Similar Expression Patterns Research Articles

Temporal Changes Toward Cellular Senescence in Rat Dental Pulp Stem Cells Induced by Long-Term In Vitro Culture

Rat dental pulp stem cells (DPSCs) can be used to elucidate mesenchymal stem cell (MSC) applications in regenerative medicine. However, information on rat DPSCs during long-term passage, which could lead to replicative senescence, is limited. In this study, we investigated the phenotypic changes in DPSCs after 3–26 passages (3P–26P). The results show that cell morphology and nuclear size increase proportionally with passage number. The phosphorylated histone H2A.X (γ-H2A.X) positive cells (indicating DNA damage) increased significantly earlier than the 4-Hydroxynonenal (4-HNE) stained cells (indicating an abundance of intracellular reactive oxygen species). Compared to the cells subjected to 3P and 5P, the cells subjected to 15P showed reduced proliferation despite being positive for Ki67. Furthermore, cell growth was completely arrested after 26P. The senescence markers, senescence-associated β-galactosidase (SA-β-gal) and p16, exhibited similar expression patterns that were not correlated with those of p21 and urokinase-type plasminogen activator receptor (uPAR). Nearly all cells expressed SA-β-gal and p16 after 26P, whereas only half expressed p21 and uPAR. These results will contribute to understanding the characteristics of DPSCs toward replicative senescence, which are applicable to elucidate mechanisms related to regenerative medicine and stem cell aging.

Evolutionary changes leading to efficient glymphatic circulation in the mammalian brain

The functional significance of the morphological and genetic changes that occurred in the brain during evolution is not fully understood. Here we show the relationships between evolutionary changes of the brain and glymphatic circulation. We establish a mathematical model to simulate glymphatic circulation in the cerebral hemispheres, and our results show that cortical neurons accumulate in areas of the cerebral hemispheres where glymphatic circulation is highly efficient. We also find that cortical folds markedly enhance the efficiency of glymphatic circulation in the cerebral hemispheres. Furthermore, our in vivo study using ferrets reveals sulcus-dominant cerebrospinal fluid (CSF) influx, which enhances the efficiency of glymphatic circulation in the enlarged cerebral hemispheres of gyrencephalic brains. Sulcus-dominant CSF influx is mediated by preferential expression of aquaporin-4 in sulcal regions, and similar expression patterns of aquaporin-4 are also found in human cerebral hemispheres. These results indicate that evolutionary changes in the cerebral hemispheres are related to improved efficiency of glymphatic circulation. It seems plausible that the efficiency of glymphatic circulation is an important factor determining the evolutionary trajectory of the cerebral hemispheres.

Two NPC1 homologous proteins are involved in asexual reproduction, pathogenicity, and lipid trafficking in Phytophthora sojae

Niemann-Pick type C (NPC) disease is characterized by lysosomal lipid storage disorders and defects in lipid trafficking, primarily due to mutations in the NPC1 protein. Two NPC1 homologous proteins are present in the genome of Phytophthora sojae, named as PsNPC1-1 and PsNPC1-2. Both proteins exhibit high sequence identity, consistent conserved functional domains, similar gene expression patterns, and comparable subcellular localization. Deletion of a single PsNPC1 gene did not result in significant phenotypic changes. However, simultaneous deletion of both PsNPC1 genes led to reduced mycelial growth, decreased sporangial production, impaired pathogenicity, and an inability to release normal zoospores in P. sojae. Furthermore, dysfunction of PsNPC1s did not completely block the absorption and utilization of exogenous sterols by P. sojae. While lipidome analysis revealed that the relative contents of fatty acyls, sphingolipids and saccharolipids were significantly elevated in the double-gene deletion mutant, alongside obvious alterations in glycerophospholipid and glycerolipid metabolism. Additionally, we observed a significant down-regulation of PsCDP-AP protein along with its interactions with both PsNPC1s. Deletion of PsCDP-AP also impaired asexual reproduction and virulence of P. sojae. These findings demonstrate that both PsNPC1 proteins may collaborate with other key regulators to modulate asexual reproduction, pathogenicity and lipid trafficking in P. sojae.

Genome-wide association study identified BnaPAP17 genes involved in exogenous ATP utilization and regulating phosphorous content in Brassica napus.

BnaPAP17s associated with root-secreted APases activity were identified by genome-wide association study, and those were induced by Pi-deficiency. BnaPAP17s were involved in improving exogenous organophosphorus utilization as secreted APases. Deficiency of available phosphorus (P) in soil has become an important limiting factor for yield and quality in oilseed rape (Brassica napus). In many soils, organic P (Po) is the main component of the soil P pool. Po must be hydrolyzed to inorganic P (Pi) through acid Phosphatase (APases), and then taken up by plants. However, root-secreted APases (SAP) activity, as a quantitative trait, plays an important role in soil Po utilization; those genetic loci are not clear in B. napus. In this study, we performed a genome-wide association study for SAP activity under Pi-deficiency using a panel of 350 accessions of B. napus and more than 4.5 million polymorphic single nucleotide polymorphisms (SNPs). Thirty-five significant SNPs associated with SAP activity were identified. BnaA01.PAP17 (BnaA01g27810D) was a candidate gene underlying lead SNP (ChrA01_19576615). We experimentally verified that both BnaA01.PAP17 and its three homologous genes had similar expression pattern in response to Pi-deficiency. The dynamic changes in BnaPAP17s expression level were opposite to those of Pi concentration in both roots and leaves, suggesting their potential utility as Pi marker genes in B. napus. Transient expression of BnaPAP17s in tobacco leaves proved that BnaPAP17s were located in the apoplast as secreted APases. The overexpression of BnaPAP17s enhanced SAP activity in response to Pi-deficiency and resulting in increased P content in plants when ATP was supplied as the sole P resource. Taken together, these results suggest that BnaPAP17s contributed to SAP activity, thus having a function in extracellular Po utilization in B. napus.

Glypican-3 and Cytokeratin-19 Expression in Pancreatic Cancer in a Canadian Population.

Background/Objectives: One study of pancreatic ductal adenocarcinoma has found expression of glypican-3 (GPC3) and cytokeratin-19 (CK19) determined by immunohistochemistry to be associated with higher stage and grade disease, with a more adverse prognosis. The reported 44% rate of GPC3 expression in pancreatic cancer raises the important possibility that targeted immunotherapies currently in development for hepatocellular carcinoma may also prove useful for GPC3-expressing pancreatic cancers. The present study aims to determine if a similar expression pattern of these markers and stage/grade/prognostic associations is present in our Canadian patient population. Methods: Patients with a pancreatic surgical resection for adenocarcinoma or neuroendocrine tumor (NET) were identified from pathology records over a 5-year period. Immunohistochemistry for GPC3 and CK19 was performed on archived tumor tissue and the proportion of positive cells and intensity of staining were recorded. Grade, stage, and overall survival were compared in patients with NETs that were CK19-positive versus -negative. Results: All 72 pancreatic adenocarcinomas and 20 NETs tested were negative for GPC3, apart from a single case of pancreatic adenocarcinoma. All 72 adenocarcinomas were positive for CK19 expression. Half of the NETs were positive for CK19. There was no correlation between CK19 expression in NETs and tumor grade, lymph node metastasis, distant metastasis, or overall survival. Conclusions: We are skeptical of the reported prognostic value of GPC3 and CK19 in pancreatic adenocarcinomas. CK19 as a prognostic marker in NETs has potential for further study. The results with our protocol for GPC3 immunohistochemistry suggest that pancreatic cancer may be a less promising target for GPC3-targeted immunotherapies than previously thought.

Proteomic profiling of oral squamous cell carcinoma tissues reveals altered immune-related proteins: implications for personalized therapy

ABSTRACT Objectives Oral squamous cell carcinoma poses a substantial global health challenge marked by rising mortality rate. Recently, immunotherapy has shown promising results in cancer management by enhancing immune response. Thus, identifying additional immune-related markers is critical for advancing immunotherapy treatments. Methods Data-independent acquisition (DIA) mass spectrometry approach was used to explore differentially expressed immune-related proteins in oral cancer tissues compared to adjacent non-cancerous tissues. Functional significance was identified through Gene Ontology, pathway, and network analysis. Gene expression of identified proteins was validated using transcriptomic data. Results DIA analysis identified 29,459 precursors corresponding to 3429 proteins. Among these, 1060 proteins were differentially expressed, with 166 being immune-related. Differentially regulated proteins were involved in innate immune response, mitochondrial ATP synthesis, and neutrophil degranulation. Pathway analysis of immune-related proteins showed perturbation in anti-tumor immunity-related pathways such as interferon signaling, TCR signaling, PD-1 signaling, and antigen processing and presentation. Significance of these pathways was further reinforced by the strong interactions identified in the protein–protein interaction network analysis. Additionally, gene expression analysis showed similar mRNA expression patterns for key proteins involved in altered pathways, including ISG15, IFIT1/3, HLA-A/C and OAS2/3. Conclusions Further validation of these proteins could establish them as potential targets for personalized therapy.

Transcriptome analyses at specific plastochrons reveal timing and involvement of phytosulfokine in maize vegetative phase change

Successive developmental stages of representative early and late juvenile, transition, and adult maize leaves were compared using machine-learning-aided analyses of gene expression patterns to characterize vegetative phase change (VPC), including identification of the timing of this developmental transition in maize. We used t-SNE to organize 32 leaf samples into 9 groups with similar patterns of gene expression. oposSOM yielded clusters of co-expressed genes from key developmental stages. TO-GCN supported a sequence of events in maize in which germination-associated ROS triggers a JA response, both relieving oxidative stress and inducing miR156 production, which in turn spurs juvenility. Patterns of expression of MIR395, which regulates sulfur assimilation, led to the hypothesis that phytosulfokine, a sulfated peptide, is involved in the transition to adult patterns of differentiation.

Generation of Functioning Platelets from Mature Megakaryocytes Derived from CD34+ Umbilical Cord Blood Cells.

Clinically patients with thrombocytopenia are in urgent need of platelet transfusion, thus it is necessary to produce the platelets in large scale in vitro to meet the clinical needs. In this study, we developed efficient protocol to generate functioning platelets by differentiating umbilical cord blood (CB)-derived CD34+ cells into mature megakaryocytes. Under our condition, up to 85% of mature megakaryocytes were generated from CB-derived CD34+ cells, and over 75% CD42b+CD62p+ platelets were produced. The megakaryocytes at day 12 after the differentiation had the similar gene expression pattern to natural mature megakaryocytes, and AMPK and insulin signal pathway were activated to inhibit the apoptosis and benefit platelet release. There were up to 72% of the platelets that could bind with PAC1, which is the highest rate of CB CD34+ cell-derived platelets to play function in vitro to date. The recovery of hemostasis and coagulation was similar in thrombocytopenia mice injected with CB CD34+ cell-derived platelets and with human blood-derived platelets, respectively, and it is the first time to demonstrate that human CB CD34+ cell-derived platelets were functional invivo. Therefore, our findings open a new avenue to provide an in vitro efficient approach to generate functional platelets for clinical applications.

Leukemia inhibitory factor receptor inhibition by EC359 reduces atherosclerotic stenosis grade in Ldlr−/− mice

Cytokines are involved in all stages of atherosclerosis, generally contributing to disease progression. Previously, members of the Interleukin (IL)-6 cytokine family, such as IL-6, oncostatin M, and cardiotrophin-1, have been extensively studied in atherosclerosis. However, the role of leukemia inhibitory factor (LIF), member of the IL-6 family, and its receptor (LIFR), remains to be further elucidated. Therefore, the aim of this study is to provide insight in LIF receptor signalling in atherosclerosis development.Single-cell RNA sequencing analysis of human carotid artery plaques revealed that mast cells highly express LIF, whereas LIFR was specifically expressed on activated endothelial cells. A similar expression pattern of Lifr was observed in mouse atherosclerotic plaques. Next, female Western-type diet fed Ldlr−/− mice were treated with LIF receptor inhibitor EC359 (5 mg/kg s.c., n = 15) or control solvent (n = 15) three times per week for eight weeks. Stenosis grade was reduced in the aortic root of EC359 treated mice compared to control mice, but treatment did not affect plaque composition. Serum cholesterol levels were significantly reduced in EC359 treated mice, likely attributed to a reduction in VLDL cholesterol levels. Furthermore, LIF receptor inhibition reduced Pecam1 and Vcam1 expression in the aorta. Consequently, immune cell infiltration was reduced in aortic plaques of EC359 treated mice compared to control mice.Conclusively, we demonstrated that LIF receptor is a potential therapeutic target in atherosclerosis by reducing plaque size, attributed to lower serum cholesterol levels, reduced endothelial activation and less immune cell infiltration in the plaque.

The brain atlas of a subsocial bee reflects that of eusocial Hymenoptera.

The evolutionary transition from solitary life to group-living in a society with cooperative brood care, reproductive division of labor and morphological castes is associated with increased cognitive demands for task-specialization. Associated with these demands, the brains of eusocial Hymenoptera divide transcriptomic signatures associated with foraging and reproduction to different populations of cells and also show diverse astrocyte and Kenyon cell types compared with solitary non-hymenopteran insects. The neural architecture of subsocial bees, which represent evolutionary antecedent states to eusocial Hymenoptera, could then show how widely this eusocial brain is conserved across aculeate Hymenoptera. Using single-nucleus transcriptomics, we have created an atlas of neuron and glial cell types from the brain of a subsocial insect, the small carpenter bee (Ceratina calcarata). The proportion of C. calcarata neurons related to the metabolism of classes of neurotransmitters is similar to that of other insects, whereas astrocyte and Kenyon cell types show highly similar gene expression patterns to those of eusocial Hymenoptera. In the winter, the transcriptomic signature across the brain reflected diapause. When the bee was active in the summer, however, genes upregulated in neurons reflected foraging, while the gene expression signature of glia associated with reproductive functions. Like eusocial Hymenoptera, we conclude that neural components for foraging and reproduction in C. calcarata are compartmentalized to different parts of its brain. Cellular examination of the brains of other solitary and subsocial insects can show the extent of neurobiological conservation across levels of social complexity.

Genome-wide identification and functional analysis of CYP450 genes in eggplant (Solanum melongena L.) with a focus on anthocyanin accumlation

BackgroundIn plants, Cytochrome P450 (CYP450) constitutes the largest family of metabolic enzymes and plays a crucial role in various physiological processes, including plant growth and development. Eggplants are well-known for their peel’s high concentration of anthocyanin compounds that provide significant health benefits to humans. The accumulation of anthocyanins in eggplant peels is an important process during growth and development. Therefore, it is essential to identify the CYP450 genes in eggplant (SmCYPs) and analyze their expression profiles during the period of anthocyanin accumulation in the peel.ResultsA total of 180 SmCYPs were identified in the eggplant genome and classified into eight subfamilies based on phylogenetic analysis. These SmCYPs exhibited highly conserved gene structure (exon/intron) and protein motifs, especially within their respective subgroup. Sixteen pairs of genes with collinearity were identified through gene duplication analysis. Promoter cis-acting element analysis revealed that SmCYPs are involved in various responses, including growth and development, stress responsiveness, and light responsiveness. Transcriptome data analysis revealed that all SmCYPs were expressed in various eggplant tissues, such as roots, stems, leaves, flowers, and fruits; with diverse expression patterns among members. The expression patterns of SmCYPs in eggplant peel also exhibited diversity during different stages of anthocyanin accumulation. qRT-PCR analysis demonstrated similar expression patterns for 15 selected SmCYPs as observed in the transcriptome data. Metabolomics analysis further suggested that SmCYPs are involved in the biosynthesis of secondary metabolites and metabolic pathways related to flavonoid and flavone/flavonol biosynthesis. Notably, three specific SmCYPs (SmCYP73A1/75A/98A1) play a significant role in flavonoid biosynthesis, particularly in anthocyanin synthesis in eggplant.ConclusionGenome-wide identification, phylogenetic analysis, expression profile analysis, and exploration of metabolic pathways related to SmCYPs provide valuable insights into the roles of these genes in anthocyanin accumulation in various tissues and organs, including eggplant peel. The findings from this study lay a foundation for the functional analysis of SmCYPs involvement in anthocyanin accumulation in eggplant peel, providing a molecular basis for breeders to cultivate novel varieties of eggplants with high levels of anthocyanin content.

Genome-Wide Identification of the Oxidative Stress 3 (OXS3) Gene Family and Analysis of Its Expression Pattern During Ovule Development and Under Abiotic Stress in Cotton.

Oxidative Stress 3 (OXS3) encodes a plant-specific protein that makes great contributions to a plant's stress tolerance. However, reports on genome-wide identification and expression pattern analyses of OXS3 were only found for Arabidopsis, wheat, and rice. The genus Gossypium (cotton) serves as an ideal model for studying allopolyploidy. Therefore, two diploid species (G. raimondii and G. arboreum) and two tetraploid species (G. hirsutum and G. barbadense) were chosen in this study for a bioinformatics analysis, resulting in 12, 12, 22, and 23 OXS3 members, respectively. A phylogenetic tree was constructed using 69 cotton OXS3 genes alongside 8 Arabidopsis, 10 rice, and 9 wheat genes, which were classified into three groups (Group 1-3). A consistent evolutionary relationship with the phylogenetic tree was observed in our structural analysis of the cotton OXS3 genes and the clustering of six conserved motifs. Gene duplication analysis across the four representative Gossypium species suggested that whole-genome duplication, segmental duplication, and tandem duplication might play significant roles in the expansion of the OXS3 gene family. Some existing elements responsive to salicylic acid (SA), jasmonic acid (JA), and abscisic acid (ABA) were identified by cis-regulatory element analysis in the promoter regions, which could influence the expression levels of cotton OXS3 genes. Furthermore, the expression patterns of the GhOXS3 gene were examined in different tissues or organs, as well as in developing ovules and fibers, with the highest expression observed in ovules. GhOXS3 genes exhibited a more pronounced regulatory response to abiotic stresses, of which ten GhOXS3 genes showed similar expression patterns under cold, heat, salt, and drought treatments. These observations were verified by quantitative real-time PCR experiments. These findings enhance our understanding of the evolutionary relationships and expression patterns of the OXS3 gene family and provide valuable insights for the identification of vital candidate genes for trait improvement in cotton breeding.

Identification of Defined Molecular Subgroups on the Basis of Immunohistochemical Analyses and Potential Therapeutic Vulnerabilities of Pulmonary Carcinoids

IntroductionMulti-omic studies have identified three molecular separated pulmonary carcinoid (PC) subgroups (A1, A2, B) with distinctive mRNA expression profiles (e.g., orthopedia homeobox protein [OTP]/achaete-scute homolog [ASCL1]/hepatocyte nuclear factor 1 homeobox A [HNF1A]). We aimed to establish an immunohistochemical (IHC) biomarker panel that enables subgroup identification, and assessment of its potential clinical relevance. MethodsAll patients with resected pulmonary carcinoids (2003–2012) were identified from the Dutch Cancer/Pathology Registry, and tumors were revised. The IHC expression of OTP/ASCL1/HNF1A was scored in a blinded fashion in a mRNA-profiled (n = 5/subgroup) and national carcinoid cohort (n = 478). The expression of potential therapeutic targets (somatostatin receptor type 2a [SSTR2A]/delta-like canonical Notch ligand 3 [DLL3]) was assessed. Immunohistochemistry was assessed using H-scoring. ResultsOTP/ASCL1/HNF1A reported similar IHC and mRNA expression patterns in the matched primary samples. In the national cohort, IHC separated PCs into subgroups A1 (n = 224 [53%], OTPhigh/ASCL1high/HNF1Alow), A2 (n = 161 [38%], OTPhigh/ASCL1low/HNF1Ahigh), and B (n = 37 [9%], OTPlow/ASCL1low/HNF1Ahigh). In 12% of PCs, no distinct classification could be provided. Patients with A1 were enriched for older age (83% > 50 y), female individuals (83%), and peripheral location (55%) with low SSTR2A (median = 10) and high DLL3 (median = 52) expression. A2 included younger patients (34% < 40 y) and endobronchial/central (87%) tumors with high SSTR2A (median = 160), but low DLL3 (median 0) expression. Group B included more male individuals (59%) and recurrence was more frequent (19%) than in groups A1 (8%) and A2 (6%). Neuroendocrine cell hyperplasia was enriched in A1 (25%) compared with A2 (3%) and B (0%). ConclusionsAn OTP/ASCL1/HNF1A IHC panel enables the identification of molecular-defined pulmonary carcinoid subgroups with distinct clinical phenotypes and diverging therapeutic vulnerabilities that require further prospective evaluation.

Spatial and temporal expression analysis of BMP signal modifiers, Smoc1 and Smoc2, from postnatal to adult developmental stages in the mouse testis

Smoc1 and Smoc2, members of the SPARC family of genes, encode signaling molecules downstream of growth factors such as the TGF-β, FGF, and PDGF families. Smoc1 has been implicated in playing a crucial role in microphthalmia with limb anomalies in humans and mice, while Smoc2 deficiency causes dental developmental defects. Although developmental cytokines/growth factors including TGF-β superfamily have been shown to play critical roles in postnatal spermatogenesis, there are no reports analyzing the spatial and temporal expression of Smoc1 and Smoc2 in the postnatal testis. In this study, we investigated the mRNA and protein expression of Smoc1 and Smoc2 in neonatal, juvenile, and adult mouse testes by RNA in situ hybridization, immunofluorescence, and single-cell RNA-seq analysis. We show that Smoc1 and Smoc2 have distinct expression patterns in male germ cells: Smoc1 is more highly expressed than Smoc2 in the germline. In contrast, Smoc2 is highly expressed in testicular somatic cells from neonatal to juvenile stages. The Smoc2-expressing cells then switch from somatic cells to germ cells in adults. Thus, although SMOC1 and SMOC2 proteins are structurally very similar, their spatial and temporal expression patterns in the postnatal testis differ significantly, suggesting their distinct roles in reproduction.

ZmKTF1 promotes salt tolerance by mediating RNA-directed DNA methylation in maize.

The epigenetic process of RNA-directed DNA methylation (RdDM) regulates the expression of genes and transposons. However, little is known about the involvement of RdDM in the response of maize (Zea mays) to salt stress. Here, we isolated a salt-sensitive maize mutant and cloned the underlying gene, which encodes KOW DOMAIN-CONTAINING TRANSCRIPTION FACTOR1 (KTF1), an essential component of the RdDM pathway. Evolutionary analysis identified two homologs of KTF1 (ZmKTF1A and ZmKTF1B) with highly similar expression patterns. Whole-genome bisulfite sequencing revealed that mutations in ZmKTF1 substantially decrease genome-wide CHH (H = A, C, or T) methylation levels. Moreover, our findings suggest that ZmKTF1-mediated DNA methylation regulates the expression of multiple key genes involved in oxidoreductase activity upon exposure to salt, concomitant with increased levels of reactive oxygen species. In addition, insertion-deletion mutations (InDels) in the promoter of ZmKTF1 affect its expression, thereby altering Na+ concentrations in seedlings in a natural maize population. Therefore, ZmKTF1 might represent an untapped epigenetic resource for improving salt tolerance in maize. Overall, our work demonstrates the critical role of ZmKTF1 involved in the RdDM pathway in maize salt tolerance.

Exploring the longitudinal expression dynamics of midguts in adult female Amblyomma americanum ticks

BackgroundFemale ticks remain attached to their host for multiple days to complete a blood meal. This prolonged feeding period is accompanied by a significant increase in the tick’s size and body weight, paralleled by noteworthy changes to the tick midgut. While the midgut is recognized for its established role in blood storage and processing, its importance extends to playing a crucial role in the acquisition, survival, and proliferation of pathogens. Despite this, our overall understanding of tick midgut biology is limited.ResultsOur transcriptome analysis identified 15,599 putative DNA coding sequences (CDS), which were classified into 26 functional groups. Dimensional and differential expression analyses revealed four primary transcriptional profiles corresponding to unfed, slow-feeding, transitory (from slow- to rapid-feeding), and rapid-feeding stages. Additionally, comparing the current dataset with previously deposited transcriptome from other tick species allowed the identification of commonly expressed transcripts across different feeding stages.ConclusionOur findings provide a detailed temporal resolution of numerous metabolic pathways in the midgut of A. americanum adult females throughout the feeding process, highlighting the dynamic transcriptional regulation of the tick’s midgut as feeding progresses. Furthermore, we identified conserved transcripts across three different tick species that exhibit similar expression patterns. This knowledge not only enhances our understanding of the physiological processes within the tick midgut but also opens up potential avenues for developing control methods that target multiple tick species.

Molecular characterization of CeOLE6, a diverged SH oleosin gene, preferentially expressed in Cyperus esculentus tubers.

CeOLE6, a tuber-specific gene in tigernut, encodes a diverged SH oleosin that functions in oil accumulation via homo and heteromultimerization. Tigernut (Cyperus esculentus L.) is a rare example accumulating high levels of triacylglycerols (TAGs) in underground tubers; however, the mechanism underlying is poorly understood. Given essential roles of oleosins (OLEs) in oil accumulation, in this study, structural and functional analyses were conducted for CeOLE6, an oleosin gene preferentially expressed in tigernut tubers. Phylogenetic analysis revealed that CeOLE6 encodes a diverged oleosin in Clade SH, which also includes CeOLE4 and -5. Further synteny analysis and sequence comparison indicated that CeOLE6 is more likely to be a whole-genome duplication (WGD) repeat of CeOLE4, which underwent rapid evolution and deletion of the typical C-terminal insertion for SHs. Nevertheless, CeOLE6 retains the capacity of oligomerization and oil accumulation, because (i) CeOLE6 could not only interact with itself but also with CeOLE2 and -5, two tuber-dominant members belonging to Clades SL and SH, respectively, and (ii) overexpressing CeOLE6 in tobacco leaves could significantly enhance the TAG content. Though CeWRI1 exhibits a similar expression pattern as CeOLE6 during tuber development, both CeWRI1 and -3 could not activate the CeOLE6 promoter, implying that they are not transcription factors contributing tuber-specific activation of CeOLE6. These findings not only provide insights into CeOLE genes in tuber oil accumulation, but also lay a foundation for further genetic improvement in tigernut and other species.

No two clones are alike: characterization of heterologous subpopulations in a transgenic cell line of the model diatom Phaeodactylum tricornutum

BackgroundConjugation-based episome delivery is a highly efficient method used to transfer DNA into the diatom Phaeodactylum tricornutum, facilitating the production of recombinant proteins and high-value metabolites. However, previous reports have indicated phenotypic heterogeneity among individual cells from clonally propagated exconjugant cell lines, potentially affecting the stability of recombinant protein production in the diatom.ResultsHere, we characterized the differences between subpopulations with distinct fluorescence intensity phenotypes derived from a single exconjugant colony of P. tricornutum expressing the enhanced green fluorescent protein (eGFP). We analyzed the expression cassette sequence integrity, plasmid copy number, and global gene expression. Our findings reveal that lower copy numbers and the deletion of the expression cassette in part of the population contributed to low transgene expression. Gene co-expression analysis identified a set of genes with similar expression pattern to eGFP including a gene encoding a putative Flp recombinase, which may be related to variations in fluorescence intensity. These genes thus present themselves as potential candidates for increasing recombinant proteins production in P. tricornutum episomal expression system.ConclusionsOverall, our study elucidates genetic and transcriptomic differences between distinct subpopulations in a clonally propagated culture, contributes to a better understanding of heterogeneity in diatom expression systems for synthetic biology applications.

Graph attention automatic encoder based on contrastive learning for domain recognition of spatial transcriptomics

Spatial transcriptomics is an emerging technology that enables the profiling of gene expression in tissues while preserving spatial location information. This innovative approach is anticipated to provide a comprehensive understanding of the spatial distribution of different cells within tissues and facilitate in-depth analysis of tissue structure. To accurately recognize spatial domains from spatial transcriptomics, we have introduced a generalized deep learning method called GAAEST (Graph Attention-based Autoencoder for Spatial Transcriptomics). Our proposed approach effectively integrates both spatial location information and gene expression data from spatial transcriptomics. Specifically, it leverages spatial location details to construct a neighborhood graph and employs a graph attention network-based encoder to embed gene expression information into a spatially informed space. At the same time, to further optimize the learned potential embedding, self-supervised contrastive learning is introduced to capture spatial information at three levels: local, global and contextual feature of spots. Finally, the decoder reconstructs gene expressions, which are then clustered to identify spatial domains with similar expression patterns and spatial proximity. Based on our experiments conducted on multiple datasets, GAAEST consistently outperforms existing state-of-the-art methods. The proposed GAAEST demonstrates excellent capabilities in spatial domain recognition, positioning it as an ideal tool for advancing spatial transcriptomics research.

A novel investigation of NANOG and POU5F1 associations in the pluripotent characterization of ES-like and epiblast cells

The transcription factors NANOG and POU5F1 (OCT4) play crucial roles in maintaining pluripotency in embryonic stem (ES) cells. While their functions have been well-studied, the specific interactions between NANOG and POU5F1 and their combined effects on pluripotency in ES-like and Epiblast cells remain less understood. Understanding these associations is vital for refining pluripotent stem cell characterization and advancing regenerative medicine. In this matter, we investigated the associations between NANOG and POU5F1 in maintaining pluripotency in ES-like and Epiblast cells and how these interactions contribute to the distinct pluripotent states of these cells. In the present paper, we examined the pattern of NANOG expression by the immunocytochemical method in embryonic stem-like (ES-like) cells and compared it with its expression pattern in embryonic stem cells (ESCs). Similarly, we examined the expression pattern of POU5F1 in ES-like cells, ESCs, and epiblast cells and compared the expression pattern of these two genes with each other. On the other hand, using Fluidigm Biomark system analysis, we compared the amount of NANOG mRNA in these three cell lines and differentiated and undifferentiated Spermatogonial stem cells in several passages. Microscopic observations indicated the cytoplasmic expression of NANOG in the considered cells; moreover, they showed a similar expression pattern of NANOG with POU5F1 in the experimented cells. It has also been suggested that the more limited the cell’s pluripotency, the lower the expression of these two genes. However, the decrease in NANOG expression is less than that of POU5F1. Fluidigm real-time RT-PCR analysis also confirmed these results. During the experimental process, protein-protein (PPI) network analysis shows a significant association of NANOG with other stem cell proteins, such as POU5F1. Our findings reveal distinct yet overlapping roles of NANOG and POU5F1 in maintaining pluripotency in ES-like and Epiblast cells. The differential binding patterns and functional interactions between these factors underscore the complexity of pluripotency regulation in different stem cell states. This study provides new insights into the molecular mechanisms governing pluripotency and highlights potential targets for enhancing stem cell-based therapies.