Spatiotemporal Expression Patterns Research Articles

Multi-omics integration analysis identifies pathogenic candidate genes of congenital heart defects with vascular malformation

Abstract Background Congenital heart defects (CHD) with vascular malformation harbors numerous genetic and epigenetic aberrations of DNA copy numbers and methylation. Proteomic deregulation by the aberrations plays key driver roles in heterogeneous progression of CHD. However, it still remains largely unknown about integrative analysis of genetic, epigenetic regulation and proteome in CHD. Purpose We employed a multi-omics approach to explore pathogenic candidate genes in CHD with vascular malformation. Methods Previously, we gathered WES data for 216 CHD patients with vascular malformation and 100 healthy controls. Meanwhile, we carried out DNA methylation analysis from 11 patients and 5 controls. Here, we profile proteomics of vascular tissues from 11 patients and 5 controls. Integration analysis of DNA copy-number-correlated (CNVcor) genes, methylation-correlated (METcor) genes and differentially proteins reveal the innovative and crucial pathogenetic genes. We then detected spatiotemporal expression patterns of candidate genes in zebrafish embryos using in situ hybridization and qPCR, and verified the roles in vascular development by morpholino-mediated gene knockdown. Results 90 differentially expressed proteins (DEPs) were obtained using proteomics and were mainly related to cell adhesion and migration, angiogenesis, amino acid metabolism, and HIF-1 signaling pathway through GO and KEGG pathway enrichment analysis. 15 hub proteins were identified by WGCNA analysis (Fig 1). Additionally, we found that the frequencies of the aberrancies of the CNVcor genes were significantly correlated with the METcor genes (Fig 2 C). We obtained 1924 differentially CNVcor genes and 952 differentially METcor genes. Then, we computed the Pearson’s correlation coefficients of all three-omics by pairwise comparison and found that the distribution of correlations was bimodal (Fig 2 D). Meanwhile, 28 overlapped genes were observed among three-omics. We screened out 11 candidate genes (TIMP1, BGN, LTBP1, AKR1C3, LOXL3, MAOA, MAOB, GNA12, IFI30, CNTNAP1, IGF2BP1) according to the node degree of PPI network analysis between 28 genes and known vascular development related genes. We further figured out LOXL3, MAOA, CNTNAP1 and IGF2BP1 are the novel important genes for vascular malformation, of which CNTNAP1 is the joint gene of three-omics. Through expression patterns and gene knockdown in zebrafish embryos, we revealed that LOXL3, MAOA, CNTNAP1 and IGF2BP1 affect vascular development (Fig 2 E-G). Conclusions We identify novel key candidate genes (LOXL3, MAOA, CNTNAP1 and IGF2BP1) potentially related to CHD with vascular malformation and elucidate the possible molecular pathogenesis of CHD. Our multi-omics integrative analysis provides new insights on the multi-layered etiology of CHD, which might be helpful in developing clinical diagnosis and precaution for CHD.Figure 1Figure 2

A spatiotemporal transcriptomic atlas of mouse placentation

The placenta, a temporary but essential organ for gestational support, undergoes intricate morphological and functional transformations throughout gestation. However, the spatiotemporal patterns of gene expression underlying placentation remain poorly understood. Utilizing Stereo-seq, we constructed a Mouse Placentation Spatiotemporal Transcriptomic Atlas (MPSTA) spanning from embryonic day (E) 7.5 to E14.5, which includes the transcriptomes of large trophoblast cells that were not captured in previous single-cell atlases. We defined four distinct strata of the ectoplacental cone, an early heterogeneous trophectoderm structure, and elucidated the spatial trajectory of trophoblast differentiation during early postimplantation stages before E9.5. Focusing on the labyrinth region, the interface of nutrient exchange in the mouse placenta, our spatiotemporal ligand–receptor interaction analysis unveiled pivotal modulators essential for trophoblast development and placental angiogenesis. We also found that paternally expressed genes are exclusively enriched in the placenta rather than in the decidual regions, including a cluster of genes enriched in endothelial cells that may function in placental angiogenesis. At the invasion front, we identified interface-specific transcription factor regulons, such as Atf3, Jun, Junb, Stat6, Mxd1, Maff, Fos, and Irf7, involved in gestational maintenance. Additionally, we revealed that maternal high-fat diet exposure preferentially affects this interface, exacerbating inflammatory responses and disrupting angiogenic homeostasis. Collectively, our findings furnish a comprehensive, spatially resolved atlas that offers valuable insights and benchmarks for future explorations into placental morphogenesis and pathology.

Transcriptomic Analysis of Genes Associated with Stinger Development at Different Life Stages of Apis mellifera.

Stingers, evolved from ovipositors, are an important defense organ for the Apidae, Vespidae, and Formicidae species. However, the molecular mechanism of stinger development remains unclear. Here, we show that the earliest time point for the appearance of stingers in Apis mellifera is at the 1-day-old worker pupal stage based on morphological observations and anatomy from the pre-pupal to adult stages. To discover the genes related to stinger development, we first comprehensively compared the stinger transcriptome at different stages and screened 1282, 186, and 166 highly expressed genes in the stingers of 1- and 5-day-old worker pupae and newly emerged worker bees (NEBs), respectively, then identified 25 DEGs involved in the early stage of stinger development. We found that Dll was a key candidate gene in the early development of A. mellifera stingers by combining analyses of the protein-protein interaction network and spatiotemporal expression patterns. An RNAi experiment showed that about 20% of individuals exhibited tip bending in the piercing parts of their stingers in the Dll-dsRNA-treated group, with the morphology presenting as side-side or front-back tip bending. This indicates that Dll plays a vital role in the early development of A. mellifera stingers. Together, our study provides insight into the molecular mechanism of Hymenoptera stinger development and an inspiration for the molecular breeding of gentle honeybee species with stinger abnormalities.

Analysis of rosmarinic acid synthase (RAS) gene family and functional study of SmRAS1/2/4 in Salvia miltiorrhiza

Rosmarinic acid synthase is an essential enzyme involved in the biosynthesis of rosmarinic acid (RA), which facilitates the coupling of phenylpropanoid and tyrosine-derived pathway products. Our study identified six SmRAS genes in Salvia miltiorrhiza, with SmRAS1 being the only one functionally characterized to date. Real-time quantitative PCR was employed to analyze the expression profiles of the SmRAS gene family, revealing that SmRAS1/2/4 are predominantly expressed in the roots, which are the medicinal components of S. miltiorrhiza. SmRAS2 and SmRAS4 exhibited significant responses to abscisic acid (ABA), gibberellin (GA3), and methyl jasmonate (MeJA) stimuli, while SmRAS1 had notable responses to GA3 and MeJA. β-glucuronidase (GUS) staining in transgenic Arabidopsis thaliana confirmed a spatiotemporal expression pattern of SmRAS1/2/4 that was consistent with the qRT-PCR results. SmRAS1/2/4 are primarily localized to the cytoplasm and plasma membrane. Our findings suggested that the overexpressions of SmRAS1 or SmRAS4 led to increased levels of salvianolic acid B (SalB) and RA, with a concomitant decrease in the Danshensu (DSS) content, which served as a substrate. In contrast, RNA interference lines exhibited a downward trend in the content of these substances. Interestingly, no significant changes were detected in the SalB, RA, or DSS contents due to the overexpression of SmRAS2 or RNA interference lines. Collectively, our study demonstrated that SmRAS1 and SmRAS4 are key regulators of RA and SalB biosynthesis in S. miltiorrhiza, while SmRAS2′s role appears less impactful, suggesting a complex regulatory network that influences the medicinal properties of this plant.

A role of Lhx2 in the migration and axonal projection of cortical postmitotic neurons in the cortical upper layer of the mouse neocortex

The transcription factor LHX2 contains a LIM domain and plays an important role in the development of the vertebrate nervous system. Although much research has been conducted on the function of Lhx2 during cerebral development, its role in postmitotic neuron differentiation in the cerebral cortex remains unknown. Therefore, this study was conducted to determine the function of Lhx2 in dynamic and elaborate developmental processes, including neurogenesis. We first created and confirmed an Lhx2-BAC Gfp transgenic model to three-dimensionally confirm the spatiotemporal expression pattern of Lhx2 during brain development. On this basis, we used the bilateral in utero electroporation technique to express the dominant-negative form of LHX2. LHX2 was confirmed to be important for the migration and callosal projection of postmitotic neurons that form the upper layer of the cerebral cortex during neurogenesis. Additionally, transcriptome analysis confirmed that LHX2 affected the genes involved in neuronal migration and axonal projection. We demonstrated that Lhx2 is important for postmitotic neurons in the cerebral cortex, which migrate to normal positions and extend nerve axons. Taken together, our findings can provide important clues to understanding the relationship between human Lhx2 gene mutations and brain developmental diseases.

Genome-wide investigation of glycoside hydrolase 9 (GH9) gene family unveils implications in orchestrating the mastication trait of Citrus sinensis fruits

Mastication trait of citrus significantly influences the fruit’s overall quality and consumer preference. The accumulation of cellulose in fruits significantly impacts the mastication trait of citrus fruits, and the glycoside hydrolase 9 (GH9) family plays a crucial role in cellulose metabolism. In this study, we successfully identified 32 GH9 genes from the Citrus sinensis genome and subsequently conducted detailed bioinformatics analyses of the GH9 family. Additionally, we profiled the spatiotemporal expression patterns of CsGH9 genes across four distinct fruit tissue types and six crucial developmental stages of citrus fruits, leveraging transcriptome data. Parallel to this, we undertook a comparative analysis of transcriptome profiles and cellulose content among diverse fruit tissues spanning six developmental stages. Furthermore, to identify the pivotal genes involved in cellulose metabolism within the GH9 family during fruit maturity, we employed correlation analysis between cellulose content and gene expression in varying tissues across diverse citrus varieties. This analysis highlighted key genes such as CsGH9A2/6 and CsGH9B12/13/14/22. Collectively, this study provides an in-depth analysis of the GH9 gene family in citrus and offers novel molecular insights into the underlying mechanisms governing the mastication trait formation in citrus fruits.

Reproductive development in Trithuria submersa (Hydatellaceae: Nymphaeales): the involvement of AGAMOUS-like genes

Main conclusionIn the early diverging angiosperm Trithuria submersaTsAG1 and TsAG2 are expressed in different flower organs, including bracts, while TsAG3 is more ovule-specific, probably functioning as a D-type gene.Species of Trithuria, the only genus of the family Hydatellaceae, represent ideal candidates to explore the biology and flower evolution of early diverging angiosperms. The life cycle of T. submersa is generally known, and the “reproductive units” are morphologically well described, but the availability of genetic and developmental data of T. submersa is still scarce. To fill this gap, a transcriptome analysis of the reproductive structures was performed and presented in this work. This analysis provided sequences of MADS-box transcription factors, a gene family known to be involved in flower and fruit development. In situ hybridization experiments on floral buds were performed to describe the spatiotemporal expression patterns of the AGAMOUS genes, revealing the existence of three AG genes with different expression domains in flower organs and in developing ovules. Trithuria may offer important clues to the evolution of reproductive function among early angiosperms and Nymphaeales in particular, and this study aims to broaden relevant knowledge regarding key genes of reproductive development in non-model angiosperms, shaping first flower appearance and evolution.

Involvement of extracellular vesicle microRNA clusters in developing healthy and Rett syndrome brain organoids

Rett syndrome (RTT) is a neurodevelopmental disorder caused by de novo mutations in the MECP2 gene. Although miRNAs in extracellular vesicles (EVs) have been suggested to play an essential role in several neurological conditions, no prior study has utilized brain organoids to profile EV-derived miRNAs during normal and RTT-affected neuronal development. Here we report the spatiotemporal expression pattern of EV-derived miRNAs in region-specific forebrain organoids generated from female hiPSCs with a MeCP2:R255X mutation and the corresponding isogenic control. EV miRNA and protein expression profiles were characterized at day 0, day 13, day 40, and day 75. Several members of the hsa-miR-302/367 cluster were identified as having a time-dependent expression profile with RTT-specific alterations at the latest developmental stage. Moreover, the miRNA species of the chromosome 14 miRNA cluster (C14MC) exhibited strong upregulation in RTT forebrain organoids irrespective of their spatiotemporal location. Together, our results suggest essential roles of the C14MC and hsa-miR-302/367 clusters in EVs during normal and RTT-associated neurodevelopment, displaying promising prospects as biomarkers for monitoring RTT progression.Graphical

Systematic Identification and Characterization of O-Methyltransferase Gene Family Members Involved in Flavonoid Biosynthesis in Chrysanthemum indicum L.

Chrysanthemum indicum L. capitulum is an enriched source of flavonoids with broad-ranging biological activities, mainly due to their anti-inflammatory, anti-cancer, immune regulation, anti-microbial activity, hepatoprotective, and neuroprotective effects. The O-methylation of various secondary metabolites has previously been demonstrated to be mainly catalyzed by S-adenosyl-L-methionine-dependent O-methyltransferase (OMT) proteins encoded by the OMT gene family. However, limited comprehensive study was published on the OMT gene family, especially the CCoAOMT subfamily, involved in the O-methylation of flavonoids in Chrysanthemum. Here, we analyzed the spatiotemporal expression patterns of C. indicum OMT genes in leaf and flower at different developmental stages. Transcriptome sequencing and qRT-PCR analysis showed that COMTs were mainly highly expressed in capitulum, especially in full bloom, while CCoAOMTs were mainly highly expressed in leaves. Correlation analysis of OMT gene expression and flavonoids accumulation revealed that four OMTs (CHR00029120, CHR00029783, CHR00077404, and CHR00078333) were putatively involved in most methylated flavonoids biosynthesis in the capitulum. Furthermore, we identified a true CCoAOMT enzyme, CiCCoAOMT1, and found that it catalyzed O-methylation of quercetin and luteolin at the 3'-OH position. In summary, this work provides an important theoretical basis for further research on the biological functions of OMTs in C. indicum.

Spatiotemporal Mapping of the Oxytocin Receptor at Single-Cell Resolution in the Postnatally Developing Mouse Brain.

The oxytocin receptor (OXTR) has garnered increasing attention for its role in regulating both mature behaviors and brain development. It has been established that OXTR mediates a range of effects that are region-specific or period-specific. However, the current studies of OXTR expression patterns in mice only provide limited help due to limitations in resolution. Therefore, our objective was to generate a comprehensive, high-resolution spatiotemporal expression map of Oxtr mRNA across the entire developing mouse brain. We applied RNAscope in situ hybridization to investigate the spatiotemporal expression pattern of Oxtr in the brains of male mice at six distinct postnatal developmental stages (P7, P14, P21, P28, P42, P56). We provide detailed descriptions of Oxtr expression patterns in key brain regions, including the cortex, basal forebrain, hippocampus, and amygdaloid complex, with a focus on the precise localization of Oxtr+ cells and the variance of expression between different neurons. Furthermore, we identified some neuronal populations with high Oxtr expression levels that have been little studied, including glutamatergic neurons in the ventral dentate gyrus, Vgat+Oxtr+ cells in the basal forebrain, and GABAergic neurons in layers 4/5 of the cortex. Our study provides a novel perspective for understanding the distribution of Oxtr and encourages further investigations into its functions.

Identification and functional analysis of a new cold induced LncRNA44154

With the development of high-throughput sequencing technology, numerous unknown long noncoding RNAs (lncRNAs) have been discovered. Preliminary research found that after being subjected to low-temperature enviroment, lncRNA44154 expression in the subcutaneous fat of pigs significantly increased. In order to analyze its biological function and regulatory mechanism, qRT-PCR was used to detect its spatiotemporal expression pattern. The nucleoplasmic isolation and FISH were used to locate it subcellular. Through overexpression or interference assays discussed the effect of lncRNA44154 on adipocyte proliferation and differentiation. RNA-seq was used to screen the biological pathways that lncRNA44154 may be involved in. RNA-pull down was used to find the interaction protein and double luciferase reporter gene detection experiment was used to validate the target gene. It was found that the expression of lncRNA44154 had tissue specificity and played a role in adipocyte proliferation. It was expressed in both cytoplasm and nucleus. After overexpression of lncRNA44154 in adipocytes, the expression level of Cyclins B, D, and E were all up-regulated and the cell proliferation ability was significantly enhanced. The number of positive cells increased significantly and the DNA replication activity was significantly enhanced. LncRNA44154 can promote the proliferation of porcine preadipocytes by encouraging cells to enter the S phase of the cell cycle. The result of interfering with lncRNA44154 is the opposite. Further analysis revealed that lncRNA44154 regulates intracellular lipid metabolism, inhibits the production of intracellular lipid droplets, and inhibits preadipocyte differentiation. RNA-seq results indicate that lncRNA44154 may be involved in biological pathways such as AMPK, PPAR and cAMP signaling pathway. It may work by regulating the RPS4X gene. This research may provide a new perspective for investigating the regulation of adipose metabolism by lncRNAs.

Functional allocation of Mitogen-activated protein kinases (MAPKs) unveils thermotolerance in scallop Argopecten irradians irradians

Global warming has significantly impacted agriculture, particularly in animal husbandry and aquaculture industry. Rising ocean temperatures due to global warming are severely affecting shellfish production, necessitating an understanding of how shellfish cope with thermal stress. The mitogen-activated protein kinases (MAPK) signaling pathway plays a crucial role in cell growth, differentiation, adaptation to environmental stress, inflammatory response, and managing high temperature stress. To investigate the function of MAPKs in bay scallops, a comparative genomics and bioinformatics approach identified three MAPK genes: AiERK, Aip38, and AiJNK. Structural and phylogenetic analyses of these proteins were conducted to determine their evolutionary relationships. Spatiotemporal expression patterns were examined at different developmental stages and in various tissues of healthy adult scallops. Additionally, the expression regulation of these genes was studied in selected tissues (hemocyte, gill, heart, mantle) following exposure to high temperatures (32 °C) for different durations (0 h, 6 h, 12 h, 24 h, 3 d, 6 d, 10 d). The spatiotemporal expressions of AiMAPKs were ubiquitous, with significant increases in AiERK expression observed at the umbo larval stage (3.09-fold), while Aip38 and AiJNK were identified as potential maternal effect genes. In adult scallops, different gene expression patterns of AiMAPKs were observed across eight tissues, with high expressions in the foot and gill, and lower expressions in the striated muscle. Following high temperature stress, AiMAPKs expressions in the gill and mantle were mainly up-regulated, while in the hemocyte, they were primarily down-regulated. These findings indicate time- and tissue-dependent expression patterns with functional allocation in response to different thermal durations. This study enhances our understanding of the function and evolution of AiMAPKs genes in shellfish and provides a theoretical basis for elucidating the energy regulation mechanism of bay scallops in response to high temperature stress.

Evolution and Spatiotemporal Expression of ankha and ankhb in Zebrafish.

Craniometaphyseal Dysplasia (CMD) is a rare skeletal disorder that can result from mutations in the ANKH gene. This gene encodes progressive anksylosis (ANK), which is responsible for transporting inorganic pyrophosphate (PPi) and ATP from the intracellular to the extracellular environment, where PPi inhibits bone mineralization. When ANK is dysfunctional, as in patients with CMD, the passage of PPi to the extracellular environment is reduced, leading to excess mineralization, particularly in bones of the skull. Zebrafish may serve as a promising model to study the mechanistic basis of CMD. Here, we provide a detailed analysis of the zebrafish Ankh paralogs, Ankha and Ankhb, in terms of their phylogenic relationship with ANK in other vertebrates as well as their spatiotemporal expression patterns during zebrafish development. We found that a closer evolutionary relationship exists between the zebrafish Ankhb protein and its human and other vertebrate counterparts, and stronger promoter activity was predicted for ankhb compared to ankha. Furthermore, we noted distinct temporal expression patterns, with ankha more prominently expressed in early development stages, and both paralogs also being expressed at larval growth stages. Whole-mount in situ hybridization was used to compare the spatial expression patterns of each paralog during bone development, and both showed strong expression in the craniofacial region as well as the notochord and somites. Given the substantial overlap in spatiotemporal expression but only subtle patterning differences, the exact roles of these genes remain speculative. In silico analyses predicted that Ankha and Ankhb have the same function in transporting PPi across the membrane. Nevertheless, this study lays the groundwork for functional analyses of each ankh paralog and highlights the potential of using zebrafish to find possible targeted therapies for CMD.

Genetic variation and molecular profiling of congenital malformations of the female genital tract based on whole-genome sequencing.

Congenital malformations of the female genital tract (CM-FGT) are characterized by abnormal development of the fallopian tubes, uterus, and vagina, often accompanied by malformations in the urinary system, bones and hearing. However, no definitive pathogenic genes and molecular genetic causes have been identified. We present the largest whole-genome sequencing study of CM-FGT to date, analyzing 590 individuals in China: 95 patients, 442 case-controls, and 53 familial controls. Among the patients, 5.3% carried known CM-FGT-related variants. Pedigree and case-control analyses in two dimensions of coding and non-coding regulatory regions revealed seven novel de novo copy number variations, 12 rare single-nucleotide variations, and 10 rare 3' untranslated region (UTR) mutations in genes related to CM-FGT, particularly highlighting ASH1L as a pathogenic gene. Single-cell sequencing data showed that the majority of CM-FGT-related risk genes are spatiotemporally specifically expressed early in uterus development. In conclusion, this study identified novel variants related to CM-FGT, particularly highlighting ASH1L as a pathogenic gene. The findings provide insights into the genetic variants underlying CM-FGT, with single-cell sequencing data revealing spatiotemporal specific expression patterns of key risk genes early in uterine development. This study significantly advances the understanding of CM-FGT etiology and genetic landscape, offering new opportunities for prenatal screening.

AMPK-mediated regulation of cardiac energy metabolism: Implications for thermotolerance in Argopecten irradians irradians

AMPK is a key regulator of metabolism in eukaryotes across various pathways related to energy regulation. Although extensive investigations of AMPK have been conducted in mammals and some model organisms, research on AMPK in scallops is comparatively limited. In this study, three AMPK family genes (AiAMPKα, AiAMPKβ and AiAMPKγ) in scallop Argopecten irradians irradians were identified through genome scanning. Structure prediction and phylogenetic analyses of AiAMPKs were performed to determine their structural features and evolutionary relationships. Spatiotemporal expression patterns of AiAMPKs at different developmental stages and in healthy adult tissues were analyzed to elucidate the function of AiAMPKs in bay scallops’ growth and development. The spatiotemporally specific expression of AiAMPKs implied their important roles in growth and development of bay scallops. Heat stress experiment was performed to determine the regulations of AiAMPKs in four kinds of thermosensitive tissues. Expression profiles revealed distinct molecular mechanisms of AiAMPKs in different tissues in response to heat stress: significant down-regulations in mobile hemocytes, but dominant up-regulations occurring in stationary gills, mantles and hearts. Functional verification including knock-down of AiAMPKα and inhibition of AiAMPK was separately conducted in the thermotolerant tissue heart at the post-transcription and translation levels. The thermotolerant index Arrhenius break temperature (ABT) showed a significant decrease and the rate-amplitude product (RAP) peaked earlier in the individuals after RNAi targeting AiAMPKα, displaying an earlier transition to anaerobic metabolism under heat stress, indicating an impairing ability of aerobic metabolism. After AiAMPK inhibition, widespread down-regulations of genes in key energy metabolism pathways, RNA polymerase II-mediated transcription, and aminoacyl-tRNA synthesis pathways were obviously observed, revealing the post-translational inhibition of AiAMPK hindered cardiac energy metabolism, basal transcription and translation. Overall, our findings provide evidences for exploring the molecular mechanisms of energy regulation in thermotolerant traits in bay scallops under ongoing global warming.

Physiological roles of Arabidopsis MCA1 and MCA2 based on their dynamic expression patterns

Determining the mechanisms by which plants sense and respond to mechanical stimuli is crucial for unraveling the detailed processes by which plants grow and develop. Mechanosensitive (MS) channels, including MCA1 and its paralog MCA2 in Arabidopsis thaliana, may be essential for these processes. Although significant progress has been made in elucidating the physiological roles of MS channels, comprehensive insights into their expression dynamics remain elusive. Here, we summarize recent advancements and new data on the spatiotemporal expression patterns of the MCA1 and MCA2 genes, revealing their involvement in various developmental processes. Then, we describe findings from our study, in which the expression profiles of MCA1 and MCA2 were characterized in different plant organs at various developmental stages through histochemical analyses and semiquantitative RT‒PCR. Our findings revealed that MCA1 and MCA2 are preferentially expressed in young tissues, suggesting their pivotal roles in processes such as cell division, expansion, and mechanosensing. Lastly, we discuss the differential expression patterns observed in reproductive organs and trichomes, hinting at their specialized functions in response to mechanical cues. Overall, this review provides valuable insights into the dynamic expression patterns of MCA1 and MCA2, paving the way for future research on the precise roles of these genes in planta.

Transcriptome-level analysis of gene expressions in different tissues of tomato and key gene identifications during seed germination

Exploring the spatiotemporal expression patterns of genes is the foundation for studying their functions and molecular mechanisms. Genes with stable spatiotemporal expression patterns are often used as reference genes for quantitative real-time polymerase chain reaction (qRT-PCR) and genes are significantly highly expressed in a specific tissue or stage often represent their unique biological functions. Tomato is an important economic crop and a model plant for scientific researches. However, the existing reference genes in tomatoes are not stable enough and researches on tissue-specific genes have been more focused on flowers and fruits, with less emphasis on studying the seed germination process. In this study, a total of 26 tissue/organ samples from the entire growth cycle of tomatoes were collected from Micro-Tom (Solanum lycopersicum cv. Micro-Tom). After RNA sequencing, a transcriptome database of the entire growth cycle of tomatoes was constructed. Solyc01g107870.3 gene was identified as a novel reference gene through systematic analysis and validated by qRT-PCR. Moreover, tissue-specific genes in seed and 8 days past germination (DPG) seedlings are screened. Among them, SlLEA16 gene belongs to late embryogenesis abundant protein (LEA) gene family, having subcellular localization in the nucleus and cytoplasm, and playing important roles in seed germination initiation. The above experimental results provide data support for studying tomato gene spatiotemporal expression patterns and can also serve as an experimental basis for investigating the deeper mechanisms of tomato seed germination processes.

Spatiotemporally distinct roles of cyclooxygenase-1 and cyclooxygenase-2 at fetomaternal interface in mice.

Embryo implantation is crucial for ensuring a successful pregnancy outcome and subsequent child health. The intrauterine environment during the peri-implantation period shows drastic changes in gene expression and cellular metabolism in response to hormonal stimuli and reciprocal communication with embryos. Here, we performed spatial transcriptomic analysis to elucidate the mechanisms underlying embryo implantation. Transcriptome data revealed that lipid metabolism pathways, especially arachidonic acid-related (AA-related) ones, were enriched in the embryo-receptive luminal epithelia. Cyclooxygenases (COXs), rate-limiting enzymes involved in prostaglandin production by AA, were spatiotemporally regulated in the vicinity of embryos during implantation, but the role of each COX isozyme in the uterus for successful pregnancy was unclear. We established uterine-specific COX2-knockout (uKO) and COX1/uterine COX2-double-KO (COX1/COX2-DKO) mice. COX2 uKO caused deferred implantation with failed trophoblast invasion, resulting in subfertility with reduced pregnancy rates and litter sizes. COX1/COX2 DKO induced complete infertility, owing to abrogated embryo attachment. These results demonstrate that both isozymes have distinct roles during embryo implantation. Spatial transcriptome and lipidome analyses revealed unique profiles of prostaglandin synthesis by each COX isozyme and spatiotemporal expression patterns of downstream receptors throughout the endometrium. Our findings reveal previously unappreciated roles of COXs at the fetomaternal interface to establish early pregnancy.

An approach to analyze spatiotemporal patterns of gene expression at single-cell resolution in Candida albicans-infected mouse tongues.

Candida albicans is a fungal pathobiont inhabiting multiple mucosal surfaces of the human body. Immunosuppression, antibiotic-induced microbial dysbiosis, or implanted medical devices can impair mucosal integrity enabling C. albicans to overgrow and disseminate, causing either mucosal diseases such as oropharyngeal candidiasis or life-threatening systemic infections. Profiling fungal genes that are expressed in the infected mucosa or in any other infected organ is paramount to understand pathogenesis. Ideally, these transcript profiling measurements should reveal the expression of any gene at the single-cell level. The resolution typically achieved with current approaches, however, limits most gene expression measurements to cell population averages. The approach described in this report provides a means to dissect fungal gene expression in infected tissues at single-cell resolution.

JrPPO1/2 play distinct roles in regulating walnut fruit browning by different spatiotemporal expression and enzymatic characteristics

Polyphenol oxidase (PPO) activity drives walnut fruit browning, but the roles of its only two-family genes, JrPPO1 and JrPPO2, remain unclear. This study explores the spatiotemporal expression and enzymatic characteristics of JrPPO1 and JrPPO2 in walnut. Treatment with the PPO activator CuSO4 and H2O2 accelerated fruit browning and up-regulated JrPPO1/2 expression, whereas treatment with the PPO inhibitor ascorbic acid delayed browning, down-regulating JrPPO1 and up-regulating JrPPO2 expression. Compared to mJrPPO1, mJrPPO2 can exhibited better enzyme activity at higher temperatures (47 °C) and in more acidic environments (pH 4.25). mJrPPO2 exhibited a higher substrate specificity over mJrPPO1, and the preferred substrates are catechol, chlorogenic acid, and epicatechin. Additionally, mJrPPO2 adapted better to low concentration of oxygen (as low as 1.0% O2) and slightly elevated CO2 levels compared to mJrPPO1. Subcellular localization and spatiotemporal expression patterns showed that JrPPO1 is only expressed in green tissues and located in chloroplasts, while JrPPO2 is also located in chloroplasts, partly associated with membranes, and is expressed in both green and non-green tissues. Silencing JrPPO1/2 with virus-induced gene silencing (VIGS) reduced fruit browning, maintained higher total phenols, and decreased MDA production. Notably, silencing JrPPO1 had a greater impact on browning than JrPPO2, indicating JrPPO1's greater contribution to PPO activity and fruit browning in walnut fruits. Consequently, JrPPO1 can be effectively regulated both at the molecular level and by manipulating environmental conditions, to achieve the objective of controlling fruit browning.