression patterns of cell type-specific lncRNAs exclusively detected with ELATUS and unveil biologically important lncRNAs, such as AL121895.1, a previously undocumented cis-repressor lncRNA, whose role in breast cancer progression is unnoticed by traditional methodologies. Our results emphasize the necessity for an alternative scRNA-seq workflow tailored to lncRNAs that sheds light on the multifaceted roles of lncRNAs.

Characterization of HTLV-1 Infectious Molecular Clone Isolated from Patient with HAM/TSP and Immortalization of Human Primary T-Cell Lines

Human T-cell leukemia virus (HTLV-1) is the etiological agent of lymphoproliferative diseases such as adult T-cell leukemia and T-cell lymphoma (ATL) and a neurodegenerative disease known as HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). While several molecular clones of HTLV-1 have been published, all were isolated from samples derived from patients with adult T-cell leukemia. Here, we report the characterization of an HTLV-1 infectious molecular clone isolated from a sample of a patient with HAM/TSP disease. Genetic comparative analyses of the HAM/TSP molecular clone (pBST) revealed unique genetic alterations and specific viral mRNA expression patterns. Interestingly, our clone also harbors characteristics previously published to favor the development of HAM/TSP disease. The molecular clone is capable of infection and immortalization of human primary T cells in vitro. Our studies further demonstrate that the HTLV-1 virus produced from primary T cells transfected with pBST or ACH molecular clones cannot sustain long-term expansion, and cells cease to proliferate after 3–4 months in culture. In contrast, long-term proliferation and immortalization were achieved if the virus was transmitted from dendritic cells to primary T cells, and secondary infection of 729B cells in vitro was demonstrated. In both primary T cells and 729B cells, pBST and ACH were latent, and only hbz viral RNA was detected. This study suggests that HTLV-1 transmission from DC to T cells favors the immortalization of latently infected cells.

Identification and Molecular Analysis of Proteins Associated with Non-spinning Disease in the Silkworm Bombyx mori L Induced by BmNPV and BmDNV

Aims: The present study combines morphological and molecular biology investigations to uncover the underlying causes of the non-spinning syndrome in silkworms. To understand the molecular mechanism and immune mechanism after infection of viral pathogen in silkworm Bombyx mori L. Study Design: Infected silkworm samples were collected from the field and isolated the genomic DNA and total proteins. Analyzed the samples by using genomic tools and proteomic tools. Place and Duration of Study: Central Sericultural Research and Training Institute, Mysuru, Karnataka. 01.03.2023 to 31.07.2023 total 04 months. Methodology: Our findings indicate that infections of BmDNV and BmNPV are associated with abnormal growth of the silk gland, leading to the non-spinning phenotype. To confirm pathogen infection in non-spinning silkworms, we collected genomic DNA and conducted PCR analysis using specific primers for DNV and NPV. Results: Our results showed amplification of both viral infections in the isolated samples. SDS-PAGE analysis revealed differential regulation of several proteins involved in physiological and immunological processes in the hemolymph of non-spinning silkworms. MALDI-TOF/MS was used to identify an up-regulated protein of interest, which was found to be a Lepidopteran low molecular weight lipoprotein. This protein is known to play an important role in insect immune mechanisms. Conclusion: Major disease-causing agents such as BmDNV and BmNPV infections and investigate their impact on protein expression patterns in the hemolymph, shedding light on the factors contributing to non-spinning behavior in silkworms.

Exposure to High Concentrations of Tetrabromobisphenol A Slows the Process of Tissue Regeneration and Induces an Imbalance of Metabolic Homeostasis in the Regenerated Intestines of Apostichopus japonicus

Background: Tissue regenerative capacity following evisceration, potentially influenced by environmental contaminants and intestinal microflora, is essential for the financial success of Apostichopus japonicus farming. However, the morphological structure, gut microbiome composition, and genes expression pattern of the regenerated gut after exposure to high levels of TBBPA remain poorly unclear. Methods: In this research, the effect of TBBPA exposure on tissue regeneration in A. japonicus was investigated through a comprehensive multi-omics approach. Results: Our results showed that the integrity, the intestinal wall thickness, and the villi length of the regenerated intestines in A. japonicus decreased after treatment with high levels of TBBPA. The findings from PCoA and NMDS analyses revealed that the microbial community composition was significantly altered following exposure to high concentrations of TBBPA in the regenerated intestines of A. japonicus. The KEGG pathway enrichment analysis indicated that the DEGs (differentially expressed genes) were predominantly enriched on metabolism and immunity-related signaling pathways after exposure to high levels of TBBPA. These included pathways involved in the PPAR signaling pathway, ECM receptor interaction, glycerolipid metabolism, and fatty acid degradation. Interestingly, the results have demonstrated that there are 77 transcript factors that were significantly different after exposure to TBBPA. Conclusions: These results suggested that high levels of exposure to TBBPA induces an imbalance of the metabolic homeostasis by regulating the expression levels of transcription factors in the regenerated intestines of A. japonicus.